Apha 13302

DR MATHIAS WERNBOM (Orcid ID : 0000-0003-0411-3611)
Accepted Article
Article type : Review Article
Muscle fibre activation and fatigue with low-load blood

flow restricted resistance exercise – An integrative
physiology review
Mathias Wernbom1,2, Per Aagaard3
1
Center for Health and Performance, Department of Food and Nutrition and Sport Science,
University of Gothenburg, Gothenburg, Sweden. 2Department of Health and Rehabilitation,
Institute of Neuroscience and Physiology, Sahlgrenska Academy, University of Gothenburg,
Gothenburg, Sweden. 3Department of Sports Sciences and Clinical Biomechanics, SDU
Muscle Research Cluster (SMRC), University of Southern Denmark, Odense M, Denmark.
Short title: Muscle fibre use in ischemic exercise
Corresponding author: Mathias Wernbom, Center for Health and Performance, Department
of Food and Nutrition and Sport Science, University of Gothenburg, Sweden. Box 300, SE-
405 30 Göteborg, Sweden. E-mail: mathias.wernbom@gu.se
Abstract
Blood flow restricted resistance exercise (BFRRE) has been shown to induce increases in
muscle size and strength, and continues to generate interest from both clinical and basic
research points of view. The low loads employed, typically 20-50% of the one repetition
maximum (1RM), make BFRRE an attractive training modality for individuals who may not
tolerate high musculoskeletal forces (e.g. selected clinical patient groups such as frail old
adults and patients recovering from sports injury) and/or for highly trained athletes who have
reached a plateau in muscle mass and strength.
This article has been accepted for publication and undergone full peer review but has not
been through the copyediting, typesetting, pagination and proofreading process, which may
lead to differences between this version and the Version of Record. Please cite this article as
doi: 10.1111/apha.13302
This article is protected by copyright. All rights reserved.
It has been proposed that achieving a high degree of muscle fibre recruitment is
important for inducing muscle hypertrophy with BFRRE, and the available evidence suggest
Accepted Article
that fatiguing low-load exercise during ischemic conditions can recruit both slow (type I) and
fast (type II) muscle fibres. Nevertheless, closer scrutiny reveals that type II fibre activation
in BFRRE has to date largely been inferred using indirect methods such as electromyography
(EMG) and magnetic resonance spectroscopy (MRS), while only rarely addressed using more
direct methods such as measurements of glycogen stores and phosphocreatine (PCr) levels in
muscle fibres. Hence, considerable uncertainity exists about the specific pattern of muscle
fiber activation during BFRRE.
Therefore, the purpose of this narrative review was (1) to summarize the
evidence on muscle fibre recruitment during BFRRE as revealed by various methods
employed for determining muscle fibre usage during exercise, and (2) to discuss reported
findings in light of the specific advantages and limitations associated with these methods.
Keywords: Occlusion, ischemia, muscle hypertrophy, motor units, fatigue
1. Introduction, background and purpose
2. Definitions and limitations
3. Neural control of muscle force development

3.1. Muscle fibre types and the size principle of motor unit recruitment
3.2. Can deviations from the size principle occur in voluntary movements?
3.3. Rate coding of motor units in voluntary movements
3.4. Can high threshold motor units fire at tetanic frequencies during maximal
voluntary contractions?
3.5. Type of activation versus firing rates for high threshold motor units
3.6. The possible roles of spike-frequency adaptation and small muscle afferents in
limiting maximal firing rates
3.7. The role of small muscle afferents in limiting muscle activation during fatiguing
exercise with large muscle mass

4. Fatigue and recovery in the muscle fibre types with ischemic exercise
4.1. Fatigue in the muscle fibre types during ischemic and anaerobic exercise
4.2. Possible contributions of low-frequency fatigue and neuromuscular
transmission
Accepted Article
failure to torque decreases in ischemic exercise
4.3. Force recovery in the muscle fibre types after ischemic and anaerobic exercise
5. Methods to investigate muscle fibre activation and use

5.1. Glycogen depletion
5.2. Phosphocreatine depletion
5.3. Electromyography (EMG)
5.3.1. Surface EMG
5.3.2. Intramuscular EMG
31
5.4. P-MRS
5.5. Studies on combined EMG and 31P-MRS
6. Studies on muscle fibre activation and use in BFRRE

6.1. Studies on glycogen depletion in BFRRE
6.2. Studies on phosphocreatine depletion to assess muscle fibre use in BFRRE
6.3. Summary of studies on glycogen and phosphocreatine depletion in BFRRE
6.4. EMG studies on BFRRE
6.4.1. Surface EMG
31
6.5. P-MRS studies on muscle fibre recruitment in BFRRE
6.6. Can the order of muscle fibre recruitment be changed during cuff occlusion?
7. Conclusions and implications

7.1. Order of muscle fibre recruitment in BFRRE
7.2. Patterns of muscle fibre use and fatigue in BFRRE
7.3. Consequences for hypertrophic adaptations in BFRRE
7.4. Consequences for neural adaptations to BFRRE
7.5. Consequences for interpretations of surface EMG during BFRRE
7.6. Consequences for the possible role of ”metabolic stress” in BFRRE
8. References
1. Introduction, background and purpose
Resistance exercise with low-to-moderate loads combined with blood flow restriction (BFR)
has been shown to induce increases in muscle size comparable to those seen with
conventional heavy resistance exercise in untrained individuals1-3, although a recent meta-
analysis4 suggests that the muscle strength gains are somewhat lower with BFR resistance
exercise (BFRRE). Nevertheless, the marked neuromuscular adaptations observed with

BFRRE continue to generate considerable interest from both clinical and basic research
points of view since the loads used in BFRRE typically are only ~20-50% of the 1 repetition
Accepted Article
maximum (1RM), to compare with the ~70-85% of 1RM that is often recommended and used
in conventional strength training (reviewed by Wernbom et al5 and Hughes et al6).
The stimuli for the muscle hypertrophy resulting from BFRRE, also known as
occlusion training or ischemic strength training, are not well understood at present. However,
it has been proposed that achieving a high degree of muscle fibre recruitment in BFRRE is
important1,5,7,8, and a number of studies have reported evidence suggesting that fatiguing low-
load exercise during ischemic conditions can recruit both slow (type I) and fast (type II)
muscle fibres (reviewed by Wernbom et al5, see also Suga et al9 and Takada et al10). The
physiological significance of type II fibre recruitment is that these fibres typically increase
their cross-sectional area to a greater extent than type I fibres with conventional heavy
strength training11-14, and that when measured at 37°C, human type II fibres have 3-fold
greater maximal contraction velocities and 4-fold higher maximal power outputs than type I
fibres15. Against this background, the considerable hypertrophy in both type I and type II
fibres reported by Nielsen et al16 after a short period of low-load high frequency BFRRE is of
great interest.
Taken together, it would appear that there is good evidence for the recruitment
of both fibre types with BFRRE. Nevertheless, closer scrutiny reveals that type II fibre
activation in BFRRE has to date largely been inferred via indirect methods such as
electromyography (EMG) (e.g., Moritani et al17; Takarada et al1,18; and Wernbom et al19) and
magnetic resonance spectroscopy (MRS) (e.g., Suga et al9 and Takada et al10), and only rarely
with more direct methods such as the measurement of glycogen and phosphocreatine (PCr)
levels in muscle fibres (e.g. Krustrup et al20; Cumming et al21). Moreover, in contrast to

Nielsen et al16, other studies have shown fibre hypertrophy and myonuclear addition
primarily in the type I myofibres with low-load BFRRE22-24.

Accepted Article
Therefore, the purpose of this review is to summarize the evidence on muscle
fibre recruitment in BFRRE as revealed by the various methods that have been employed for
determining muscle fibre use during exercise, and discuss reported findings in the light of
some of the advantages and limitations of these methods.
However, because motor unit recruitment, firing rates, substrate use and fatigue
all affect the physiological stimuli which the muscle fibres are exposed to, it was deemed
necessary to provide an introductory background of the physiology of these processes
(Sections 3 and 4) as well as a discussion of different methods to investigate motor unit
recruitment and muscle fibre use (Section 5). In section 6, the evidence of motor unit
recruitment and muscle fibre use in BFRRE will be outlined in detail. Finally, Section 7
discusses the possible influences of motor unit recruitment and firing rates on the resulting
neuromuscular adaptations to systematic training periods of BFRRE.
2. Definitions and limitations
For the purposes of this review, BFRRE is defined as exercise that is performed with typical
resistance exercise movements (e.g., knee extensions, leg presses, squats, elbow flexions)
with concurrent addition of BFR, most often provided by a tourniquet cuff around the
exercising limb although elastic wraps can also be used. In the current text low-to-moderate
load BFRRE will refer to a load range of ~20-50% of 1RM, unless otherwise indicated. In
some studies, the experimental resistance training protocols consisted of isometric muscle
contractions instead of dynamic resistance exercise. These exercise protocols were also
considered for this review. With isometric contractions, the intensity is typically expressed as

a percentage of the maximal voluntary isometric contraction (MVC) torque in the exercise,
and the load range referred to here is 20-50% of MVC.

Accepted Article
Because the most effective mode of blood-flow restricted training for increases
in strength and muscle size appears to be of the resistance exercise type (reviewed by
Wernbom et al5), the scope of this review is limited to BFRRE. With regard to the degree of
BFR, both very low levels of vascular restriction (e.g. Sumide et al25) and near-complete
occlusion (e.g. Shinohara et al26; Takada et al10) have been successfully employed to induce
measurable gains in muscle strength and mass. Therefore, studies covering this wide
physiological range are included and discussed in this review.
3. Neural control of muscle force development
The body of knowledge reviewed in this section is based on a vast number of studies that
have made motor unit recordings with needle or fine-wire electrodes, and additionally on
studies which have employed depletion of glycogen and PCr to investigate muscle fibre use.
Recent studies using sophisticated decomposition of surface electromyographic signals have
also been included as sources of information. See Section 5 for discussion on the some of the
advantages and disadvantages with these methods.
3.1. Muscle fibre types and the size principle of motor unit recruitment
It is generally accepted that with voluntary efforts, motor units are usually recruited in an
orderly manner based on the size of the motor neurons, so that small motor units comprised
of type I muscle fibres are recruited first, and with greater demands of force and/or velocity,
larger motor units with type II muscle fibres are increasingly recruited27-30. According to their
fatigue and contractile characteristics, motor units are usually classified into three major

types: slow fatigue-resistant (S) units, fast fatigue-resistant (FR) units, and fast fatigable (FF)
units31-33.
Accepted Article
However, this classification was originally based on studies on cat
gastrocnemius muscles, and attempts to reproduce this scheme in humans have resulted in a
more continuous distribution of contractile properties, without clustering into discrete groups
(reviewed by Heckman & Enoka30). It deserves to be noted, however, that in the paper where
Burke et al32 corroborated their proposed classification system, they also remarked on the
wide and continous range of twitch contraction times found within each class of units, noting
that while the distribution of fatigue resistance of fast units had a bimodal pattern (i.e., FR
and FF), it was essentially also continuous. They also found a few units that were
intermediate between FR and FF and which could not be classified as either type. This later
led to the identification of a fourth type of unit termed F(int), which is intermediate between
FR and FF, and which is sometimes also included in the proposed scheme of motor units34-36.
Furthermore, it is worth mentioning that the human studies which failed to
identify discrete groups of motor units (see Heckman & Enoka30) typically were performed
on small muscles in the hand and the foot. Burke36 suggested that in large limb muscles,
”distinct” types of motor units may be found while in smaller, distal muscles their
distributions are more continuous. In line with this notion, Garnett et al37 studied the human
medial gastrocnemius muscle using intramuscular electrical microstimulation and reported
that the motor units identified could be divided into S, FR and FF units. Nevertheless, the
distribution of contraction times of muscle fibre bundles varies even among relatively large
limb muscles in humans, with some muscles displaying a wide spectrum of contraction times
while others have less wide distributions38. The data reported by Buchtal & Schmalbruch38
suggest that the medial gastrocnemius muscle may indeed be one of the muscles which has

discernible gaps between the clusters of shorter, moderate and longer contraction times,
whereas many other muscles display a more continuous distribution.

Accepted Article
The motor units examined in intact human skeletal muscle correspond to the
three major fibre types that they typically consist of: type I, type IIA and type IIX; and in
addition the intermediate type IIAX which is a hybrid form containing both type IIA and IIX
myosin39-41. Thus, in a typical muscle, the type I fibres are activated first, followed by type
IIA and type IIAX and finally type IIX fibres42,43. With ATPase and immunohistochemical
methods, further hybrid types with varying extents of co-expression of type I and type IIA
myosin can be identified: type IC, type IIC and sometimes also type IIAC41,44. However,
these are normally rarely (~0.5-2%) seen in muscle biopsy samples even when all two or
three subtypes are considered together (see Staron41).
It should be noted that 100% pure type IIX fibres are also often rare, especially
in some types of athletes. As an extreme example, Andersen et al45, using electrophoretic
single fibre analysis, found only 0.1% pure type IIX fibres in the vastus lateralis muscles of
sprinters, in contrast to the 10.5-18.8% type IIX fibres they observed with traditional myosin
ATPase histochemistry methods. With gel electrophoresis analysis Andersen & Aagaard46
reported myosin heavy chain (MHC) IIX isoform content to be 9.3 ± 5.9% (±SD) in
untrained yet habitually physically active males. Using similar methods, Klitgaard et al47
found only 1% pure type IIX fibres in the biceps brachii of bodybuilders, as compared to
12% in control subjects, and that ATPase staining overestimated the percentage type IIX in
both groups. Ennion et al39 reported only 3% pure type IIX in the vastus lateralis of normal
healthy volunteers. It is thus clear that traditional staining methods result in an overestimation
of type IIX fibres, as most of the type IIX fibres identified by ATPase and
immunohistochemistry methods also express varying amounts of type IIA myosin41,45,47. In
recognition of this continuum, type IIAX fibres have sometimes been further subdivided into

IIAx, IIAX and IIXa depending on the balance between the type IIA and IIX
expression44.48,49.
Accepted Article
Interestingly, there appears to be a functional differentiation even among type I
myofibres, as indicated by the recent discovery of two metabolically distinct populations,
termed type I-1 and type I-2 fibres50. The former subtype has a smaller cross-sectional area
(CSA), a higher density of lipid droplets, and a tendency towards lower glycogen content
compared to type I-2 fibres. The subtype I-2 has similar lipid content and CSA to that of type
IIA fibres50. The implications of these two groups of type I fibres are not entirely clear, but
given the rather wide range of glycolytic capactity within the type I fibre population as
judged by the activities of key glycolytic enzymes51 and in vivo glycolytic rates52,53, we
speculate that the type I-2 fibres have a significantly greater glycolytic capacity than the type
I-1 fibres, perhaps somewhere in between that of type-I-1 fibres and type IIA fibres,
respectively. This would be in line with their intermediate position between these two fibre
types in the fibre type continuum as proposed by Prats et al50.
The size principle of motor unit recruitment was first demonstrated by
Henneman and colleagues in the 1950s and 1960s54,55, and essentially verified also in humans
in studies using motor unit recordings obtained with intramuscular electrodes56-61. Notably,
the size principle has also been found to operate in humans during both stretch reflex induced
contractions and with transcranial stimulation of the motor cortex62,63.
It has been generally held that motor unit recruitment in relatively large limb
muscles occurs up to ~85% of MVC, while in hand muscles recruitment is complete already
at ~50-60% of MVC27,28,64. However, accumulating evidence suggests recruitment of new
units up to as high as ~88-96 % of MVC in limb muscles such as tibialis anterior, soleus,
biceps brachii and vastus lateralis65-69. Nevertheless, Christie et al67, Oya et al68 and De Luca
& Hostage69 all employed slow ramp maximal contractions, which may well entail some

fatigue already during the contraction (see Section 3.5.), which in turn can raise the
recruitment threshold of the highest threshold units (discussed in Section 3.4.). As a result,
Accepted Article
the approximate upper limit for motor unit recruitment during more normal and
physiologically relevant contraction durations (e.g. 1-3 seconds) remains to be fully defined.
3.2. Can deviations from the size principle occur in voluntary movements?
Possible exceptions to the size principle have been suggested, mainly falling into the
categories of very rapid (ballistic) contractions, lengthening (eccentric) contractions and
muscle actions during changed afferent input such as cutaneous stimulation (reviewed by
Bawa et al70). Although some studies have reported deviations from the size principle during
ballistic and eccentric contractions71,72, subsequent studies by other researchers have for the
most part failed to confirm these findings (reviewed by Chalmers29, Heckman & Enoka30, and
Bawa et al70, but see also Grimby73 for some possible reasons behind different results).
Current evidence thus supports that the size principle operates in most types of human
voluntary movements, including ballistic and eccentric contractions.
However, an unequal balance of excitatory and inhibitory inputs onto motor
neurons may give rise to departures from the orderly recruitment74. Some of the more
dramatic changes in recruitment order in both animals and humans have been observed with
electrical stimulation of cutaneous afferents, resulting in elevated recruitment thresholds in
the lower-threshold units and vice versa for higher-threshold units61,75,76. Masakado et al76
observed that nearly all the units that normally were recruited above 30% of MVC had
lowered thresholds after cutaneous stimulation. It is important to note that the findings in
these studies do not fulfill criteria for a systematic overall reversal of the recruitment order77.
For example, cases of increased and unchanged thresholds among the high-threshold units

can also be found in the data of Masakado et al76, where the typical decrease among the high-
threshold (>30% of MVC) units seems to have been on the order of 5-10%.
Accepted Article
In other words, while some high-threshold units may become recruited earlier
than normal, a significant number of low-threshold units still appear to be recruited before
these high-threshold units. Furthermore, electrical stimulation of cutaneous afferents might be
considered a non-physiological condition compared with normal tactile stimulation78,79. Still,
the findings in the studies of Kanda et al75, Garnett & Stephens61 and Masakado et al76
constitute possible proof-of-principle that some deviations from the normal recruitment order
can occur when inputs from afferents are unequally distributed on the alpha-motorneuron
pool during voluntary contractions. Interestingly, increasing evidence suggests that such
alterations can also occur during painful contractions80,81. Even so, the physiological
relevance of these findings remains to be determined, as most of the human studies cited
above concerned contractions performed at only a few percent of the MVC, the study of
Masakado et al76 being a notable exception.
3.3. Rate coding of motor units in voluntary movements
The other major mechanism that the central nervous system (CNS) uses to regulate muscle
force development is “rate coding”, which refers to the modulation of the firing rates
(synonymous terms: discharge rates, firing frequencies) of the active motor units27,28. In most
muscles, force typically is increased by both motor unit recruitment and increased firing rates
up to ~85% of MVC, but above this level further increases in muscle force are considered to
be accomplished entirely by rate coding (reviewed by Duchateau et al28). A generalized
scheme of the size principle for the recruitment of motor units and the accompanying
increases in firing rates is depicted in Figure 1 below. Rate coding has a direct influence on
the force developed by the muscle fibres innervated by the firing motor neuron. In the human

quadriceps, the torque evoked by neuromuscular electrical stimulation (NMES) rises with
increasing stimulation frequency and reaches a plateau of maximum torque at ~50-60 Hz

Accepted Article
when the interval between pulses is constant82-84. Notably, studies on bundles of human
muscle fibres which were stimulated in vitro at 37°C demonstrated a similar force plateau at
~50-60 Hz85,86.
Interestingly, even at 100% of MVC, the average maximum firing rates
observed in human limb muscles during maximal contractions in vivo are typically only ~25-
30 Hz64,87, which is considerably lower than the 50-60 Hz required for maximum torque
development with NMES. As noted by Enoka & Fuglevand64, this suggests that either the
force is not maximal during these presumed maximal contractions or that the force exerted by
a motor unit does not depend solely on its average discharge rate. There may be several
explanations for the discrepancy between the maximal firing rates observed during MVC in
vivo and the stimulation frequencies required for maxinal isometric torque with NMES.
Firstly, the CNS may use very short intervals between the first firings, resulting
in double discharges (doublets) when a contraction is initiated, especially when the intent is
to perform the contraction as fast and strong as possible60,88,89. It is known that doublets and
triplets markedly enhances force when added to a submaximal firing rate via the so-called
catch-like property of skeletal muscle27,90,91. This force enhancement may last up to a few
seconds before it dissipates27,64. Such short bursts of extra impulses both in the beginning and
later on in the contraction89 may in part explain the paradox of how high forces can be
reached during maximal voluntary contractions despite seemingly suboptimal firing rates
when averaged across af few seconds.

However, it has been reported that average maximal firing rates in the vastus
lateralis can increase with strength training92,93, and that several measures of activation levels
Accepted Article
of the quadriceps correlate well with the measured maximal firing rates in the vastus
lateralis94. Moreover, although earlier studies reported near-maximal activation of the
quadriceps during MVC (e.g. Woods et al95), several more recent studies have found that
even with maximal voluntary effort, quadriceps muscle activation is typically only about 85-
90% during isometric and slow dynamic contractions96-101, although still others have found
near maximal (~95%) voluntary activation in the vastus lateralis (e.g. Pucci et al102).
The study of Pucci et al102 is of particular interest, as it demonstrated not only
high voluntary activation of the quadriceps but also relatively high maximal firing rates
during MVC (~42 Hz). Notably, the discharge rates increased drastically from ~20 Hz to ~42
Hz between 75% and 100% of MVC. This clearly indicates the importance of approaching
maximal voluntary activation in order to reach near-maximal firing rates even when these are
expressed as mean values for all units. Collectively, these findings on voluntary activation
and motor unit discharge rates strongly suggest that firing rates in the quadriceps during
MVCs are often not high enough to elicit the maximum torque capacity of this muscle in
subjects without resistance training experience.
Secondly, it is important to note that many of the investigations on firing rates
versus force development have reported average maximal rates. It is known that human
muscles contain motor units with a wide spectrum of contraction times of the muscle fibres
ranging from slow to fast38, which is probably explained by the continuum of fibre types and
subtypes discussed earlier in Section 3.1. In the study of Buchtal & Schmalbruch38, there
were 3-4 fold differences in contraction times between the fastest and the slowest bundles of
muscle fibres in the biceps brachii and triceps brachii when the bundles were activated in situ
in human subjects by intramuscular stimulation. In line with these estimates, Faulkner et al15

reported a 3-fold difference in maximal shortening velocity between bundles of human type
II fibres and bundles of type I fibres stimulated in vitro at 37° C.

Accepted Article
The short contraction times of the type II fibres means that these require much
higher discharge rates to reach tetanic tension than type I fibres68,103,104. This notion is
supported by the in vitro data in the report of Moulds et al86. Although 50-60 Hz was required
for generating maximal force in a given biopsy sample, 80% of maximum force was reached
already at ~15 Hz in bundles consisting of 80% type I fibres, whereas bundles made up of
57% type II fibres required ~30 Hz to reach 80% of maximal force86. In line with this,
Grimby et al103,105 reported that fast motor units with contraction times of 40 ms had maximal
firing rates of ~50 Hz (calculated from periods of 250 ms) in vivo, whereas slow units with
contraction times in the range of 60-90 ms had maximal firing rates of ~30 Hz. The fastest
units observed by Grimby, Hannerz and colleagues65,105 were characterised by maximal firing
rates of 65 Hz. Even greater ranges were found by Bellemare et al104, who reported that the
mean maximal firing rates for the biceps brachii and the adductor pollicis varied largely in
proportion to their respective twitch contraction and half relaxation times and that for each
muscle, the firing rates distribution covered approximately a fourfold range about the mean
value. Given the wide range of firing rates during maximal contractions, it can be argued that
reporting only average maximal firing rates provides relatively limited information on motor
unit behaviour during contractions of maximal efforts.
Thirdly, the time window used for calculating the discharge frequency has
varied considerably between studies; e.g. from the shortest interval of four102 and five
consecutive spikes67,94,106,107 to the greatest number of discharges in 250 ms intervals65,108 and
1 second intervals104,109. This can in part explain the discrepancies in the maximal firing rates
between different studies even when the same muscles have been investigated. For the biceps
brachii, Christie et al67 reported 37 Hz compared with 31 Hz in Bellemare et al104, whereas

Gydikov & Kosarov109 found no units firing higher than 24 Hz even at 100% of MVC.
However, Gydikov & Kosarov109 investigated firings rates only when steady-state forces had
Accepted Article
been reached, which probably also contributed to the low values. The importance of the
criteria for firing rate calculations is further underscored by the study of Van Cutsem et al89,
who observed that the interspike intervals increased for the first four discharges during
ballistic contractions, and that the increase was particularly marked for the interval between
the third and fourth spike. The increase in interspike intervals with increasing numbers of
successive discharges is likely at least in part due to spike-frequency adaptation, discussed in
Section 3.6.
Taken together, these observations help explain the discrepancy between the
maximal firing rates required for maxinal force development during electrical stimulation and
those observed during MVCs in vivo. Nevertheless, as will be outlined below, it may well be
that not all motor units tetanize during maximal effort contraction.
3.4. Can high threshold motor units fire at tetanic frequencies during maximal
voluntary contractions?
A current debate revolves around the questions whether high threshold units reach firing rates
sufficiently high for tetanic tension during voluntary maximal activation, and if the discharge
frequencies of these high threshold units really are higher than those of the earlier recruited
low-threshold units. In a series of investigations spanning over several decades, De Luca and
coworkers presented evidence and arguments which indicate that the firing rates of earlier
recruited motor units are greater than those of later recruited motor units during voluntary
isometric contractions even up to 100% of MVC69,110-113. Several other investigations have
arrived at similar results68,114,115. This observed property of higher firing rates in the earlier

versus the later recruited units all the way up to maximum force development has been
termed the ”onion skin” phenomenon111.

Accepted Article
However, as reviewed by Enoka & Duchateau116 and Piotrkiewicz & Türker117,
other researchers have reported results which support that high-threshold units can reach
higher firing rates than low-threshold units during near-maximal to maximal
contractions65,67,68,105,106,108,109,118–120. Christie et al67 observed that motor units recruited at
higher forces tended to demonstrate higher maximal discharge rates, although the relationship
was weak (r2 = 0.08).
For several reasons, these contrasting findings are not necessarily mutually
exclusive. The first one relates to the previous point on whether the motor unit firing rates
observed during MVCs in humans are truly maximal. As argued above, this often appears not
to be the case for the quadriceps in untrained subjects. Given that motor units with the highest
recruitment thresholds generally also have the fastest contraction times71,103,105, these must be
activated with the highest discharge rates to reach tetanic tension, as discussed in Section 3.3.
with reference to type I and type II fibres. There is some evidence from other muscles than
the quadriceps that this is possible, but typically only for short periods in untrained subjects.
In a series of studies on the tibialis anterior and on the extensor digitorum brevis
muscles, Grimby, Hannerz and co-workers65,71,103,105,121 found populations of very high-
threshold units that typically fired intermittently in bursts lasting 100-200 ms with short
intervals in between, and that these could continue during maximal voluntary efforts for only
brief periods before ceasing to fire. This typically happened after a few seconds when the
discharge rates of these units had dropped to 25-30 Hz. As noted in the previous section, the
highest frequencies observed for this type of units were in the range of ~50-65 Hz.
Interestingly, Hannerz65 also reported that the units recruited above 80% of maximum
isometric tension in the tibialis anterior were all of the intermittently discharging type.

Similarly, Gatev et al122 reported the recruitment of intermittently firing motor units above
80% of MVC in the flexor pollicis brevis. The intermittent ”phasic” firing patterns which
Accepted Article
seem characteristic of these units73 and their near-maximal thresholds could mean that these
units are easily missed and/or difficult to follow in experimental settings. Grimby &
Hannerz71 raised a similar point in noting that continuously firing long interval motor units
were more easily identified than intermittently firing short interval motor units (the latter
presumably representing very high threshold units).
A second reason relates to a point raised by Hannerz65, who noted that
measuring ”maximum frequency” during sustained contraction based on the highest
discharge in only 250 ms increases the maximum frequencies as compared with those
obtainable from a whole second, but it does so significantly only for motor units with high
recruitment thresholds. From this and the earlier discussion in Section 3.3., it is clear that the
criteria for calculating the maximum firing rates of different units can greatly impact the
results and conclusions derived hereupon.
3.5. Type of activation versus firing rates for high threshold motor units
A third aspect, related to those discussed above, concerns the manner in which the
contraction is performed, and the impact that this has on the firing rates of fast motor units.
Grimby et al103 illustrated this by following the fastest motor unit that they observed in one of
their subjects during varying types of isometric contractions, and showed that this unit
increased its frequency in proportion to the rate of force development (Figure 5 in Grimby et
al103). Based on the criteria of Harwood et al107 of at least 5 consecutive action potentials,
approximate firing rates for this unit can be calculated. With slowly increasing contraction,
the unit fired at a relatively steady rate of ~30 Hz after about 2 seconds. During maximal
sustained contraction, the unit fired at ~50 Hz for periods of 100-200 ms. With an

accelerating but submaximal contraction, the unit reached as high as ~100 Hz for a time span
of ~50 ms. Finally, during ballistic contractions, it discharged at ~100-140 Hz for periods of
Accepted Article
~67 ms.
In the same paper by Grimby et al103, the authors provided an example of the
different dischage behaviour between two opposite types of motor units, continuously firing
long interval motor units and intermittently firing short interval motor units, possibly
consisting of type I and type IIX fibres, respectively (discussed in Grimby & Hannerz71).
During the first second of a strong sustained contraction, the intermittently firing unit
discharged at 40 Hz, which dropped to 20 Hz during the next second of the contraction. In
contrast, the continuosly firing unit maintained 20 Hz during the first 2 seconds of the same
sustained contraction. With prolonged contraction, only the continuously firing unit
responded.
Compelling evidence for the existence of intermittently firing high threshold
units in humans was presented by Warmolts and Engel123,124, who performed open biopsy
electromyography in the biceps brachii and the quadriceps in patients with low-grade
neuropathy but with otherwise normal levels of muscle strength. The biopsies were taken
from the muscles after motor unit patterns had been recorded in the same areas during
different types of voluntary contractions. A few samples were almost entirely (96-100%)
composed of type I and type II fibres, respectively. When the pre-biopsy recordings were
analyzed, it was found that the almost pure type I areas had sustained discharges throughout
the entire force range, with maximum rates of 18-20 Hz at full voluntary efforts. Conversely,
the nearly pure type II areas were only active during strong or sudden activations, discharged
only for brief periods between 0.5 to 5 seconds, and reached peak frequencies of between 16-
50 Hz. Although caution is warranted due to reinnervation of denervated muscle fibres,
Grimby125 has argued that abnormal firing behaviour is seen only during the early phase of

reinnervation, and that at later time points the firing behaviour is no different from normal
motor units.
Accepted Article
Further support for the importance of the type of activation on motor unit
discharge rates comes from intramuscular electrode studies on dynamic movements, which
have shown markedly greater firing rates with concentric contractions compared with
isometric contractions, even when using the same relative intensity for each type of
contraction107,126. Harwood et al107 showed that low-load (25% of MVC) dynamic elbow
extensions with maximal effort resulted in greater firing rates in the anconeus muscle than
isometric contractions at 100% of MVC (39 Hz vs 24 Hz). Moreover, Grimby73 noted that
although lower-threshold units could also reach very high firing rates during ballistic
contractions, higher-threshold units reached the greatest discharge frequencies. Similarly, the
results of Desmedt & Godaux60 suggest higher firing rates in higher-threshold versus lower-
threshold units during ballistic contractions.
Collectively, this evidence clearly indicates that high-threshold units can reach
higher discharges rates than low-threshold units during brief maximal contractions,
particularly during accelerating, fast dynamic and ballistic contractions. This conclusion is in
line with Enoka & Duchateau116, who with reference to gradual increases in force stated that
”recordings of motor unit activity with fine-wire electrodes in which action potentials can be
observed directly often show that the peak discharge rate achieved during such tasks is
greater for later recruited motor units”.
It must again be stressed that the time periods during which the highest
threshold units reach their peak rates are typically very brief. Summarizing their results on
maximal voluntary contractions in the tibialis anterior and the extensor digitorum brevis
muscles, Grimby73 noted that ordinary subjects did not drive the highest thresholds units for
more than a few seconds, despite having done their utmost to the point of being very

exhausted. Possible causes of this inability in untrained subjects to maintain the firing in the
highest threshold motor units include motor unit adaptation (also known as spike-frequency
Accepted Article
adaptation, see below) and a selective increase in the recruitment thresholds for these units
due to fatigue (reviewed by Grimby73,127, see also Grimby et al105).
3.6. The possible roles of spike-frequency adaptation and small muscle afferents in
limiting maximal firing rates
With slow ramp contractions and maximal contractions lasting longer than a few seconds,
high-threshold motor units do not necessarily reach higher discharge rates than low-threshold
units. It deserves to be noted that the MVCs in the studies of De Luca and colleagues69,110-113
were performed with gradually increasing effort for about 10 seconds before the MVC was
reached, i.e. a rate of 10% MVC/s. During such conditions, the onion skin pattern of firing
rates of the recruited units may well prevail over other patterns. Interestingly, Erim et al112
noted that during such contractions, the subjects failed to reach their pretest MVCs, which
were performed during 3 seconds with no restrictions on the rate of torque increase. Erim et
al112 conceded that their subjects could have been affected by fatigue during their slow ramp
contractions. Inspection of their figures suggests that their subjects only reached ~90% of
MVC during the 10 s ramp contractions.
Since the same type of slow ramp contractions were used in many of the reports
from De Luca and coworkers, it is possible that the highest threshold units never reached
maximal firing rates in these studies. This possibility is supported by the findings of Oya et
al68, who studied motor unit recruitment in the soleus muscle during slow ramp contraction at
a rate of 10% MVC/s. They noted that while higher threshold units reached higher maximal
firing rates than lower-threshold units, the motor units recruited above 89% of MVC showed
peak discharge rates which were as low as those units recruited at or below 30% of MVC. A

similar tendency may be discerned in the results of Christie et al67, who also used slow-ramp
MVC contractions at a rate of 10% MVC/s. In Fig 2 in Christie et al67, the motor units
Accepted Article
recruited at or above ~87% of MVC seem to generally fire about ~10 Hz slower than the
units recruited between ~80-86%. The surface EMG data in the same study reveal a clear
trend towards a decreased mean power frequency (MPF) during the last 3 seconds of the 10-
second ramped-up MVC, suggesting development of fatigue.
Further evidence for the negative effects of previous muscle activity on the
maximal firing rates comes from a study by Van Cutsem and Duchateau128, who observed
decreased discharge rates in motor units during ballistic contractions which were preceded by
3-4 second isometric contractions at 25% of MVC compared to contractions initiated from a
relaxed state. Interestingly, the lower firing rates during ballistic contractions performed
immediately after low-intensity contractions were noted also in high-threshold motor units
that were not recruited at 25% of MVC, which suggests that the 25%-contraction negatively
affected the entire motor unit population. Harwood et al129 also reported lower firing rates in
maximal velocity contractions when these were preceded by submaximal fatiguing
contractions. Taken together, these findings strongly suggest that near-maximal firing rates
may not be reached in the highest-threshold motor units during slow isometric ramp
contractions, and that this type of contraction does not reflect the behaviour of these units
during the very activities that they are primarily designed for, i.e. brief very strong and fast
contractions.
The explanation may at least in part reside in the phenomenon of spike-
frequency adaptation in the motor units, which refers to the decrease in motor unit firing rate
as a function of time130. It has been shown that this acute phenomenon has an early rapid
phase which lasts ~2 seconds, which is then followed by a late phase of adaptation, with most
of the decline taking place during the initial phase130. One of the underlying mechanisms is

afterhyperpolarization, which is longer in slow motor units and shorter in fast units131-133. The
extent of spike-frequency adaptation appears to be greatest in the fast-fatigable units and least
Accepted Article
in the slow fatigue-resistant units130. This may in part explain both why very high firing rates
cannot be maintained during MVCs, and why maximal discharge rates are not seen in the
highest-threshold units during slow ramp contractions. This notion conforms well to the
observations that these units have a much higher fatigability also in humans37 and that type
IIXa and IIAx muscle fibres have the longest recovery times after PCr depletion due to
intense exercise49.
However, with training and extraordinary motivation, it is possible to drive
high-threshold units for longer than a few seconds at a time73. Given that the discharge rates
of intermittently firing units can decline already after the first second of high-force
contractions103, an improved ability to drive very high-threshold units seems likely to be one
of the major mechanisms among the host of so called neural adaptations, and the strength
increases that result from systematic training. It would also appear that if the adaptations are
especially pronounced in these units, they could easily be missed in studies on firing rates
before and after a period of strength training if only mean maximal values are reported. This
risk is underscored by the estimate that motor units innervating type IIX myofibres may
comprise only about 4% of the motor units while contributing with as much as ~18% of the
muscle fibres in a limb muscle such as triceps brachii64.
Another pathway which can impact on motor unit discharge rates originates
from small diameter afferents in the muscle. Classic physiological experiments have shown
that fatiguing contractions result in lower firing rates in the motor units, that complete
occlusion of the blood flow to the muscles after exercise maintains the suppression of firing
rates, and that these recover to normal values only once the circulation is restored95,134.
Bigland-Ritchie and colleagues134 suggested that group III and IV muscle afferents were

ideally suited to mediate such reflex modulations in motor unit firing rates during acute
fatigue and it is now well accepted that increased firing in these afferents leads to reduced
Accepted Article
motor neuron firing, The precise pathway for this effect is not known135, but it has been
proposed that group III/IV afferents act upstream of the motor cortex to inhibit voluntary
descending drive136.
How group III and IV afferents affect the different populations of motor units is
also not entirely clear, but it has been suggested that they decrease the firing rates of fast
fatiguing units in particular137, which in humans would be those with type IIX and IIAX
fibres. In contrast, others have provided arguments and evidence suggesting that group III/IV
afferents primarily affect low-threshold units, leading to slowing or derecruitment of these
small units80,81. However, the observation that saline injection and post-exercise ischemia
affect group III/IV afferent input differently (discussed in Martin et al.138) warrants caution in
extrapolating findings from saline injection studies to fatigue occuring during physiological
conditions. Interestingly, firing of group III and IV afferents may also influence
afterhyperpolarization of spinal motor neurones and thereby possibly also affect spike-
frequency adaptation, although the effects of this input during repetitive motor unit firing
remain to be elucidated (discussed in Windhorst et al137).
Spike-frequency adaptation may in part explain some paradoxical findings in
the literature on motor unit firing rates during fatigue. Garland et al139 observed that during an
isometric contraction of the triceps brachii at 20% of MVC held to fatigue, the motor units
which were active already from the beginning of the task (presumably S and perhaps also
some FR units) typically displayed marked decreases in their firing rates throughout the
contraction even as task failure neared, although by then some early recruited units had
started to increase their rates again. In contrast, motor units which were recruited later in the
contraction, which were identified as higher-threshold units, typically increased their firing

rates throughout the remainder of the task, with only a few exceptions. Figure 1 in Garland et
al139 also suggests that some of the lower-threshold units may have ceased to fire after
Accepted Article
reaching their lowest discharge rates, but this was not commented by the authors. It is
relevant to note that blood flow in the triceps brachii can become occluded already at 25% of
MVC and that sustained contractions at 20% of MVC are followed by pronounced
hyperemia140,141, suggesting that the muscle blood flow is insufficient at this load. Therefore,
the findings of Garland et al139 have possible implications for the activation of motor units
during ischemic exercise as well as for the interpretation of EMG results from BFRRE
studies, which will be discussed later (Section 7.5.).
3.7. The role of small muscle afferents in limiting muscle activation during fatiguing
exercise with large muscle mass
Studies have shown that skeletal muscle afferent input (including group III/IV afferents) to
the CNS increases with the amount of active muscle mass142-145. Interestingly, this muscle
mass dependence of afferent input to the CNS also affects endurance and muscle activation in
different tasks. Thus, Matkowski et al146 found greater losses in MVC torque and voluntary
activation (assessed by interpolated twitch analysis) after unilateral compared to bilateral
isometric knee extensions sustained to fatigue at 20% of unilateral and bilateral MVC,
respectively. Notably, despite similar ratings of perceived exertion (RPE) at the end of the
tasks (20 on the Borg 6-20 RPE scale), endurance time was shorter and EMG increases in the
quadriceps were reduced in the bilateral task. Resting doublet twitch force decreased after the
unilateral task only.
Similarly, Rossman et al147 demonstrated greater peripheral fatigue (e.g.
amplified decreases in MVC and twitch force) in unilateral dynamic knee extensions than in
bicycle exercise when both were performed to task failure. In another study, Rossman et al148

reported greater peripheral fatigue in unilateral than in bilateral dynamic knee extensions
performed to task failure, as judged by greater EMG increases and decreases in MVC and
Accepted Article
twitch forces. Rossman et al148 suggested that the CNS tolerates a greater magnitude of
peripheral fatigue when the source of group III/IV afferent feedback is limited to a small
muscle mass. Taken together, these findings are highly relevant for the activation of motor
units during BFR training with small and large muscle mass exercises, as discussed in greater
detail in Section 7.2.
4. Fatigue and recovery in the muscle fibre types with ischemic exercise
4.1. Fatigue in the muscle fibre types during ischemic and anaerobic exercise
A discussion on the fatigue processes in the muscle fibres during ischemic conditions is
motivated by the fact that fatigue is inherent and constitutes a major driving factor behind
muscle fibre recruitment in BFRRE (reviewed by Wernbom et al5). In an early study on
human muscle during ischemia, Buchtal & Schmalbruch38 showed that 20-45 minutes of
circulatory occlusion eliminated electrically induced twitch contractions of slowly
contracting muscle fibres in the biceps brachii, the triceps brachii and the gastrocnemius
muscles, while faster contracting fibres still responded to twitch stimulation with
characteristically short contraction times. In contrast, Dietz149 suggested that high threshold
motor units rather than low threshold units are inhibited during acute ischemia. Nevertheless,
studies on rat muscles have shown a greater decline in twitch force in slow muscle than in
fast muscle during the first 45 minutes of ischemia, particularly during the first 15-30
minutes150,151. Likewise, Gossen et al152 reported markedly greater declines in twitch force in
slow contracting motor units than in fast units during very low-force (~5-10% of MVC)
voluntary ischemic contractions in human subjects.

The available evidence thus suggests that during rest and very low-force
contractions, slow motor units are relatively more sensitive than fast motor units to ischemia
Accepted Article
of short to moderately long durations. This may be explained by the greater oxygen
consumption at rest in slow muscle compared with fast muscle153. Studying stimulated low-
force contractions in rat soleus muscle under anaerobic conditions, Sahlin et al154 concluded
that the decline in relative force was similar to that previously observed in fast twitch muscle
and that the soleus cannot be termed fatigue-resistant under anaerobic conditions. In should
be noted that with very low-force (~5-10% of MVC) voluntary ischemic contractions, it is
likely that only type I fibres are used155. With greater voluntary efforts due to higher loads
and/or fatigue in type I fibres, the type II fibres most likely will also be recruited and undergo
fatigue during ischemic exercise, and probably eventually to a greater degree than type I
fibres, as discussed elsewhere in this review.
The mechanisms of fatigue in muscle fibres during ischemic contractions are
probably in part related to the effect of hypoxia on PCr levels. Ischemia directly induces
muscle fatigue in humans, and the decrease in twitch force with increasing occlusion pressure
is proportional to the decrease in tissue oxygenation156. Intramuscular oxygen partial
pressures (pO2) have been correlated to the PCr levels during rest and exercise in humans157.
Bylund-Fellenius et al157 proposed that the correlation between the pO2 and PCr can be
explained by the equilibrium maintained by creatine kinase, since a direct relationship was
also found between the ATP/ADP ratio and PCr. During high energy demand, ATP is
initially relatively constant while PCr breaks down to creatine (Cr) and inorganic phosphate
(Pi)158. Increased Pi is one of the major causes of fatigue and can together with decreased
calcium (Ca2+) release from the sarcoplasmic reticulum (SR) explain much of the decline in
muscle force159.

Recent evidence also shows that increased hydrogen ions (i.e., lower pH) can
influence fatigue by acting synergistically with elevated Pi to depress force160,161 and that by
Accepted Article
considering these factors together with compromised Ca2+ release from the SR, it is possible
to account for much of the observed losses in force and velocity162. Strong support for a
synergistic role for pH on fatigue-induced force depression can be found in an in vivo study
on humans by Hultman et al163, who used neuromuscular electrical stimulation (NMES) to
evoke contractions during circulatory occlusion at a force corresponding to 50% of MVC,
with or without oral administration of ammonium chloride (NH4Cl) to vary intracellular pH
during exercise. Hultman et al163 showed that lower pH levels at the same degree of PCr
depletion were associated with a significantly lower force production after a 75-s ischemic
continuous isometric contraction, ending at 45% vs 55% respectively of the initial force level.
This investigation was one in a series of landmark studies in which Hultman
and colleagues investigated muscle fibre metabolism in the human quadriceps during
complete occlusion (~250 mm Hg) combined with NMES contractions52,163-169. These studies
helped to shed light particularly on the regulation of glyogenolysis and on the fatigue
processes during anaerobic exercise. Of particular interest for the physiology of BFR
exercise, Greenhaff et al52 reported more than 10-fold increases in glycogenolytic rates in
type I fibres in vastus lateralis muscles after NMES combined with occlusion compared with
fibres obtained after NMES alone. In contrast, the glycogenolytic rates in type II fibres with
occlusion were only marginally (not significant) higher than those observed during the free
blood flow condition. The NMES protocol included 20 maximal 1.6 s isometric contractions
with 1.6 s rest between each contraction, with a stimulation frequency of 50 Hz.
It can be argued that repeated 50 Hz stimulation is not physiological, but studies
from the same group in which 52 contractions with 20 Hz were combined with occlusion
resulted in even higher lactate levels in mixed muscle samples than in Greenhaff et al52 as

well as very low PCr values in both type I and type II fibres (Hultman & Sjöholm166,
Söderlund & Hultman167, Söderlund & Hultman169). The lactate in mixed muscle increased to
Accepted Article
128.9 mmol per dry weight, corresponding to ~29.6 mmol per kg wet weight, assuming a
77% water content in human skeletal muscle170. Such extreme lactate values in mixed muscle
fibres are only possible if glycogenolysis is extensive in both fibre types. Thus, ischemia can
drastically increase lactate production and decrease muscle fibre pH in type I fibres also
during muscle contractions at physiological firing rates.
Additional evidence for an important role of low pH in ischemic exercise
fatigue can be found in the investigation of Hultman & Sjöholm166. In their report, it was
shown that isometric force was still relatively well maintained (~80% of baseline) after 26
ischemic contractions, when PCr had already reached minimal levels. After 39 and especially
after 52 contractions, the force decline was much greater and mirrored the continued increase
in lactate levels, while PCr was essentially constant. However, it is possible that the low PCr
levels and consequent Pi increases may after a delay have resulted in Ca2+-Pi-precipitation in
the SR, causing decreased Ca2+ release from the SR158,159. In addition, the decreased pO2 may
reduce Ca2+ release within a few minutes as will be discussed below. Interestingly, the force
decline in the study of Hultman & Sjöholm167 was also related to decreases in ATP, which
dropped to as low as 37% of baseline levels, and to the decline in the ATP/ADP ratio. Very
low ATP levels, which can be especially pronounced in type IIX fibres following exhaustive
muscle exercise, may affect force and velocity negatively via multiple mechanisms, several
of which ultimately decrease the Ca2+ release from the SR158. Collectively, the drops in PCr
and ATP, and the corresponding changes in Pi, pH and ADP along with decreased Ca2+
release, probably explain a large part of the peripheral fatigue accumulated during ischemic
contractions.

Of note, Tesch et al171 and Söderlund & Hultman169 demonstrated 11-13%
higher resting PCr concentrations in type II than in type I fibres. This was confirmed and
Accepted Article
extended by Karatzaferi et al49, who showed that the resting PCr levels were highest in type
IIXa fibres (composed of 50-100% type IIX MHC) and lowest in type I fibres, with the type
IIXa fibres displaying ~33% higher values than type I. In contrast, type IIA fibres and type I
fibres had similar PCr levels, resulting in 11% greater PCr content in the type II fibres
overall. In the same study by Karatzaferi et al49, 25 seconds of maximal sprint cycle exercise
produced similar low absolute PCr levels the different fibre types immediately post-exercise,
with the type I fibres only slightly higher than the type II fibres. These findings strongly
suggest that the percentage drop in PCr levels can be greater in type II fibres with exhaustive
anaerobic and ischemic exercise and that the corresponding increase in Pi will also be greater
in type II fibres, particularly in the fastest subtypes. This may contribute to the greater torque
decreases seen with repeated concentric contractions in individuals with a high percentage of
type II fibres170, and to the greater fatigue in individuals with higher percentage type IIX
compared to persons with lower type IIX fibre proportion but who otherwise demonstrate the
same overall % of type II fibres172.
In addition to decreased PCr and pH levels, cellular ischemia can also suppress
Ca2+ release from the SR. Sun et al174 demonstrated that hypoxia (1% O2) markedly depresses
Ca2+ release and force development in comparison to normoxia (20% O2), with intermediate
forces at 5% O2. Interestingly, the effects of hypoxia were mediated by a decreased
production of hydrogen peroxide (H2O2) from NADPH oxidase 4 (Nox4). Nox4 was found to
produce H2O2 in direct proportion to the pO2, and the authors further reported that consequent
oxidation of a small set of cysteine thiols on the ryanodine receptor (RyR1) results in
increased RyR1 activity and Ca2+ release in isolated sarcoplasmic reticulum and in cultured
myofibres and enhanced contractility of intact muscle. They proposed that Nox4 thus acts as

an O2 sensor in skeletal muscle. The effects of pO2 on Ca2+ release and contractile force were
seen after only 10 minutes of hypoxia. Furthermore, a study on rat skeletal muscle showed
Accepted Article
that slow red muscle fibres have >2-fold greater expression of Nox4 and >2-fold higher
baseline production of H2O2 than fast white muscle fibres175. This raises the intriguing
possibility that greater Nox4 expression and baseline activity may contribute to the seemingly
greater pO2-sensitivity of force production in type I myofibers.
4.2. Possible contributions of low-frequency fatigue and neuromuscular
transmission failure to torque decreases in ischemic exercise
A persistant lowering in the ratio between stimulated forces at low stimulation frequencies
(typically 20 Hz) and those obtained at higher frequencies (50-100 Hz) may be observed after
the exercise-induced metabolite changes have reverted back to baseline levels. This is
denoted as low-frequency fatigue, reflecting an impairment in the electromechanical coupling
and consequent reductions in Ca2+ release176,177. In support of this phenomenon, Wernbom et
al178 reported a decreased 20/50 Hz ratio in the vastus medialis 3 minutes after an acute
fatiguing low-load BFRRE bout, when the partial BFR (~55% of complete occlusion
pressure) was still maintained. The 20/50 Hz ratio was reduced by 34% (Wernbom,
unpublished results).
Importantly, decreased myofibrillar Ca2+ sensitivity by factors such as increased
Pi and lowered pH can cause a similar low-frequency fatigue158. The acute changes reported
by Wernbom et al177 could therefore be due to decreased myofibrillar Ca2+ sensitivity and/or
decreased Ca2+ release. In support of the former alternative, 20/50 Hz ratio was no longer
significantly changed at 1 hour post-exercise. However, Sieljacks et al179 reported a reduced
20/100 Hz ratio in the quadriceps as a whole at 1 hour and 1 day, respectively, after a similar
acute BFRRE protocol as that employed by Wernbom et al178. Thus, there is evidence for

both acute and prolonged low-frequency fatigue with acute BFRRE performed with multiple
sets to failure.
Accepted Article
Wernbom et al178 further reported 61-62% reductions in MVC torque during the
first 2 minutes post-exercise, and no improvement between the 1 and 2 minute post-exercise
MVCs during maintained partial BFR. Part of the decline during post-BFRRE MVCs with
occlusion still in place may be explained by a reduced central excitatory drive. With a similar
protocol, we observed that EMG in the quadriceps during 1 minute post-BFRRE MVC with
maintained BFR was markedly lower compared with pre-exercise MVCs (~30% reduction,
Wernbom unpublished data 2007). Similarly, Yasuda et al180,181 reported reduced EMG
activity in the biceps brachii during post-exercise MVCs with sustained partial BFR, and
even greater decreases in EMG after exercise with complete occlusion. Motor unit
derecruitment and reduced firing rates would act in concert with an impaired
electromechanical coupling and decreased Ca2+ sensitivity, which in turn would contribute to
decrements in MVC torque.
However, the acutely reduced 20/50 Hz ratio seen after fatiguing BFRRE by
Wernbom et al178 could also in part be due to a decreased excitability of the muscle fibres
and/or transmission failure in some fibres. Importantly, both these factors may affect the
EMG signal amplitude165,182. Hultman & Sjöholm165 measured EMG in the quadriceps during
a continuous electrically stimulated (20 Hz) contraction which lasted 75 seconds, and showed
decreased EMG in parallell with decreased force, thus demonstrating that decrements in
EMG can have a peripheral origin. The contraction started at ~70% of MVC and ended at
~50% MVC. At 50-75% of MVC an almost complete occlusion of the blood flow occurs in
the quadriceps due to the high intramuscular pressures183,184, thus the EMG decreases in the
investigation of Hultman & Sjöholm165 were likely caused by the combined effects of
ischemia and the prolonged contraction. In line with this scenario, Cupido et al185 showed in

the human biceps brachii that continuous 20 Hz stimulation in combination with ischemia
(300 mm Hg) caused a decrease in the M-wave size after about 60 s, which continued to
Accepted Article
decrease steeply toward values approaching zero during the next ~60 s.
Working with the tibialis anterior and soleus muscles, Galea186 confirmed these
findings with 20 Hz and 30 Hz stimulation. In parallell with the decline in M-wave
amplitude, tetanic torque decreased, reaching 4-20% of control values after 3 minutes. In
contrast, little or no changes were seen at the same time points during 20 Hz stimulation with
free circulation185,186. Studying voluntary low-force contractions in the soleus, Leonard et
al187 found a 52% reduction in M-wave readings after 105-126 coupled concentric-eccentric
repetitions performed in approximately 3-4 minutes during ischemia (300 mm Hg). A
lowered M-wave size suggests a decreased effectiveness of neuromuscular transmission
and/or action potential propagation in muscle fibres188, respectively, although how M-wave
changes should be interpreted is debated189. Nevertheless, the consistent decreases in M-wave
size reported by Leonard et al186, Cupido et al184 and Galea185 as well as in the evoked EMG
responses by Hultman & Sjöholm165 are very similar to those seen with high-frequency
fatigue190. Thus, acute EMG and M-wave decreases of this magnitude seem to be associated
with a reduced muscle fibre excitability and/or neuromuscular junction (NMJ) transmission
failure.
With regard to the latter, Dahlbäck et al191 studied impulse transmission during
ischemic contractions in human extensor digitorum communis muscles. In the recordings of
action potentials from pairs and sometimes also three and four muscle fibres that belonged to
the same motor unit, they found that one of the fibres in a pair dropped out after a certain
number of discharges, and that a total block typically occurred after between 3500-7000
impulses. The block was ascribed to failure of neuromuscular transmission at the motor end-
plate. Notably, at a rate of ~10 Hz, a 50% block of one of the potentials occurred after 6.5

minutes, and in recordings of units with three and four fibres, the first fibres dropped out
already after 2.5-3 minutes, which corresponds to a total of about 1500-1800 muscle fibre
Accepted Article
action potentials. Another interesting finding was that when there were only 30 seconds of
rest with free circulation between the periods of ischemic contractions, the time until these
changes occurred was ~35% shorter. Dahlbäck et al191 emphasized that the synthesis of
acetylcholine is an aerobic process which is probably negatively affected by the lowered pO2
during ischemia. They concluded that during ischemia, the synthesis of acetylcholine is likely
inhibited and that the block occurs when synaptic stores of acetylcholine have been emptied.
Presynaptic failure of neuromuscular transmission can also occur in the motor
axon, especially at the branch points. Sieck & Prakash192 proposed that the probability of
axonal conduction block may be greater in FF units, due to the typically larger size and
greater number of axonal branches in these. They also suggested that this might explain the
observation of Sandercock et al193 of failure to evoke muscle fibre action potentials in some
of the muscle fibres in a given motor unit. The findings of Sandercock et al192 of blocked
action potentials mainly concerned the responses to a 80 Hz stimulation protocol which
induced high-frequency fatigue. Nevertheless, during high-frequency fatigue, there appears to
be more or less parallell decreases in M-wave, EMG and force (Figure 5 in Bigland-
Ritchie190). Notably, ischemia is known to slow motor nerve conduction velocity within 10-
15 minutes during resting conditions194,195. Given this and the striking similarities between
the effects of high-frequency fatigue and the fatigue induced by low-to-moderate load
ischemic contractions on these parameters, it seems possible that axonal conduction block
can occur in some branches during ischemic exercise.
Ultimately, neuromuscular transmission efficacy depends on the end-plate
potential (EPP), which in turn can be impaired by both pre-synaptic and post-synaptic
mechanisms192. Muscle fibre excitability is an important post-synaptic factor, and the

combination of lowered acetylcholine release and decreased fibre excitability can result in
EPP rundown and eventually transmission failure196. It seems clear that ischemic exercise can
Accepted Article
negatively affect both pre and post-synaptic mechanism, which may explain the findings of
Dahlbäck et al191. Moreover, results from rat muscle studies show that FF and F(Int) units are
generally more prone than FR and S units to M-wave decreases during repetitive
stimulation192. Studies on rat muscles also indicate that fast fibres have a much greater Na+-
K+ leak/pump ratio than slow fibres, and the resulting faster rise in extracellular K+ and
decreased Na+ gradient results in an accelerated loss of excitability in fast fibres197. Even so,
it seems reasonable to suggest that repetitive stimulation during ischemia can lead to
transmission failure also in some type I and type IIA muscle fibres, given that the S and FR
units are likely recruited much more extensively than F(Int) and FF units during low-force
contractions. This notion is supported by the fact that the motor units investigated by
Dahlbäck et al191 were primarily very low-threshold ones (see also Section 6.6).
The observation that some fibres, probably including type I fibres, can ”drop
out” due to neuromuscular transmission failure already after 2.5-3 minutes of relatively
moderate firing (~10 Hz) during ischemia has implications for muscle fatigue also during
low-load BFRRE. Although these studies investigated muscle contractions during complete
vascular occlusion, their findings suggest the possibility that the decreases in EMG in post-
BFRRE MVCs observed by Yasuda et al180,181 and in our labs (Wernbom et al. unpublished
results 2007) could in part have been due to transmission failure in some of the muscle fibres,
in addition to varying degrees of reduced excitability in active muscle fibres and lower firing
rates in many of the motor units.
Regardless of the relative contributions from each of the above fatigue
mechanisms, it is clear that there is a substantial peripheral fatigue component in BFRRE
which does not fully recover until free blood flow is restored. This notion is in line with

studies showing that if blood flow occlusion is maintained or applied immediately after
fatiguing exercise, there is no recovery of static and dynamic torque134,172. The dynamic peak
Accepted Article
torque results of Colliander et al172 even suggest a ~10% further decrease after a 1 minute rest
period with complete occlusion. This observation sheds light on the seemingly small
magnitude of post-exercise recovery of MVC reported by Yasuda et al181 and Wernbom et
al178 while partial BFR was still maintained. Fatigue will undoubtedly affect the force
development at the muscle fibre level, and hence also influence motor unit recruitment
patterns as well as modulate the mechanical and metabolic stimuli involved in BFRRE, as
discussed in detail below.
4.3. Force recovery in the muscle fibre types after ischemic and anaerobic exercise
In addition to depletion, Söderlund & Hultman169 also studied the recovery of PCr and ATP
levels in type I and type II fibres at different time points after ischemic contractions.
Although PCr levels were extremely low in both fibre types immediately post-exercise, a
faster repletion of PCr occurred in type I fibres than in type II fibres after 60 s recovery (to
~59% of baseline vs ~37%) with intact circulation. After 5 minutes of rest, PCr was no longer
significantly lower than baseline, although visual inspection of their graphs suggests that for
type II fibres, PCr was still ~22% lower while the type I fibres were back to pre-exercise
levels. Interestingly though, at 15 minutes post exercise, type II fibres had PCr levels which
were ~18% higher than baseline. The ATP levels were reduced to 60% and 53% in type I and
II fibres respectively immediately post-exercise, increasing to ~76-81% and ~61-63% at 20-
60 s post-exercise. At 5 minutes, the corresponding values were ~87 for type I fibres, and
~73% for type II fibres. At 15 minutes, ATP was no longer significantly lower than baseline
in type I fibres (~94% of pre) while type II fibres had still not recovered (~77%). Curiously,
these values are slightly lower than previously reported by Söderlund & Hultman167 using an

identical exercise protocol, where ATP levels were also reduced at 15 minutes post-exercise
in the type II fibres to ~91% of pre-exercise levels.

Accepted Article
These results suggest that full recovery of PCr levels in type II fibres after
severely depleting exercise may take considerable time, on the order of 10 minutes.
Furthermore, recovery of ATP after depleting ischemic exercise seems to take even longer
time in the fast fibres (>15 minutes). Unfortunately, no analysis of type II fibre subtypes was
conducted in the study by Söderlund & Hultman169. However, yet other studies have shown
that the rate of PCr recovery differs between the fibre types and subtypes after strenuous
anaerobic exercise, being quicker in both type I and pure type IIA fibres than in hybrid type
IIAX and almost pure IIX fibres49. After 1.5 minutes of recovery, PCr in type I fibres were
~64% of baseline levels, while type IIA levels were ~57%. In contrast, PCr levels in type
IIAx and IIXa (15-50% and 50-100% type IIX MHC, respectively) had only recovered to
~37% and ~33%. The rate of recovery of PCr after strenuous high-repetition isokinetic
exercise was positively correlated with the content of the oxidative enzyme citrase synthase
(CS), a marker of myocellular oxidative capacity198. Furthermore, type IIA fibres have
markedly higher oxidative capacity compared with type IIX fibres51,199-202. Taken together,
these results show that type I fibres replenish their PCr levels faster than type II fibres, and
that type IIA in turn recover considerably faster than both the hybrid type IIAX fibres and
pure IIX fibres.
Nevertheless, the significance of exercise-induced depletions and subsequent
recovery in PCr and ATP levels for acute muscle force capacity is not entirely clear. Sahlin &
Ren203 investigated the time courses of recovery for both MVC and various muscle
metabolites (including ATP, ADP, PCr and lactate) after sustained isometric knee extensions
at 66% of MVC which were held to fatigue (to ~50% of MVC). They showed that although
PCr was markedly depleted immediately post-exercise in mixed muscle samples and that

neither PCr nor lactate recovered to pre-exercise levels even with 4 minutes of rest (~85% of
pre-exercise for PCr), MVC was no longer significantly depressed at 2 minutes post-exercise
Accepted Article
(~95% of pre-exercise) and was fully recovered at 4 minutes of rest. Notably, MVC had
recovered to 80% already after 15 seconds of rest and to 90% after 30 seconds. Data from a
study with a very similar exercise protocol204 suggests that at these time points, PCr recovers
to only ~30% and ~40% of the baseline levels. Altogether, these observations confirm that of
Hultman & Sjöholm166 that even quite large degrees of PCr depletion are by themselves not
necessarily sufficient for large decrements in torque development, and therefore that there has
to be other factors behind the recovery of muscle force capacity in addition to PCr repletion.
In their study on repeated high-repetition isokinetic exercise, Jansson et al198
found that peak torque recovery between the sets was positively related not only to PCr
content relative to the resting values (r=0.63) but also to the degree of restoration of the
ATP/ADP ratio (r=0.69). Importantly, both these parameters as well as lactate recovery
correlated with CS activity, which in turn correlated with torque recovery (r=0.69). Using a
similar isokinetic protocol, Colliander et al172 showed that individuals with a dominance of
slow fibres (57% relative area of type I fibres) had an almost complete recovery of torque
after already 60 seconds of rest. In contrast, a group with a type II relative area of 70% had
only recovered to 80% of pre-exercise torque. As both groups had a mix of fast and slow
fibres, the differences in the in situ force recovery between type I and type II fibres were
likely even greater. Therefore, it seems clear that short rest intervals have a much more
negative effect on the recovery of force development in type II fibres than in type I fibres,
and that the recovery is probably more prolonged in the fastest subtypes of type II fibres (IIX,
IIAX) compared with type IIA.

With regard to BFRRE, it is important to note that the rest periods between sets
are not only typically brief (30-60 s), but also that most models of BFRRE involve occlusion
Accepted Article
also during the inter-set rest periods, which means that torque recovery between sets is
limited (see further below). However, it is likely that the force recovery in type I fibres is still
faster than in type II fibres. It should also not be overlooked that some studies have reported
signs of muscle damage and prolonged torque decrements lasting up to 48 hours post-
exercise after low-load BFRRE178,179,205 accompanied by low-frequency fatigue for 24 hours
post-exercise179. Cumming et al21 reported heat shock protein (HSP) responses from the same
acute BFRRE investigation as Wernbom et al178, and found that the torque decrements at 24
hours post-exercise were highly correlated with the cytoskeletal levels of HSP27 (r=0.87).
These findings may have implications for mechanisms of muscle damage, torque decrements
and low-frequency fatigue with BFRRE. A suggested scheme for some of the possible factors
behind the development of fatigue in BFRRE is depicted in Figure 2.
5. Methods to investigate muscle fibre activation and use
5.1. Glycogen depletion
Glycogen depletion has a central place in the history of the study of motor unit recruitment
and muscle fibre use. In the 1960s and 1970s, measurements of glycogen deposits in muscle
fibre sections provided direct evidence for the existence of different types of motor units in
both animals31,32,206 and humans37. Glycogen depletion was also used in many of the classic
studies on muscle fibre recruitment in various types of exercise in humans154,207-212 and
continues to be used up to the present day to map muscle fibre use with various exercise
protocols (e.g. Cumming et al21, Andersen & Gruschy-Knudsen213). In exercise studies,
glycogen content is usually determined semi-quantitatively with assessments of periodic acid

schiff (PAS) staining intensity on muscle fibre sections. Importantly, PAS staining intensity
on sections correlates very strongly (r=0.91-0.93) with direct measurements of glycogen

Accepted Article
concentration in the same muscle fibres210,214.
In contrast to endurance and intermittent exercise, relatively few studies had
examined glycogen depletion and repletion patterns in muscle fibres with resistance type
exercise up until the late 1990s215,216. Surprisingly, this is still largely true, especially
regarding the influence of resistance exercise variables on glycogen utilization and repletion
in the different muscle fibre types. Among the few studies on this subject, Robergs et al214
and Tesch et al216 demonstrated that glycogenolytic rates in mixed muscle fibres and in fast
fibres increased with increasing exercise intensity. This is supported by data from Söderlund
et al168, who showed that 50 Hz stimulation of the vastus lateralis resulted in a nearly doubled
rate of glycogenolysis in type II fibres compared with 20 Hz, which is consistent with the 50-
60 Hz required for maximal force in type II fibres. Current models of the regulation of
glycolysis during muscle contractions strongly suggest that both increased Ca2+
concentrations and elevated levels of ADP and AMP play important roles in activating
glycolysis217. More marked increases in Ca2+, ADP and AMP would thus appear to explain
the greater glycogenolytic rates in high-force contractions compared with low-force
contractions.
Robergs et al214 studied the effects of two resistance exercise protocols with
knee extensions at 70% and 35% of 1RM respectively, which were approximately matched
for total work (6 sets of 6 repetitions vs. 6 sets of 12-13 repetitions). Interestingly, although
the rate of glycogenolysis was greater for the higher load, the two protocols resulted in
similar degrees of glycogen depletion in both type I and type II fibres. Additionally, the data
suggested that glycogen utilization tended to be greater in the type II fibres, especially for the
high-load regime. Also of note, the rate of glycogen resynthesis during the first 2 hours post-

exercise was high particularly in the type II fibres. The higher rate of resynthesis in fast fibres
may in part have been due to glycogen synthesis from lactate218-220, which is considerably
Accepted Article
faster in type II fibres than type I fibres at least in rodents221 and which appears to take place
via a reversal of pyruvate kinase action218,219. As will be discussed later, an enhanced
glycogen repletion may also have influenced the results of studies on glycogen depletion with
BFRRE. However, Robergs et al214 did not differentiate between subtypes of type II fibres.
Tesch et al216 investigated three different resistance exercise protocols, each
consisting of 5 sets of 10 concentric-only repetitions for the knee extensors at 30%, 45% and
60% of 1RM respectively. Because they found very few type IIX fibres in their myosin
ATPase stained muscle sections, Tesch et al216 analyzed type IIAX and IIX fibres together.
With the two lower loads, only type I and type IIA fibres displayed reductions in glycogen,
but with 60% of 1RM, type IIAX+IIX fibres also showed glycogen loss. Interestingly,
Robergs et al214 reported no significant difference in type II fibre depletion between 70% and
35% of 1RM, and inspection of the absorbance data indeed suggests little difference: a ~52%
vs a 44% decrease in muscle glycogen levels in type II fibres with 70% of 1RM and 35% of
1RM, respectively. This suggests that type II fibres were recruited and used to almost the
same extent with the lower load protocol. In contrast, Tesch et al216 showed considerably
greater depletion in type IIA as well as type IIAX+IIX fibres with 60% of 1RM compared
with both 30% and 45% of 1RM.
Both Robergs et al214 and Tesch et al216 used subjects with resistance-training
experience, so a difference in training status seems unlikely to be an important factor behind
the divergent results. However, a greater number of repetitions was performed for the low-
load protocol in Robergs et al214 compared to Tesch et al216, ~77 vs 50 repetitions. An
important additional difference was that the repetitions were continuous concentric-eccentric
in the former study and concentric-only in the latter. As noted by Robergs et al214, an elevated

intramuscular pressure therefore remained throughout the duration of each repetition,
resulting in reduced blood flow, whereas the protocol of Tesch et al216 almost certainly
Accepted Article
allowed a higher muscle blood flow due to the muscle relaxation between each concentric
repetition (see Wernbom et al5 for further discussion of this point).
Taken together, the results of Tesch et al216 and Robergs et al214 demonstrate
that there is substantial type II fibre usage already at 30-35% of 1RM. However, the data in
Robergs et al214 suggest a relatively greater use of type II fibres with the low-load protocol in
their study, probably at least in part due to the maintained intramuscular restriction of blood
flow and resulting fatigue in type I fibres, which in turn may have caused compensatory
recruitment of more type II fibres, as discussed elsewhere in this review.
Robergs et al214 also showed linear decreases in glycogen levels in both type I
and type II fibres which were proportional to the total work performed during exercise. This
may be viewed as both a strength and a weakness. On one hand, a given pattern of glycogen
depletion may provide a rough estimate of how much a specific fibre type has been used in a
given exercise protocol. An important advantage is that full recovery of glycogen occurs
slowly (hours), thus biopsies do not have to be obtained immediately (within seconds) post-
exercise. On the other hand, because depletion is dependent on duration, it may require
several minutes before a measureable decrease is seen222,223, and as discussed elsewhere in
this review, the rate of glycogen depletion is usually much slower in type I than in type II
fibres with intermittent high-force contractions.
For example, Andersen & Gruschy-Knudsen213 found a much greater
percentage of glycogen depleted type IIX fibres compared to type I fibres after 340 maximal
concentric contractions (~70% vs 15%). Gollnick et al155 reported that repeated contractions
of the quadriceps at 20% of MVC or more sustained to fatigue resulted in glycogen depletion
of type II but not type I fibres. Similarly, Tupling et al224 reported non-significant decreases

in glycogen in type I fibres but substantial depletion in type II fibres with 5 second
intermittent contractions at 60% of MVC. The finding of seemingly very little type I fibre use
Accepted Article
at intensities 20% of MVC in the studies of Gollnick et al155 and Tupling et al224 is difficult
to explain but given that the contractions were performed many times and over periods of
many minutes, a considerable contribution from oxidative metabolism to the later bouts of
exercise is plausible, as observed by Ingemann-Hansen et al225. In contrast to these results
from repeated contraction studies, Tesch & Karlsson226 observed substantial and almost equal
lactate contents in type I and type II fibres after single contractions at 25%, 50% and 75% of
MVC maintained to fatigue. These and other investigations show that glycogen depletion
patterns need careful interpretation within the specific context that they occur.
Furthermore, it is known that muscle fibre type distribution can vary
significantly between the superficial and the deep parts as well as between the origin and the
insertion in human limb muscles such as the vastus lateralis and the biceps brachii227-229.
Interestingly, muscle fibre areas have also been reported to differ between the superficial and
deep layers in the vastus lateralis, with the mean fibre area being larger in the deep layers230.
It has been suggested that this is an indication of functional differences between the various
parts of the muscle230 (c.f. Section 5.3.1. for further discussion of such possible differences).
Accordingly, the possibility of differences in fibre activation patterns between different layers
in the same muscle must be kept in mind when choosing the sites of biopsy sampling and
when interpreting glycogen depletion patterns induced by various exercise protocols.
5.2. Phosphocreatine depletion
Pioneering studies on PCr depletion in human muscle fibres after acute exercise bouts were
published in the late 1960s and early 1970s231-233. Rehunen et al234 were the first to
investigate exercise-induced PCr changes in type I and II fibres separately, and seven years

later Tesch et al171 were the first to report on fibre-type specific changes in PCr with high-
force contractions. Tesch et al171 found reductions in PCr in type I and type II fibres to 41%
Accepted Article
and 31% of baseline levels immediately after 30 maximal concentric contractions at 180°/s,
thus providing clear evidence of both type I and type II fibre recruitment with isokinetic
resistance exercise. After 60 seconds of recovery, the levels were 69% and 49% of baseline
levels, indicating not only less decline in type I fibres but also a quicker recovery. It is
important to note that the concentric-only contraction were separated by 1.2 second rest
periods in the investigation of Tesch et al171, unlike with conventional resistance exercise,
where the concentric and eccentric phases follow each other with little or no relaxation in
between.
Since the study of Tesch et al171, surprisingly few investigations have looked at
PCr use in type I and II fibres during voluntary high-force contractions. A sensitive method
for determining muscle fibre use was developed by Beltman et al222, based on the ratio
between PCr and Cr levels. In a subsequent study, Beltman et al223 found that type I fibres
and some type IIA fibres had been recruited after 7 brief isometric contractions at 39% of
MVC, with further involvement of both type I and type IIA fibres after 7 contractions at 72%
of MVC. Only at 87% of MVC did type IIAX fibres display significantly lower PCr/Cr ratios
(type IIAX included fibres with 15-100% type IIX MHC). Employing this method, Beltman
et al99 also investigated muscle fibre use with maximal concentric and eccentric contractions
at 60°/s, as well as maximal isometric contractions, each performed for 10 repetitions. In this
study, the type IIAX included all fibres with 25-100% of type IIX MHC. Interestingly, while
all fibre types investigated showed decreased PCr/Cr ratios with all contraction types, the
shift was less in type IIAX fibres after eccentric contractions, indicating less recruitment of
these fibres during maximal eccentric exercise.

The paucity of research on fibre type use in resistance exercise as measured by
PCr depletion may in part be explained by the experimental difficulties associated with
Accepted Article
obtaining muscle biopsies quickly enough to ensure valid assessments of immediately post-
exercise PCr levels. For example, Beltman et al223 excluded muscle samples which had been
obtained ≥10 s post-exercise, which should be compared to the estimated half-time recovery
of 30 s for PCr levels. However, as indicated from the previous discussion on fibre type
differences in PCr recovery times, this half-time is almost certainly different between the
various fibre types and subtypes, with an approximate order from the longest to the shortest:
type IIX > type IIAX > type IIA > type I. Even so, the narrow time frame available for
muscle biopies for post-exercise PCr assessments means that the use of PCr depletion is
restricted to investigators who have developed great skill and specific routines for obtaining
muscle biopsies as quickly as possible (few seconds) following cesseation of exercise.
Furthermore, based on the finding that muscle PCr levels in muscle samples can
decrease with 13-19% with 1-4 weeks of storage even when the samples are stored at -70°C,
it has been recommended that PCr levels should be analyzed within 24 hours after the biopsy
sampling235. While it is well recognized that a delay between sampling and freezing of the
biopsy can affect muscle PCr values, the decreases with storage appear to be less known, and
many papers on PCr changes with exercise have not reported the time elapsed between
freezing and subsequent analyses of PCr levels. Both the delay between sampling and
freezing and between freezing and biochemical analyses could conceivably have affected the
results on resting PCr levels as well as changes resulting from exercise and recovery in many
of the papers in the literature reporting on these changes. Finally, it deserves to be noted that
measurements of PCr levels from muscle biopsy samples are subject to essentially the same
interpretive caveats as glycogen measurements with reference to possible differences in the
activation patterns between different regions of the same muscle.

5.3. Electromyography (EMG)
5.3.1. Surface EMG

Accepted Article
In exercise physiology, the type of EMG used is usually surface EMG with bipolar electrodes
applied on the skin over the working muscles. In surface EMG research, the amplitude is
often used to infer the level of ”neural drive” to the muscles, although the limitations of
surface EMG for this purpose have been known for several decades (reviewed by Duchateau
et al28, Farina et al236, and Vigotsky et al237). For example, Moritani et al238 cautioned that the
change in the surface EMG should not automatically be attributed to changes in either motor
unit recruitment or in firing rates as the EMG amplitude is further influenced by the
individual muscle fibre potential, degree of motor unit synchronization, and fatigue.
With the exception of sophisticated decomposition techniques235, it is probably
not possible to separate the contributions of recruitment of new motor units and increased
firing rates to increases in surface EMG amplitude during exercise, especially with dynamic
contractions. Even with advanced decomposition techniques, it can be very difficult to extract
neural strategies, as surface EMG provide only a limited representation of the active muscle
fibres and is biased towards the motor units with the largest surface action potentials, which
are large and superficial motor units236. Furthermore, surface EMG decomposition is possibly
sensitive to overlapping action potentials and phase cancellation117,240, although this has been
disputed241. In addition, decomposition techniques are still limited to mostly isometric
contractions and can not detect deep motor units236. For further discussion on these issues, the
reader is referred to the aforementioned papers, and to Farina et al242,243 and De Luca et al244.
A particularly serious criticism against using surface EMG to infer acute
changes in neural drive was raised by Dimitrova, Dimitrov and colleagues, who performed
simulation studies which suggested that peripheral changes in the muscle fibres themselves
due to fatigue may influence EMG amplitude to a much greater extent than changes in

”neural drive” 245,246. Specifically, lengthening of the intracellular action potential (IAP)
profile and the after-potential can cause increases in integrated EMG and EMG root mean
Accepted Article
square (RMS) amplitudes without alterations in neural input and recruitment of additional
motor units245,247. However, due to the lack of human data, Dimitrova & colleagues245-247
based their simulations of the effects of changes in the IAP profile primarily on data derived
from a study by Hanson & Person248 on frog muscle fibres investigated at room temperature.
Interestingly, Hanson249 noted that the after-potential in rat muscles was
considerably smaller than in the frog muscles studied earlier and that while the effects of
repetitive stimulation of rat extensor carpi radialis muscle on IAP parameters were of the
same kind as those in frog muscles, they were less pronounced. In rat soleus muscle fibres,
there were only very small changes249. In a subsequent study on both human and rat muscle
fibres, Hanson250 concluded that ”the intracellularly recorded potentials in human muscles
were not markedly changed during repetitive stimulation, while in the rat intercostal muscles
the same type of change was seen as earlier observed in fast limb muscle of rat and in frog
muscle fibres. Thus the human muscle fibres resemble, in this respect as well, the slow soleus
muscle of the rat”.
Because human muscle fibres apparently show much less fatigue-induced
changes in IAP than both frog and rat muscle fibres (except perhaps for rat soleus fibres), it is
possible that the relative degree of lengthening of the IAP profile was exagerrated in the
simulation studies of Dimitrova and colleagues245-247. Thus, the surface EMG amplitude
recorded from human skeletal muscle during fatiguing exercise may not be as affected by
changes in IAP profile as suggested in these simulation studies245-246, although the degree of
influence under hypoxic and ischemic conditions awaits further studies. In contrast, other
simulation studies suggest that amplitude cancellation of single motor unit action potentials
(MUAPs) can impact negatively on surface EMG amplitude, an effect which may be more

marked during fatiguing contractions251, although a later simulation study from the same
group reported negligible effects of fatigue on cancellation252. Surface EMG could thus still
Accepted Article
provide a useful, if crude, global measure of muscle activation, especially if EMG is properly
normalized30,253,254. However, recent results strongly indicate that the size of the MUAPs is
even more important than ”neural drive” for summated EMG amplitude and that normalized
EMG amplitude analysis may not be appropriate for inferring differences in ”neural drive”
between muscles255.
Nevertheless, EMG recording may with these limitations in mind provide clues
regarding possible differences in activation patterns in a single muscle. For example, Christie
et al67 showed weak correlations between motor unit firing rates and surface EMG RMS
amplitude over the entire range of isometric force development in the human biceps brachii.
Instead, they suggested that the overall trend of a slow initial increase followed by more rapid
increases in RMS amplitude at higher forces fitted with the later recruitment of superficial
motor units. A similar explanation was proposed by Moritani & Muro256, who also studied
the biceps brachii and who observed disproportionate increases in EMG RMS amplitude over
force output in the higher part of the force range investigated (0-80% of MVC). This scheme
is consistent with findings of Clamann257, who reported that motor units deep in the biceps
muscle had the lowest recruitment thresholds, and that those closest to the surface were the
last to be recruited. Combined, the results from these investigations on EMG in the human
biceps brachii strongly suggest that late recruited large superficial units may greatly
contribute to the overall surface EMG amplitude both by virtue of their size and by being
located nearer to the recording electrodes.
The deep-to-superficial recruitment pattern may also apply to the quadriceps.
Accordingly, it has been established that superficial layers in the vastus lateralis contain both
larger motor units258 and a greater percentage of type II muscle fibres229 than deeper layers,

and results from other studies are also consistent with a generally greater prevalence of type
II fibres in the superficial layers compared with the deeper layers in this muscle227,228,259. The
Accepted Article
vastus medialis muscle appears to have an even greater difference in fibre type distributions
between superficial and deep layers, with type II fibre dominance in the former and type I
dominance in the latter227. Furthermore, it has been reported that the vastus lateralis has a
greater percentage of type II fibres than the vastus intermedius259 and that while the rectus
femoris has a type II fibre dominance in all its parts, the superficial part has the greatest
percentage of type II fibres227. Interestingly, data from scanning studies indicate that the
vastus intermedius is the primarily recruited muscle in dynamic knee extension exercise at
low loads260-262, with all quadriceps muscle components being recruited at higher
loads260,261,263. Situated deep to the vastus lateralis and the rectus femoris, the vastus
intermedius is not accessible to surface EMG except at the most distal part264.
Collectively these findings suggest that a deep-to-superficial activation pattern
may to some extent exist not only within but also between muscle bellies in the quadriceps.
Thus, it is not surprising that numerous studies have demonstrated a non-linear relationship
between isometric knee extension force and surface EMG amplitude for the quadriceps
muscle264-269 although this is not always the case, see Alkner et al268 for discussion. In the
studies which have shown a non-linear relationship, the EMG-force curve typically seems to
be marked by a somewhat flatter slope at lower force levels accompanied by a steeper slope
at high to maximum force levels (see e.g. Thorstensson et al265, Rabita et al269, Watanabe &
Akima264).
Based on the findings of large superficial motor units in the vastus lateralis258
and that recruitment in this muscle may occur up to 95% of MVC69, it seems reasonable to
suggest that the relatively steep increase in EMG RMS activity seen between high and
maximal force development in the quadriceps may at least in part be caused by the

recruitment of very high-threshold superficial motor units containing type II fibres, possibly
type IIX. However, as discussed previously, Pucci et al102 showed that the discharge
Accepted Article
frequency in the vastus lateralis increased steeply from ~20 Hz to ~42 Hz when the torque
increased from 75% to 100% of MVC. Therefore, we suggest that higher firing rates may
also contribute to the steep increase in EMG RMS often seen between high and maximal
force development in the human quadriceps. Other factors such as motor unit
synchronization, amplitude cancellation and lengthening of the intracellular action potential
profile are also likely to affect the EMG amplitude, but their relative importance remains to
be determined.
An often overlooked phenomenon in conventional surface EMG is that EMG
amplitude during concentric contractions can exceed that observed in isometric
contractions270-273, especially with maximal efforts at fast contraction velocities270 and in the
initial phases of concentric contractions during conventional heavy resistance exercise272,273.
In the studies of Amiridis et al270 and Beutler et al273, EMG RMS activity recorded during
concentric knee extensor contractions temporarily reached up to ~1.6-1.8 times higher levels
than during isometric MVCs. It is important to note that in both these studies, EMG was
normalized to a 1 second period during the MVC which was either centered around the
greatest activation or after the force had reached a steady plateau. In contrast, the peak
dynamic EMG values were measured during time periods on the order of a few tenths of a
second. It is therefore likely that spike-frequency adaptation and consequent reductions in
maximal firing rates (Section 3.6.) affected the sEMG amplitude negatively to a much greater
extent in the MVCs than in the fast and accelerating concentric contractions in the studies of
Amiridis et al270 and Beutler et al273.
Initial spike-frequency adaptation, together with shorter time windows for
analysis of the EMG signal, probably also at least in part explains the greater EMG

amplitudes during fast vs slow concentric contractions which have been reported in several
studies270,274,275. Intriguingly though, EMG in the quadriceps can increase with ~20-30%
Accepted Article
during sets of repeated maximal-effort high-velocity pure concentric repetitions, peaking
after about 8-12 consecutive concentric contractions276. Given that high-velocity concentric
knee extensions already typically display the greatest EMG values274,275, it is highly unlikely
that this increase reflects improved muscle activation during the set. In this context, it is
interesting that changes in M-wave amplitude have been correlated with changes in EMG102.
Therefore, changes in muscle fibre excitability may at least in part explain the results of
Tesch et al276. In addition, it has been suggested that acute increases in M-wave and EMG
amplitudes may be misleading since intracellular muscle fibre action potential and muscle
fibre excitability can decrease at the same time189,236.
Taken together, it is apparent that there are many pitfalls in interpreting EMG
amplitude results247. Accordingly, it is suggested that possible changes in motor unit firing
rates and muscle fibre excitability (including transmission failure) must be taken into account
when interpreting EMG results from acute experiments and training interventions.
Furthermore, as surface EMG recording may not detect deep motor units236, there is a
theoretical possibility that some motor units might undergo drastic decreases in discharge
frequencies and/or in muscle fibre excitability and that some fibres may even drop out during
ongoing exercise without these events being detected.
Intramuscular electrodes are preferable over surface EMG to study the behaviour of single
motor units28, and many of the classic studies on motor unit recruitment have employed
intramuscular electrodes primarily in the forms of concentric needles and different variants of
fine wire electrodes28,277. Intramuscular electrodes have also been of central importance in the

efforts to determine the functions of the various muscles in the human body, as documented
in classic textbooks278. Intramuscular EMG can help distinguish between low-threshold and
Accepted Article
high-theshold motor units based on recruitment order and spike amplitudes17,279 and is
probably less affected by intracellular changes in muscle fibres than surface EMG245.
However, intramuscular electrodes do not necessarily reveal with certainty the
types of muscle fibres that are active. Furthermore, it is very difficult to use intramuscular
electrodes with dynamic movements, as they easily move out of position73. Needle and fine-
wire electrode EMG also have the obvious disadvantage of being invasive methods, and can
typically only track a few motor units at any given time, hence raising concerns about general
representivity. For example, the motor unit action potential amplitude detected with
concentric needle electrodes depends on muscle fibres located within a radius of 0.5 mm
from the active surface of the electrode280. Unless several electrodes are used (e.g. Moritani et
al279), whether the motor units recordings are representative must therefore be questioned.
5.4. 31P-MRS
Phosphorus-31 magnetic resonance spectroscopy (31P-MRS) is a non-invasive method which
has been used to study muscle metabolism in humans since the early 1980s281. Direct
measurements of pH and PCr in human muscle biopsies have confirmed the validity of 31P-
MRS for measuring exercise-induced changes in pH and PCr282, although other biopsy
studies have shown that decreases in pH and ATP may be somewhat overestimated with 31P-
MRS283. 31P-MRS has also been used for assessing muscle fibre recruitment. The peak of Pi
shifts during exercise according to changes in muscle pH, and the occurrence of ”split” Pi
peaks in the 31P spectra during exercise has been used to infer that both type I and type II
fibres have been recruited284. Specifically, the low pH peak of Pi is presumably caused by the

lower pH of type II fibres during exercise due to the greater dependence on glycolysis in
these fibres, whereas the high pH peak primarily reflects type I fibres284--287.
Accepted Article
Indeed, muscle biopsy studies have revealed that type II fibres have markedly
higher glycolytic rates during exercise than type I fibres in humans52,53,288. Furthermore,
several studies have shown a much faster disappearance of the high pH Pi peak vs the low pH
Pi during recovery after fatiguing exercise, which has been interpreted as reflecting the much
higher oxidative capacity of type I fibres compared with type II fibres, especially type IIX284-
286,289
. This interpretation is consistent with the observations that PCr recovery is dependent
on the magnitude of oxygen supply204,290,291, and that the higher the oxidative capacity of the
muscle fibre, the faster the recovery of PCr after exercise198.
Interestingly, studies have reported up to three Pi peaks in fatiguing exercise
during free flow conditions284,289, but in four out of seven subjects in the study of
Vandenborne et al284, only two peaks were observed. In Mizuno et al289, three Pi peaks were
observed only in 2 out of 14 subjects. The two peaks reported by Vandenborne et al284 were
interpreted as slow oxidative fibres (i.e. type I) and fast oxidative glycolytic fibres (type IIA)
respectively in some subjects, and fast oxidative glycolytic fibres and fast glycolytic fibres
(type IIX) in others. The recovery of the intermediate pH Pi peak was also much faster than
the low pH peak. This is in agreement with the higher oxidative capacity of the type IIA
fibres compared with type IIX fibres51,199-202, and also with biopsy studies on PCr content and
recovery after exhaustive sprint cycle exercise (discussed in Section 4.7.), where PCr
recovery is quicker in both type I and pure type IIA fibres than in hybrid type IIAX and
almost pure IIX fibres49.
Even so, criticism was raised already in early studies292-294 against data from
investigations where the signal had not been properly localized to one muscle belly.
Subsequent studies with localized 31P-MRS provided evidence for the split Pi being produced

by the different fibre types within the same muscle as opposed to pH differences between
different muscle bellies284,295. Experiments with relatively selective neuromuscular blockade

Accepted Article
of type I and type II fibres during exercise provided additional strong support for fibre type
heterogenity as the explanation for the Pi splits296,297. In these studies, blockade of either fast
fibres or slow fibres during exercise made the Pi split disappear compared to control exercise
without blockade296,297. Nevertheless, the use of surface coils larger than 2.5–2.9 cm on the
forearm is likely to sample signals from more than one muscle293. Because Mizuno et
al289,296,297 used a 4-cm coil and measured signals from multiple muscles, these elegant
studies are not definitive proof against the view that split Pi peaks can originate from different
muscle bellies, as opposed to (or in addition to) reflecting the recruitment of different muscle
fibre types in a single muscle. This notion is underscored by the results from T2 weighted
MRI studies on the plantar flexors, which show non-uniform increases in activity among the
individual muscles with exercise, demonstrating the gastrocnemius medialis to be the most
active agonist in plantar flexion exercise with the knee in an extended position298,299.
Indeed, two recent studies which employed precise localization methods to the
single muscle belly of gastrocnemius medialis demonstrated split Pi with non-localized 31P-
MRS but not with localized 31P-MRS300,301, which again has raised the question whether the
occurrence of split Pi peaks in single muscles is a fact or an artifact. Even among the studies
which have reported split Pi peaks, the occurrence has been variable. Vandenborne et al295
used localized 31P-MRS on the gastrocnemius medialis and lateralis and the soleus, and
reported that with steady state low frequency exercise, 5 out of 18 investigated muscles
showed a broad peak indicating 2 or 3 components, and in 2 of these muscles the broad Pi
peak clearly resolved into two distinct Pi peaks. With maximal effort high-frequency
exercise, the Pi line width was wider than the PCr width in all muscles, indicating more than
one component (and thus fibre type) in the Pi peak, but no split peaks were observed. This is

in clear contrast to their earlier study with the same maximal protocol but with a less precise
localization technique (87-90% of the signal from one muscle), in which 7 out 7 subjects
Accepted Article
displayed split Pi peaks284. Collectively, these results clearly indicate that signal
contamination from less active muscles may play a significant role in the observation of split
Pi peaks.
As an additional factor, the pattern of fibre type distribution also appears to play
a significant role in whether split Pi peaks are observed or not. Thus, Mizuno et al289 reported
that only subjects with a relatively even distribution of type I and type II muscle fibres
displayed split Pi peaks during exercise, whereas subjects with a predominance of type I or
type II fibres showed a single peak, at high and low pH respectively. Although this finding
should be viewed with caution due to the non-localized spectra, it suggests that a relative
abundance of both type I and II fibres is necessary to cause the ”compartments” in the spectra
that form the basis for split Pi peaks. Support for this possibility may also be found in the
results of a study by Houtman et al287, who observed a pH gradient during sustained isometric
contraction between the medal and lateral parts of tibialis anterior, and who reported double
Pi peaks in all their subjects during 30% of MVC sustained to fatigue. In addition, the failure
of some studies to find split Pi peaks with localized spectra of one muscle may be due to the
exercise not being strenuous enough to result in the recruitment of fibre populations
sufficiently different from each to cause Pi splitting.
A probably more consistent indication of recruitment of both type I and II fibres
during exercise is increased line width of the Pi peak, which is based on the same muscle
fibre pH heterogenity as split Pi peaks. Rossiter et al302 found that while splitting into low and
high pH peaks occurred in 5 out 11 subjects during exercise, all of them showed increases in
the Pi peak line width. Rossiter et al302 further noted that ”simple splitting of the Pi peak into
two separate, distinct regions may be an oversimplification. Although the Pi peak may be

adequately modeled by either one or two compartments, a broadening of the Pi peak in all
cases may reflect an overlapping of a number of muscle “compartments” distinguished by a

Accepted Article
range of pH values”. The difficulties with finding split Pi peaks in a single muscle are not
surprising in light of the evidence that in many muscles, there is a continuum of subtypes
within type I and type II muscle fibres, as discussed previously. Again, it should be noted that
the difference between type II and type I fibres in glycolytic rates and in accumulated lactate
levels can become substantially smaller during certain types of anaerobic exercise including
ischemic contractions.
This possibility is underscored by the findings of Tesch & Karlsson226,303, who
reported nearly as high lactate levels in type I fibres as in type II fibres in untrained subjects
after sustained contractions at 50% of MVC to exhaustion (up to 19.8-28.7 mmoles per kg
wet weight vs. 22.1-29.8 mmoles per kg ww). As previously noted, 50% of MVC is sufficient
to cause near-complete intramuscular occlusion of the blood flow in the human
quadriceps183,184. An increase to 22 mmol lactate per kg ww corresponds to a drop in pH of
about 0.5282,290. Interestingly, ~23 mmol lactate per kg ww has been reported in muscle fibres
after low-load BFRRE performed to failure225. Therefore, a significant shift in the Pi peak to
lower pH values may occur not only in type II fibres but in type I fibres as well during very
strenuous exercise, which could introduce methodological difficulties in separating the Pi
peaks. In line with this scenario, a late increased rate of deline in the pH of the high pH Pi
peak has been observed in the tibialis anterior muscle during ankle dorsiflexion isometric
contractions at 30% of MVC sustained to fatigue287,304.
Intriguingly, Houtman et al304 reported three phases of PCr usage during their
fatigue protocols. The first phase was characterized by a rapid decline in PCr, which was then
followed by a second phase of slowed PCr consumption, presumably due to increased
glycolysis and oxidative metabolism. During this phase, two distinct Pi peaks appeared which

were attributed to the contributions of type I and type II motor units. During the third and
final phase, a steep rate of decline in the pH of the high pH Pi peak was seen, and a merging
Accepted Article
of the two Pi peaks occurred in several subjects. Figure 4 in Houtman et al304 suggests that in
other subjects, the high pH Pi peak disappeared without any markedly increased rate of
decline in pH. This might be explained by trapping of Pi in mitochondria during ischemic
exercise, resulting in loss of the Pi signal (discussed in Houtman et al287). Notably, splitting
followed by merging of the two Pi peaks as well as disappearance of the high Pi peak have
been observed also during fatiguing dynamic exercise305.
In summary, it appears that split Pi peaks may reflect recruitment of both type I
and type II muscle fibres. However, the presence of inconsistent findings and the many
factors which can affect the occurrence and disappearance of the Pi peaks raise questions as
to whether determination of Pi splitting by 31P-MRS is sufficient as a stand-alone method for
assessing relative degrees of recruitment of the various fibre types.
5.5. Studies on combined EMG and 31P-MRS
MRS in conjunction with EMG is a useful approach in broadening the understanding of the
mechanisms of fatigue in health and disease306. A number of investigations have combinded
EMG and 31P-MRS to study the relationships between the myoelectrical and the biochemical
processes which underpin neuromuscular fatigue and recovery289,307-310. The studies of
Vestergaard-Poulsen et al308,309 are of particular interest with regard to muscle fibre
recruitment during fatiguing low-force exercise. In these studies, the subjects performed
isometric ankle dorsiflexion contractions at 10% and 30% of MVC to fatigue, and EMG
variables and metabolites in the tibialis anterior were monitored simultaneously. In both
studies308,309, it was reported that EMG RMS amplitude remained relatively constant or
increased only slightly during the low-force contractions until the pH value dropped below

6.8 and the PCr/ Pi ratio was between 1.0–2.0. Below these breakpoints, the EMG RMS
amplitude increased and in the latter study, a split Pi peak appeared synchronized with the
Accepted Article
increase in EMG. With further drops in pH and the PCr/ Pi ratio, EMG increased rapidly in a
hyperbolic manner to the point of exhaustion.
The findings of Vestergaard-Poulsen et al308,309 are very similar to those of
Houtman et al304, who studied tibialis anterior with an almost identical exercise protocol. In
Houtman et al304, the split Pi peaks appeared at around a pH of 6.8 in most of the subjects.
Houtman et al311 followed up their MRS study with an EMG study using the same exercise
protocol. An essentially hyperbolic EMG RMS increase was seen in several of their subjects
as they neared task failure, along with an increase and then a decrease in muscle fibre
conduction velocity. This was interpreted as recruitment of fresh type II muscle fibres due to
fatigue in type I fibres, with the type II fibres then showing fatigue toward the end of the
exercise. Importantly, this interpretation was supported by several lines of evidence.
These studies demonstrate some of the values of combining EMG and 31P-MRS
to gain better insight into the mechanisms of fatigue and the patterns of recruitment of motor
units during strenuous exercise. However, combining these two technologies is not without
problems as each of them might add unwanted noise to the recordings with the other312.
6. Studies on muscle fibre activation and use in BFRRE
6.1. Studies on glycogen depletion in BFRRE
At least two studies have shown type II fibre use in low-load BFR training via analyses of
glycogen depletion in muscle fibres from biopsies21,225. The study of Ingemann-Hansen et
al225 is arguably the first investigation of muscle fibre use in low-load BFRRE measured at
the cellular level. The exercise model was knee extensions performed in a prone position,

with intermittent working sets separated by 10 minute rest periods with free blood flow.
Muscle biopises were taken from the vastus lateralis. The combination of cuff width (7 cm)
Accepted Article
and pressure (300 mm Hg) used by Ingemann-Hansen et al225 typically results in complete
occlusion of thigh blood flow in most subjects in a supine position (Wernbom, unpublished
observations, see also Crenshaw et al313).
Unfortunately, these authors reported the load in Watts (14.7) and kilograms
(10), and not in % of MVC or 1RM. Based on the cadence (20 repetitions per minute) and
endurance time, the number of repetitions completed by their subjects in the first set of
exercise was around 55. From this and from the absolute load used (10 kg), as well as data
from our own acute experiments on dynamic knee extensions during varying degrees of
BFR19,178,314, it can be estimated that the load in the study of Ingemann-Hansen et al225 was
on the order of ~15-20% of 1RM. The subjects performed 10-16 sets of exercise until
exhaustion, and biopsies were taken immediately after set 2, 5, 8 and the last set (while the
cuff was still inflated), and 30 minutes after the last set.
The glycogen depletion data from this study indicated recruitment of both type I
and type II fibres, although the picture was complex. After the last exercise set, the glycogen
content was reduced by 55% and 39% in the type I and type II fibres, respectively.
Interestingly, the reduction was not homogenous in the type II fibres, with some fibres
showing only minor reductions (~10%) while others displayed more marked depletion
(~50%) and still others were in between these figures. Strikingly, the most depleted type II
fibres were those that had the lowest oxidative capacity, as judged by low staining for
NADH-tetrazolium reductase (NADH-TR). Conversely, the type II fibres with the highest
NADH-TR staining also had the highest glycogen content, i.e. they seemed to have been used
the least. This is a puzzling finding, given that the type II fibres which have the highest
oxidative capacity and NADH-TR staining are typically type IIA fibres and vice versa for the

type IIX fibres (type IIB in older literature)199-201,315. In other words, the fibres which were
presumably type IIX seemed to have been used to a much greater extent than those which
Accepted Article
presumably were type IIA. At a first glance, this could suggest an altered order of motor unit
recruitment (i.e., type I > type IIX > type IIA).
In order to make sense of these findings, a brief discussion of subtypes of type
II fibres is warranted. As noted above, type IIA generally have a greater oxidative capacity
than type IIX. Type IIAX, the intermediate form between type IIA and IIX, appears to be
close to IIA in oxidative capacity as judged by mitochondrial content and succinate
dehydrogenase activity200,315, but may have been interpreted as type IIX (previously called
IIB in humans) in older ATPase-based classification systems due to its intermediate to dark
myofibrillar ATPase stain at pH 4.6 (for discussion and pictures, see Billeter et al317, Staron
et al200; Sant’Ana Pereira et al40,48 and Staron44). And as discussed in Section 4.1., some of
the fibres which are classified as type IIX fibres by ATPase histochemistry often contain
significant amounts of type IIA myosin in addition to type IIX, resulting in an
underestimation of hybrid IIAX fibres and an overestimation of type IIX fibres.
Taken together, these findings may in part explain why some researchers did
not find a perfect correspondence between type IIA and type IIX staining on one hand and
NADH-TR (or lack of thereof) staining on the other199. In addition, performing histochemical
staining of myosin ATPase only at a single pH of 10.3-10.4 as done in Ingemann-Hansen et
al225 does not allow for separation between the subtypes of type II fibres, including the
relatively rare and higly oxidative type IIC and type IIAC fibres in addition to type IIA, IIAX
and IIX41,44. Nevertheless, type II fibres which stain clearly positive for NADH-TR are for
the most part type IIA fibres, while fibres that are only lightly stained are type
IIX199,202,315,316, with IIAX hybrids probably being closer to IIA than more pure IIX fibres on
NADH-TR stained sections because of their higher mitochondrial content.

Based on the above considerations we can now attempt to interpret the puzzling
findings of Ingemann-Hansen et al225. As judged by the glycogen depletion, it appears that

Accepted Article
the type I, type IIA and IIAX/IIX muscle fibres were all recruited by the BFRRE performed
in their study. But due to lower oxidative capacities in type IIX, these fibres were more
dependent on anaerobic glyogenolysis than the type IIA and perhaps also type IIAX fibres.
The type IIA fibres may have used oxidative phosphorylation of both glycogen and lipids to a
significant extent, since the lipid stores are about twice as large as in type IIX fibres200.
Interestingly, Ingemann-Hansen et al225 suggested that aerobic metabolism played a greater
role during the last bouts than during the first ones, and that this could have been facilitated
by greater oxygen stores, caused by a greater number of open capillaries. Such oxygen stores
would be used to a certain extent even though the blood flow was probably nearly completely
occluded during each bout.
The finding of greater glycogen depletion in both type IIX and type I fibres than
in type IIA fibres is not without precedent. Fridén et al320 reported that repeated 25 second
sprints on a treadmill caused glycogen depletion in type I and type IIX fibres, but not in type
IIA fibres in sprinters. It is of note that sprinting, like ischemic exercise, can drastically
increase the rate of glycogenolysis in type I fibres52,53. Assuming that the size principle holds
for both these modes of exercise, it is still difficult to explain why the presumably heavily
recruited type IIA fibres should show less glycogen depletion than both type I and IIX fibres.
However, it is known that type IIA fibres have a lower tension cost than type IIAX fibres321,
and thus probably also compared with type IIX. As noted earlier, type II fibres have been
shown to replete glycogen faster than type I fibres214,322. In an investigation on 2.5 minute
cycling at 130% of VO2peak followed by 30-s all-out exercise, glycogen levels were
essentially back to baseline already within 45-75 minutes of passive rest321.

It can therefore be speculated that due to decreased anaerobic glycolysis and
increased oxidative phosphorylation during later bouts of exercise, and perhaps also a faster
Accepted Article
resynthesis of glycogen during the several 10-minute rest periods, the type IIA fibres were
better able to maintain glycogen levels than both type I and type IIX fibres during the
intermittent bouts of ischemic exercise. Importantly, the lactate data reported by Ingemann-
Hansen et al225 from the first 1-2 sets of exercise also provide evidence of recruitment of both
type I and type II fibres types. The ~23 mmol lactate per kg ww in mixed muscle samples
reported after bout 2 is only possible if both fibre types would have been extensively used.
In the only other study to date on glyogen depletion with BFRRE, Cumming et
al21 showed lowered glycogen levels in both type I and type II fibres from the vastus lateralis
one hour after an acute bout of BFRRE. The model training used by Cumming et al21
consisted of 5 sets to failure at 30% of 1RM in the dynamic knee extension exercise, with 45
seconds between sets. The occlusion was achieved with a 135 mm cuff inflated to 90-100 mm
Hg, resulting in about 50-60% reduction of blood flow during rest324. Due to the partial
instead of complete occlusion, and to the short inter-set rest periods, the training model of
Cumming et al21 is therefore more representative of the type of BFRRE that has been
employed in most studies than that of Ingemann-Hansen et al225. Interestingly, although both
fibre types diplayed reduced glycogen at 1 hour post-exercise, the glycogen depletion was
much more marked in the type I fibres, suggesting greater recruitment of these fibres with
BFRRE.
However, the degree of glycogen depletion may reflect not only the intensity
but also the duration or total work, as discussed previously. It is thus possible that type II
fibres were also highly recruited in the study of Cumming et al21, but that the duration of
work was not sufficient to result in a marked glycogen depletion in these fibres, as the type II
fibres would presumably be active mainly during the repetitions with high levels of muscle

activity. After a certain number of repetitions and sets with high activation, the type II fibres
may have dropped out and/or developed less and less force due to fatigue, which would then
Accepted Article
lead to decreased glycogenolytic rates, as shown by Hultman & Sjöholm166. Due to the short
inter-set rest periods (45 seconds) combined with partial occlusion, it is possible that type I
fibres had a relatively faster force recovery between each set in the study of Cumming et al21,
as discussed in Section 4.3. In addition, there may well have been a faster glycogen repletion
rate in the type II fibres after exercise, as noted by Robergs et al214, who demonstrated that
glycogen resynthesis was twice as fast over the first 2 hours post-exercise in type II versus
type I fibres after both low-load (35% of 1RM) and high-load resistance exercise (70% of
1RM) matched for work.
Accordingly, the involvement of type II fibres in BFRRE may have been
underestimated in the study of Cumming et al21. However, an argument against the possibility
of a very fast repletion in type II fibres in this study is that glycogen content continued to be
lower than observed at baseline in both fibre types at 24 hours post-exercise in the BFRRE
leg, although the PAS staining showed a greater decrease in type I fibres over type II fibres at
this time point as well as at 48 hours post. A prolonged depletion is consistent with some
degree of muscle fibre damage and/or remodelling, as has been observed after eccentric
exercise318, and the HSP response reported by Cumming et al21 further supports this notion.
As damage and/or structural remodelling to some degree seemed to have impeded the post-
exercise repletion of glycogen in both fibre types, a greater use of type I fibres during the
acute BFRRE bout appears to be the most plausible explanation for the greater depletion at 1
hour post-exercise in the type I fibres reported by Cumming et al21.

6.2. Studies on phosphocreatine depletion to assess muscle fibre use in BFRRE
Two studies to date have analyzed phosphocreatine levels in biopsies before and after low-
Accepted Article
load BFRRE. In addition to glycogen and lactate, Ingemann-Hansen et al225 also measured
PCr in mixed muscle fibres, and biopsy samples obtained after sets 2-8 showed marked
depletion of PCr, down to about 21% of the baseline level. This 80% reduction exceeds the
61-74% decreases in PCr previously reported in mixed human skeletal (VL) muscle after 12
electrically stimulated high-force contractions at 20 Hz and 50 Hz during concurrent
occlusion167, and approaches the values reported by Söderlund & Hultman169 after 52
ischemic contractions at 20 Hz and by Greenhaff et al52 after 20 ischemic contractions at 50
Hz (both down to ~6-7% of resting values). It has been shown that baseline levels of PCr are
only slightly higher in type II fibres49,52,168. In light of these observations, and given the low-
to-extremely-low post-exercise PCr levels in both fibre types reported by Söderlund et al168
and Greenhaff et al52, there must have been an extensive recruitment of not only type I fibres
but also type II fibres in the ischemic/occlusion exercise study by Ingemann-Hansen et al225,
in line with the glycogen depletion and lactate data discussed above.
Krustrup and coworkers20 investigated the effect of 90 seconds of cyclic knee
extensions with concentric-only contractions, with and without essentially complete
occlusion (300 mm Hg). The knee extension exercise model employed by Krustrup et al20
was the same as that of Andersen & Saltin326, who developed an ergometer for pure
concentric knee extensions with a range of motion between 90 and 160 degrees (180 degrees
= full extension). The load was 29 W and the cadence was 60 extensions per minute,
corresponding to an average joint angular velocity of ~140 degrees/s. With a work period of
90 s, the total number of repetitions thus was around 90. The load in terms of MVC or 1RM
was not reported. Given that the cadence was 3 times faster than in Ingemann-Hansen et al225
with a similar range of motion, and that the work output was 2 times greater, the approximate

load in terms of % of 1RM was probably even lower in Krustrup et al20 than the ~15-20% of
1RM we estimate for Ingemann-Hansen et al225. The fact that the subjects were able to
Accepted Article
complete 90 repetitions at a constant work rate despite 300 mm Hg of occlusion pressure is
also indicative of a work load below 20% of 1RM.
Summarizing their findings, Krustrup et al20 reported that when blood flow was
occluded, fibre recruitment was different from the free flow exercise at the same load as
indicated by lower average PCr concentrations for both type I and type II fibres immediately
after exercise with occlusion, with essentially all fibres having PCr values lower than resting
levels. In contrast, only 2 out of the 5 subjects showed drops in PCr levels in some of the type
II fibres in low-load exercise without occlusion. These results clearly demonstrate that
fatiguing low-load exercise with complete occlusion recruits both type I and type II fibres.
However, the data from Krustrup et al20 also suggest that up to 11% of the type II fibres did
not show evident signs of usage (i.e. no signs of reduced PCr), in contrast to 0% for higher
load exercise (65 W at 60 RPM) performed without occlusion. The percentages of type IIA
and type IIX fibres were reported to be 23 and 20%, respectively.
Thus it appears that during occlusion exercise all three major fibre types were
recruited, including type IIX, although a few type II fibres (10%) showed no signs of use.
Consequently, it may be speculated that the occlusion exercise did not result in sufficient
degree of fatigue to recruit the last remaining type II fibres (possibly of the type IIX type),
but that with a slightly longer duration and continued effort, these would likely had been
recruited as well.

6.3. Summary of glycogen and phosphocreatine depletion in BFRRE
In summary, at least three different studies have provided in essence direct evidence of both
Accepted Article
type I and type II use with low-load BFRRE during varying degrees of occlusion. The results
of two of these studies (Ingemann-Hansen et al225, Krustrup et al20) also strongly suggest that
all fibre types and subtypes can be recruited, from type I to type IIX, although this awaits
definitive confirmation. However, in these studies, the occlusion was complete or at least
severe (300 mm Hg), and as argued previously, only the study of Cumming et al21 used a
partial occlusion training model similar to those typically used in current BFRRE training
studies. As previously discussed however, due to the 1-hour delay between the end of the
BFRRE exercise bout and the biopsy procedure in their study, it is possible that some
glycogen resynthesis occurred especially in type II fibres in the study of Cumming et al21.
It is clear that there is a need for more studies on the patterns and extent of
myofibre recruitment in relation to fibre type composition during BFRRE. PCr depletion
studies may at a first glance seem a better means to assess muscle fibre recruitment than
glycogen depletion studies, as already a few muscle contractions are sufficient to cause some
decrease in PCr99,222,223. However, PCr measurements necessitate that the muscle biopsies are
taken immediately after the exercise has stopped. Glycogen depletion can also provide
important information on muscle fibre use in BFRRE provided that the exercise intensity and
volume are sufficient (e.g. a total of 50-75 repetitions at 20-30% of 1RM) and that biopsies
are taken within minutes after exercise before any significant glycogen repletion has taken
place. Thus, assessments of glycogen depletion have an advantage over PCr depletion with
regard to the allowed delay in muscle biopsy sampling (a few minutes vs a few seconds).
Finally, with the limitations discussed previously in mind, glycogen depletion may also
provide semi-quantitative information on the contributions of the different muscle fibre types
to the total work performed in various BFRRE protocols.

6.4. EMG studies on BFRRE
Accepted Article
6.4.1. Surface EMG
A number of studies have investigated surface EMG responses to various BFRRE protocols
with different exercises, loads, set & repetition configurations and BFR pressures. As surface
EMG is an indirect method of assessesing muscle fibre use253, it is beyond the scope of this
review to list and discuss each of these studies in detail. However, some of the more
influential studies as well as some overlooked pioneering investigations will be discussed in
the following. Studies that contain data which potentially add to the overall picture of muscle
fibre activation patterns and use in BFRRE will also be discussed below.
Person & Golubovich327 appear to have been the first investigators to use
surface EMG to measure muscle activity during low-load ischemic contractions. Their
subjects performed sustained isometric contractions with their elbow flexors to fatigue with a
constant weight of 5 kg (women) or 7 kg (men) with and without vascular occlusion. In the
BFR condition, a pressure cuff was wrapped around the proximal upper arm and inflated to
30-40 mm Hg above the systolic pressure. Without occlusion, the average endurance time
was 9 minutes, which was reduced to 3.5 minutes with cuff occlusion. At task failure, the
integrated EMG amplitude increased to 185% of the initial value without occlusion and to
307% of this value with occlusion. Person & Golubovich327 concluded that artificial ischemia
accelerated and intensified the characteristic increase in EMG amplitude and decrease in
frequency during fatigue. The decline in EMG frequency discussed by Person &
Golubovich327 probably refers to EMG mean power frequency (MPF), which is known to
decrease during fatigue328. The relative load was not specified, but comparisons with an
investigation by Bonde-Peterson et al140 on the impact of occlusion on endurance time at

different loads suggest that the relative load used by Person & Golubovich327 must have been
about 20% of MVC.

Accepted Article
Myers & Sullivan329 also studied the elbow flexors during sustained isometric
contractions to fatigue. Their subjects performed contractions at 25% of MVC, with and
without 300 mm Hg of pressure in a 2.5 cm wide cuff wrapped around the proximal upper
arm. Similarly to the study of Person & Golubovich327, they found markedly reduced
endurance times and increased EMG activity with occlusion, but without specifying the
magnitude of the EMG changes. However, in his PhD thesis, Sullivan330 noted that integrated
EMG amplitude had increased by 542% at the end of the control contraction, and 285% in the
occluded condition, and that overall the EMG results were directly opposite to those of
Person & Golubovich327. The condiderably lower rise in EMG at task failure in the occluded
condition in the study of Sullivan330 is of interest and suggests a decreased excitability of
active myofibres and possibly also neuromuscular transmission failure, in addition to reduced
excitatory drive during their BFR procedure. It is difficult to explain the disparate results of
Person & Golubovich327 and Sullivan330, but they may be related to the specific criteria of
task failure used in the two studies and whether the exercise was performed with support for
the arm or not. The upper arm was supported in the study of Myers & Sullivan329, whereas
Person & Golubovich327 did not report this.
Interestingly, Person & Golubovich327 as well as Myers & Sullivan329 also
studied the effects of occluding the forearm on endurance time and EMG during the isometric
elbow flexor contractions. Both investigations found decreased endurance times, and Person
& Golubovich327 found that EMG at task failure had increased to 256% of the initial value.
Based on the assumption that the forearm muscles did not participate in the task, they argued
that the intensified changes in biceps EMG during conditions of forearm ischemia were
mediated by a reflex and irradiation effects from non-active muscles.

However, it has been estimated that the biceps brachii and brachialis combined
make up only ~57% of the total elbow flexor muscle area and thus that some forearm muscles
Accepted Article
(particularly the brachioradialis) can make substantial contributions to elbow flexor torque331.
Occluding the blood flow to these forearm muscles during a low-load static elbow flexion
contraction would almost certainly reduce endurance time and cause compensatory increases
in muscle activity in the non-occluded biceps and brachialis muscles. Even so, Myers &
Sullivan329 reported decreased endurance times also when the contralateral inactive forearm
was occluded during the task. Sullivan330 noted that EMG increased 411% in this arm. The
cause of this decreased endurance is uncertain, but may be related to group III/IV afferent
input due to metabolite accumulation and/or pressure (see Sections 3.6. and 6.6.).
Signorile et al327 conducted what seems to be the first study on surface EMG in
low-to-moderate load dynamic resistance exercise with BFR. They investigated the effects of
cuff occlusion on EMG in the biceps brachii during elbow flexion exercise performed over a
padded incline bench, at a load of 25RM (established without BFR), which was estimated to
~50-60% of 1RM. Their experimental subjects performed 2 sets of 25 repetitions, one set
with BFR and one set with free circulation, in a randomized order with 10 minutes between
sets. The pace was 3 seconds for the concentric and eccentric phases, respectively, through a
full range of motion with no stops throughout the exercise. A blood pressure cuff inflated to
midway between the diastolic and systolic pressure was used for the occluded condition.
EMG signals appear to have been recorded from the short head of the biceps. The authors
reported an 86% increase in EMG RMS in the concentric phase from the first to the last
repetition with BFR and a 39% increase in EMG from the first to the last repetition in the
non-occluded condition. In the first repetition, the EMG was non-significantly 12% elevated
in the occluded vs. free-flow condition.

The 86% increase in EMG from the first to last repetition reported by Signorile
and coworkers332 is of note. Given that a load of 50-60% of 1RM was used, this suggests that
Accepted Article
the EMG RMS amplitude reached levels similar to those that can be observed with heavy
resistance exercise. The rather large difference in EMG increases between the conditions is
surprising, given that the load was selected to allow only 25 repetitions even without
occlusion. But as the work was matched between the conditions, the effort was almost
certainly submaximal in the free-flow trial as markedly more repetitions can be done in elbow
flexion exercise with free circulation before torque failure is reached8,333, which would
largely explain the observed difference in EMG activity. Alternatively, since the short head
of the biceps is just one of several elbow flexor muscles (cf. discussion above), other elbow
flexor muscles might have fatigued before the short head of the biceps in the non-occluded
condition, limiting the increase in EMG for this particular muscle.
The study of Leonard et al187 has been discussed in Section 4.2., but merits
further mentioning. The subjects performed unilateral plantar flexions against their own
bodyweight on a 40° inclined sliding board, with 40° of knee flexion to emphasise the soleus.
Sets of 21 repetitions in 30 seconds were performed, separated by brief isometric periods
during which soleus H-reflex, M-wave and EMG measurements were obtained. During the
ischemic condition (300 mm Hg, cuff width not stated), the subjects all reached failure during
the sixth set, resulting in a total of ~105-126 coupled concentric-eccentric repetitions in
approximately 3-4 minutes. It should be noted that the ischemic exercise was preceeded by 8-
12 minutes of ischemia, in order to cause a block of proprioceptive afferents but not motor
efferents. Interestingly, during the last two measurement periods before exhaustion, EMG
RMS was drastically decreased (by ~80-85%) while muscle fibre conduction velocity and
MPF were markedly increased. The authors interpreted this as evidence of failure of slow-
twitch motor units and that the increase in EMG MPF was due to compensatory recruitment

of fast-twitch units. This interpretation is supported by the observation of increased muscle
fibre conduction velocity (see Section 5.5.). It is of note that the soleus muscle typically
Accepted Article
consists of ~80-85% type I fibres227,334,335. This type I fibre dominance likely explains why
there were sudden marked increases in muscle fibre conduction velocity with recruitment of
the type II fibres after severe fatigue in the type I fibres in the study of Leonard et al187.
Two influential studies were published 6 years later by Takarada and
colleagues1,18. Both reports had other primary focuses than EMG recording (also assessing
hormonal responses, and muscle strength and mass adaptations), but included EMG results
that were compared between occluded and non-occluded exercise protocols. In the first paper,
Takarada et al18 measured muscle activity in the vastus lateralis during 5 sets of bilateral
dynamic knee extensions at 20% of 1RM to failure combined with partial BFR, with 30-s
inter-set rest periods. This was compared to a work-matched protocol with the same number
of sets and repetitions without BFR. The BFR protocol comprised a 33-mm wide cuff inflated
to 214 mm Hg, which was maintained throughout the entire exercise including rest periods. It
was found that the relative increase in EMG amplitude during exercise with occlusion was
~1.8 times as large as that during exercise without occlusion18. However, the authors did not
report any details about this result, e.g. whether this was measured as overall mean EMG
activity or peak EMG activity.
In the second paper, Takarada et al1 measured muscle activity in the biceps
brachii during sets of unilateral dynamic elbow flexions at 40% and 80% of 1RM with and
without two degrees of partial BFR (50 and 100 mm Hg using a 90 mm wide cuff). The mean
amplitude of the integrated EMG (iEMG) was calculated from the average of the 7th to 9th
repetitions. Takarada et al1 reported that ”without occlusion, the relative iEMG during the
low-intensity exercise was lower by about 40% than that during the high intensity exercise,
indicating that the time-averaged number of muscle fibres recruited was approximately

proportional to the level of force generation. With the increase in occlusion pressure, the
relative iEMG during the low-intensity exercise gradually increased, whereas that during the
Accepted Article
high-intensity exercise was kept unchanged. Consequently, no substantial difference was
observed between the low-intensity and high-intensity exercises at occlusion pressure of 100
mmHg”. However, although 100 mm Hg significantly increased the iEMG at 40% of 1RM, it
appears that the EMG was still ~15% lower than during 80% of 1RM without occlusion.
Nevertheless, Takarada et al1 were the first to show that the EMG levels recorded during low-
intensity BFRRE could approach those observed during conventional high-intensity
resistance exercise.
Also as part of a training study, Moore et al336measured EMG activity during
elbow flexion at 50% of 1RM with and without continuous partial occlusion (100 mm Hg
with a 7 cm wide cuff). They showed a greater relative increase in EMG between the first
repetition in set #3 over set #1 in the BFR arm compared with the non-occluded arm which
performed the same amount of work (~57% vs. ~11%). Further calculations on their data
suggest that EMG activity from the first repetition in the first set to the last repetition in the
last set increased with ~92% and 47% for the BFR arm and the non-occluded arm,
respectively. Collectively, the findings from the studies by Signorile et al332, Takarada et al1
and Moore et al336 thus suggest that agonist EMG levels recorded during dynamic elbow
flexion BFRRE at 40-55% of 1RM can increase with up to ~90% during the the time course
of the exercise session, reaching levels approaching those seen with heavy resistance exercise
using exercise loads ≥80% of 1RM.
Yasuda et al180 compared the effects of four different degrees of BFR on
relative iEMG during a standardized elbow flexion exercise protocol at 20% of 1RM
consisting of 30-15-15-15 repetitions in sets 1-4. Occlusion pressures were 0 (free-flow
condition), 98, 121 and 147 mm Hg, respectively. The cuff was 30 mm wide and the dynamic

elbow flexion exercise was performed over a 45° incline bench, with each complete repetition
taking 2.4 s, with 30 s rest periods between the sets. In addition to iEMG during the exercise
Accepted Article
bouts, MVC and iEMG during MVC were measured before and 1 and 2 minutes post-
exercise, with maintained BFR during the 1-minute post MVC and free circulation during the
2 minute post MVC. From the first repetitions in the first set to the last 5 repetitions of the
fourth set of the BFRRE bouts, iEMG increased by ~90-135%, with the greatest increases
seen with 147 mm Hg and the lowest with 98 mm Hg. In the free-flow condition, iEMG
increased by only ~60%. MVC measured at 1-minute post decreased in the same order, i.e.
the largest decrements resulted from BFRRE at 147 mm Hg (~38% reduction) and the least
from free-flow exercise (15% reduction). Interestingly, the same order was also observed in
the iEMG amplitude obtained during 1-minute post-exercise MVCs: ~44% reduction at 147
mm Hg, ~32-34% at 121 and 98 mm Hg, and 10% with free circulation.
In a follow-up study, Yasuda et al181 compared the effects of varying degrees of
BFR on iEMG during 3 different elbow flexion exercise protocols at 20% of 1RM: 1 x 30
repetitions (Experiment 1), 3 x 10 repetitions (Experiment 2) and 30-15-15-15 repetitions
(Experiment 3). In Experiment 1, in two conditions restriction pressures of 160 mm Hg
(”moderate restriction”) and 300 mm Hg (”complete occlusion”) were applied, respectively,
while the third condition used free-flow circulation (control). In Experiment 2, the same three
conditions were used. In Experiment 3, the two conditions were 160 mm Hg and free
circulation. Cuff dimensions and the elbow flexion exercise were the same as in the previous
study. In addition to iEMG amplitude during exercise, MVC and iEMG activity during MVC
was measured before and 1 and 2 minutes post-exercise, as in the previous study. In
Experiment 1 and 2, exercise with complete occlusion tended to induce in the greatest
increases in iEMG from the first to the last repetitions (up to ~130%). In Experiment 3, the
highest elevation in iEMG was seen in exercise with moderate restriction (~125%). With the

most fatiguing protocols with moderate restriction and complete occlusion, the decrements in
MVC and iEMG were even greater than in the previous study, with iEMG during 1-minute
Accepted Article
post MVCs decreasing to as low as 40-50% of pre-exercise iEMG.
The EMG activity recorded during BFRRE by Yasuda et al180,181 showed
average values during the final repetitions in the most fatiguing protocols that were about 2.3
times higher than in the first repetitions. This magnitude of change is surprisingly low, as one
would expect a greater increase given that the load was only 20% of 1RM and that the ratings
of perceived exertion after the final repetitions in set 4 were 17-18 on a Borg 6-20 RPE scale,
where 17 is ”very hard”, 19 corresponds to ”extremely hard” and 20 is ”maximal exertion”
(Borg332). In a later study using almost identical methods, Yasuda et al333 reported much
greater increments in EMG activity throughout the period of acute BFR exercise. In this latter
investigation, Yasuda et al333 compared elbow flexion exercise protocols to failure with and
without BFR at 20% of 1RM. In the BFR condition, the EMG values at concentric failure
were up to 3.9 times higher than during the first repetitions. In the non-occluded trial, EMG
increased by up to 4.4-fold. Even these increases suggest a submaximal EMG level in the
highest repetitions during BFRRE compared to free-flow MVC conditions, given the low
loads used and a curvilinear EMG rise with increasing force, (see Section 5.3.1.). For
comparison, the data in Christie et al67 and Moritani & Muro256 suggest that in free flow
conditions EMG RMS activity in the biceps brachii increases ~10-fold from 20% of MVC to
100% of MVC.
The 1-minute post-exercise MVCs after BFRRE with ”moderate restriction” in
Yasuda et al180,181 reveal 45-50% reductions in EMG compared with pre-exercise MVCs, and
almost 60% decrements after BFRRE with complete occlusion. This observation may provide
insight into why EMG did not increase more during dynamic BFRRE at 20% of 1RM. The
45-60% reductions in post-exercise EMG are of a magnitude which is unlikely to be

explained by central fatigue alone, which appears to be limited during occluded exercise of
relatively short durations as judged by interpolated twitch measurements338,339. Thus, more

Accepted Article
likely contributing factors would be decreased motor unit firing rates due to spike-frequency
adaptation and inhibitory group III/IV afferent input. Lower firing rates in some motor units
during fatiguing exercise, as observed by Garland et al139, could result in lower EMG even if
no fibres were derecruited or dropped out due to propagation failure. However, we suggest
that the submaximal exercise and post-BFRRE EMG amplitudes reported in the studies of
Yasuda et al180,181 may reflect a markedly reduced excitability of the muscle fibres, and
possibly some degree of NMJ transmission failure (see Section 4.2., and 6.4.2. below).
Wernbom et al19 investigated EMG responses in the vastus lateralis and
medialis muscles during three sets of unilateral low-load knee extensions to failure at 30% of
1RM with and without BFR (100 mm Hg with a 13.5 cm cuff). In line with earlier results
with near-complete occlusion at 200 mm Hg (Wernbom et al314), the partial occlusion
reduced the number of repetitions compared to free-flow conditions by ~31% in the first set.
High EMG levels were demonstrated in both conditions, with concentric EMG increasing up
to 2.7-2.8 times from the first to the last repetitions (up to 94-102% of MVC EMG), but with
no significant differences regarding maximal concentric muscle activity between the
conditions. Interestingly, EMG increased markedly during the eccentric phase in both
conditions, up to 2.2-fold and 2.5-fold respectively, with a higher EMG in set 3 for the free-
flow condition. Wernbom et al19 were the first to report EMG increases during not only the
concentric but also the eccentric contraction phases during acute bouts of BFRRE. In
addition, it was the first study to compare dynamic low-load training to failure with and
without BFR.
In our experience, very low-load unilateral BFRRE (~20-25% of 1RM)
performed to fatigue results in a more or less hyperbolic increase in EMG RMS amplitude as

the point of concentric failure draws close (an example is shown in Figure 5a, from
Wernbom340). Similar observations have been reported by others (Farup et al8, their Fig.6).
Accepted Article
This appearance is remarkably similar to the EMG curves reported in the studies on low-force
isometric contractions (10-40% of MVC) sustained to fatigue by Vestergaard-Poulsen et
al308,309 and Houtman et al311, discussed in Section 5.5. The noticeably steeper increase in
EMG RMS amplitude in the vastus lateralis and medialis during the last 6 repetitions in the
first set may indicate an additional recruitment of large superficial motor units (presumably
innervating type II myofibres). Note that EMG activity did not seem to reach quite as high
during low-load BFR knee extensions (Figure 5a-d) as during heavy load knee extensions
without BFR (Figure 6)
Investigations by others have confirmed that EMG increases during the eccentric phase in
fatiguing BFRRE and that quadriceps EMG can reach high levels during fatiguing very low-
load knee extensions (20-30% of 1RM) irrespective of BFR341. Notably, however, available
data suggest that EMG amplitudes in the quadriceps at these loads may not reach the levels
seen with high-load (70-75% of 1RM) resistance training341,342, even when low-load training
is performed to failure without occlusion341. In the study of Loenneke et al341, EMG during
the concentric phase of BFRRE at 20% of 1RM did not exceed 60% of MVC in any of the
different BFR conditions, and EMG recorded during 20% of 1RM to failure without BFR
also did not exceed 60% of the MVC EMG activity. The increases in the concentric phase
were 114% in the 20% of 1RM to failure protocol and 45-82% in the BFRRE at 20% of
1RM.
Somewhat in contrast, Fatela et al342 reported that EMG RMS amplitude
increased up to ~100% of MVC in the rectus femoris and ~85% of MVC in the vastus
medialis by the end of 4 sets of knee extensions at 20% of 1RM. However, EMG activity at

the start of the exercise was unusually high (~50% of MVC), which is puzzling considering
the very low exercise load (20% 1RM). The exercise was performed with concentric-only
Accepted Article
repetitions in an isokinetic dynamometer set in the isotonic mode, with 1 s concentric phases
and 1 s rest between repetitions. Another difference was that knee extensions were performed
unilaterally instead of bilaterally as in Loenneke et al341. Interestingly, high-load knee
extensions at 75% of 1RM without BFR were also investigated by Fatela et al342, in which
quadriceps EMG values reached ~125% of MVC in the repetitions with the highest muscle
activity. This is somewhat similar to the findings of Amiridis et al270 and Beutler et al273,
discussed in Section 5.3.1. This increase could be due to increased motor unit recruitment
and/or elevated firing rates and/or changes in muscle fibre excitability during maximal
dynamic compared with isometric contraction conditions.
In summary, evidence suggests that EMG activity during BFRRE with loads of
40-55% of 1RM can reach levels close or comparable to those encountered with high-load
resistance exercise, at least when compared in the final repetitions of the respective exercise
sets. In contrast, yet other study reports indicate that during BFRRE with very low loads
(~20% of 1RM), peak EMG RMS levels may fall short of those seen with conventional heavy
strength training, particularly in the case of BFR training involving the elbow flexors. It is
hypothesised that this discrepancy is mainly because of the effects of decreases in firing rates
and/or muscle fibre excitability, and in addition potentially transmission failure in some
muscle fibres.
Magora et al343 studied the effects of cuff ischemia on motor unit recruitment and activity in
the biceps brachii in a low-load isometric endurance task. The subjects performed a sustained
isometric contraction against a counterload of ~20% of MVC until complete inability to

sustain target force. The task was performed first with free flow and then with cuff occlusion,
with 1 hour of rest in between. Before the ischemic contraction, a pressure cuff (width not
Accepted Article
reported) was inflated to 30 mm Hg above the systolic blood pressure. Occlusion reduced the
time to task failure by 72.5% on average (81 vs. 296 seconds). Interestingly, in the occluded
condition, there were greater numbers of large spike amplitudes and fewer low-amplitude
spikes in the EMG signal recorded at the beginning of the task. The authors concluded that
the contraction was initiated involving a higher proportion of large motor units. Curiously
though, after the initially high levels, the mean maximal spike amplitude decreased and low
amplitude spikes increased in the ischemic contraction, which was opposite to that observed
in the non-ischemic contraction. At task failure, the mean maximal spike amplitude was very
similar between the conditions.
Magora et al343 discussed the decrease in mean maximal spike amplitude during
the ischemic contractions as ”the gradual transformation of big motor units to small”. It is
unclear whether this refers to a partial loss of active muscle fibres in large motor units, thus
effectively rendering active motor units smaller, or the result of a more complete drop-out of
bigger motor units. Regardless, a drop-out of some muscle fibres due to neuromuscular
transmission failure (Section 4.2.) could explain the observed decreases in spike amplitudes.
Moritani et al17 investigated exercise with BFR using both surface and
intramuscular EMG. The subjects performed repeated hand grip contractions at 20% of MVC
for 2 s followed by 2-s rest for 4 min with either free blood circulation or arterial occlusion
given between the 1st and 2nd min (200 mm Hg on the upper arm, cuff width not reported).
The intramuscular motor unit spikes and surface EMG data indicated that mean motor unit
spike amplitude, firing rates, EMG RMS and MPF remained constant throughout the
experiment with unhindered circulation. In contrast, significant increases in mean motor unit
spike amplitude and frequency were evident during the contractions with arterial occlusion,

along with progressive increases in surface EMG RMS amplitude and decreases in MPF. The
authors interpreted the elevations in spike amplitude and frequency as suggesting new motor
Accepted Article
unit recruitment and an increased discharge rate of relatively high-threshold units.
Notably, mean spike amplitude in Moritani et al17 had increased by ~40%
during the last 15 seconds of the occlusion and at this point EMG RMS was elevated by
about 90%. The gradual increase in both parameters would appear to reflect fatigue-induced
compensatory recruitment, although altered motor unit activation patterns resulting from
pressure-induced stimulation of cutaneous receptors and other mechanically sensitive
afferents cannot be ruled out (see Section 6.6.).
6.5. 31P-MRS studies on muscle fibre recruitment in BFRRE
As with other types of exercise, the occurrence of ”split” Pi peaks in 31P-MRS spectra has
been used to infer type II fibre recruitment with strenuous BFRRE9,10,344-346. In these studies,
a unilateral plantar flexion training model was employed, involving the triceps surae muscles
(the gastrocnemius medialis and lateralis, and the soleus) and consisting of coupled
concentric and eccentric contractions. In one of the first studies on BFR resistance-type
exercise and muscle fibre recruitment, Yoshida & Watari344 reported split peaks during
occluded but not during free-flow exercise to exhaustion, suggesting recruitment of type II
fibres in the former mode of exercise but not the latter.
However, since they reported neither the workload in watts nor the absolute
load in kilograms (or relative load in % of 1RM), and due to other uncertainties in the
description of the exercise (e.g. range of motion was not noted), it is difficult to assess
relative exercise intensity in this study. More importantly, the 80 mm surface coil was placed
over the center of the calf muscle (i.e. between the gastrocnemius medialis and lateralis
muscles), which would almost certainly mean that both the individual muscle components of

the gastrocnemius were sampled, increasing the risk that the detection of split Pi peaks was
caused by contamination artifacts due to the non-localized 31P-MRS spectra rather than due to
Accepted Article
recruitment of fresh type II muscle fibres. In principle, the fact that at least two different
muscles were sampled limits the conclusions pertaining to the pattern of motor unit
recruitment that can be derived from the study of Yoshida & Watari344.
In a series of studies, Suga, Okita, Takada and colleagues9,10,345,346 investigated
low-load plantar flexion exercise combined with BFR. They used mostly an intensity of 20%
of RM, although loads of 30% and 40% of 1RM with BFR were also studied, as well as 65%
of 1RM without occlusion. BFR was achieved using a cuff around the thigh, and the
combination of very wide 18 cm cuffs and ~143-150 mm Hg of pressure was likely sufficient
to cause near-complete occlusion in their supine test position (see Crenshaw et al313), as also
indicated by the very limited recovery in PCr observed during the rest periods between sets
(see Suga et al9). Similarly to Yoshida & Watari344, split Pi was observed albeit to varying
degrees in the different studies. However, it is not immediately obvious whether the split Pi
peaks reported in the studies of Suga and colleagues9,10,345,346 truly reflected the initial
recruitment of type II fibres (type IIA) in addition to already active type I fibres, as the split
Pi peaks could also mark the recruitment of type IIX and/or IIAX fibres in addition to already
active type IIA fibres, along similar lines as the suggestions put forth by Vandenborne et
al284, (cf. Section 5.4.). This possibility was not discussed explicitly in the studies of Suga et
al9,10,345,346.
Notably, even the high load free-flow protocol (65% of 1RM) which was
investigated along with the BFRRE protocols in these studies9,345,346 did not yield dual Pi
peaks in all subjects, despite each exercise set consisting of 30-60 repetitions, for a total of
60-90 repetitions per session. This is remarkable, as is the fact that the subjects performed up
to 60 repetitions per set, since a load of 65% of 1RM would be expected to recruit type IIA

fibres already from the start of the exercise while also involving type IIAX and IIX fibres to
become active not long after. In comparison, Trappe et al347 reported that their subjects were
Accepted Article
able to perform only about 15 repetitions of unilateral plantar flexion at a load of ~70% of
1RM. Notably, the subjects in the paper of Trappe et al347 performed standing unilateral
plantar flexions with bodyweight (80 kg) and up to 10 kg of extra weight, to compare with 33
kg in supine unilateral plantar flexion for the male subjects employed by Suga et al9.
These puzzling discrepancies in load-repetition characteristics between studies
may in part be explained by differences in posture (supine vs upright) and range of motion
(ROM). Available data suggests that plantar flexion torque decreases in an essentially linear
manner with increasing plantar flexion angles, so that low-velocity concentric torque at 30
degrees of flexion (0 degrees = neutral position) is only about 25-30% of that at -10 degrees
of flexion348-350. Accordingly, Gravel et al348 concluded that the maximal strength capacity of
the plantar flexors is best represented by the ankle extensor moment measured in slight
dorsiflexed joint angles when the muscles are at elongated length. In the studies of Suga and
colleagues, the 1RM was determined concentrically as the highest weight the subject could
lift completely throughout a limited ROM (~5 cm). This probably means that the 1RM was
mainly dependent on the torque capacity at the end of the ROM, which would be
considerably lower than at the start of the movement. It is also of relevance that concentric
plantar flexion torque is markedly enhanced by preloading350 and by a preceding stretch-
shortening cycle even at submaximal loads351.
Therefore, it is likely that the plantar flexion movement employed in the studies
of Suga and colleagues9,10,345,346 was underloaded during a large part of the ROM, thus
requiring much less effort and recruitment during this part than at the end range of the
movement. In this scenario, type II fibres would not necessarily be recruited from the start of
the exercise even at a nominal load of 65% of 1RM, except for perhaps a brief activation

during the last degrees of concentric plantarflexion. This would in part explain that the
subjects were able to perform as many as 60 repetitions per set with 65% of 1RM, apparently
Accepted Article
without reaching exhaustion, and how the same subjects could perform a total of 90-120
repetitions during near-complete occlusion at 20% of 1RM, also seemingly without reaching
concentric contraction failure.
The end-exercise pH values after sets of BFRRE with near-complete occlusion
at 20% of 1RM in these studies9,10,340,341 were between ~6.84 and 6.92, with a resting pH of
about 7. Interestingly, in the studies where multiple sets of repetitions were employed at 20%
of 1RM9,10, end-exercise pH values clearly tended to be lower than in the studies using only a
single set protocol340,341: ~6.84 vs 6.91-6.92. Moreover, the percentages of subjects
displaying split Pi peaks were much greater in the studies with the lower end-exercise pH
values: 64-75% vs 31-33%. Notably, end-exercise pH in the range of 6.6-6.7 have been
reported for the gastrocnemius muscle after strenuous plantar flexion exercise295,300 and 6.7
after BFRRE at 30% of 1RM352. Therefore, pH values may well be expected to drop to ~6.6–
6.7 and possibly even lower in the gastrocnemius muscles following low-load BFRRE
protocols performed to torque failure. Given the greater incidence of split Pi peaks in these
studies once the mean pH levels dropped below 6.9 (discussed above), recruitment of type II
fibres may have occurred once pH dropped within the range of ~6.8-6.9 in the BFR plantar
flexion exercise model of Suga and colleagues9,10,345,346, suggesting that this response
reflected the firstly recruited population of type II fibres (i.e. type IIA).
Nevertheless, some puzzling observations are not easily explained by the above
scenario. For example, some of the subjects in the study of Suga et al346 were able to perform
as many as 60 successive repetitions in the high-intensity resistance exercise protocol during
free-flow conditions without displaying split Pi, and they also managed to perform 60
repetitions during near-complete occlusion at 20% of 1RM, also without generating visible

split Pi peaks. However, most of these subjects displayed split Pi peaks after 60 repetitions at
40% of 1RM with BFR, which was the most PCr depleting protocol. Therefore, it seems
Accepted Article
possible that the surprisingly late split Pi peaks observed by Suga et al346 in these subjects
were actually signifying recruitment of type IIX and/or IIAX fibres, rather than IIA fibres. In
this scenario, type IIA fibre recruitment may not have been detected for at least two possible
reasons: a relatively small pure type IIA pool in these individuals and/or only minor
differences between type I (including type I-2) and type IIA fibres in terms of intracellular
lactate accumulation.
In this context, it is noteworthy that although biopsy studies with myosin
ATPase stainings have shown that the gastrocnemius medialis and lateralis muscles are
composed of ~40-50% type II fibres227,259,334,335,353, results from a study which used a
staining method for lipid droplets and mitochondria indicate that the gastrocnemius muscles
consist of as much as 82-84% mitochondria-rich fibres, 8-9% intermediate fibres and 7-10%
mitochondria-poor fibres38. While the latter two fibre phenotypes might correspond to type
IIAX and type IIX, the discrepancy between the 50-60% type I fibres reported for the
gastrocnemius in the aforementioned studies and the large proportion (82-84%) of
”mitochondria-rich fibres” reported by Buchthal & Schmalbruch38 strongly suggests that a
considerable portion of type II fibres, presumably mostly type IIA fibres, are rich in
mitochondria and lipid droplets to an extent that approaches type I fibres. This notion is in
line with the overall slow contractile properties of the gastrocnemius reported by these same
authors38 and also with the observation that FR units (presumably consisting of type IIA
fibres) in the human gastrocnemius medialis have a fatigue resistance profile resembling that
of S units, which consist of type I fibres38.
With regard to the distribution of type II fibre subtypes in the gastrocnemius, a
biopsy study on the gastrocnemius lateralis (GL) muscle in habitually physically active

subjects showed only 3% type IIX fibres, while type IIA and type I fibres comprised 38% and
59% of the myofiber pool, respectively, as judged by ATPase staining analysis354. A study on
Accepted Article
fibre type distribution in the GL in young untrained but healthy subjects report that the GL
contained 59.4% type I fibres and 26.4% type IIA fibres, with the remaining ~14% classified
as type IIX fibres also using ATPase staining355. Assuming that both type I and type IIA
fibres in the human gastrocnemius are rich in mitochondria and lipid droplets, and that some
of the type IIX fibres in Vandenborne et al355 were in reality type IIAX, the data reported by
Vandenborne et al355 seem to match those of Buchthal & Schmalbruch38 reasonably well.
Collectively, this information suggests that type IIA is the most common type II
subtype in the human gastronemius in young healthy subjects and that type IIA fibres in the
gastrocnemius muscle are nearly as fatigue-resistant as type I fibres in the same muscle,
probably at least in part due to their abundance of mitochondria and lipid droplets. In turn,
this similarity may be of considerable significance for the balance between aerobic and
anaerobic metabolism during exercise. Specifically, it may be that the differences in lactate
accumulation and associated drops in pH between type I and type IIA fibres in the
gastrocnemius during exercise are relatively minor, thus making it difficult to detect split Pi
peaks in the 31P-MRS spectra caused by the recruitment of type IIA fibres in addition to
already active type I fibres.
We therefore speculate that the Pi splits reported in the studies of Suga and
colleagues9,10,345,350 may to a large extent reflect a progressive recruitment of type IIAX
and/or type IIX fibres, rather than type IIA fibres. This possibility is further indictated by the
observation in the study of Suga et al346 that the RPE at the end of the BFRRE protocol (1 set
of 60 repetitions at 20% of 1RM) was 6.6 out of 10 on the Borg CR-10 scale (i.e. close to
”very strong”), despite that split Pi was observed in only 31% of the subjects. In comparison,
the high load protocol (1 set of 60 repetitions at 65% of 1RM) resulted in split Pi peaks in

70% of the subjects and an RPE of 8.2. The most strenuous BFRRE protocols at 20% of 1RM
in the studies of Suga and Takada et al9,10 resulted in Pi splits in 64-75% of the subjects.
Accepted Article
However, an important limitation of the studies of Suga et al9,345,346 and Takada
et al10 is that the exact position of the MRS surface coil used to detect the metabolic changes
was not reported, the only information provided in all of the reports was that ”an 80-mm
surface coil was placed under the muscle belly of the right gastrocnemius”. If the 80 mm
surface coil was placed between the gastrocnemius medialis and lateralis muscles, as in
Yoshida & Watari344, it would probably mean that all these studies are prone to the same
criticism as the report of Yoshida & Watari344 with regard to the potential effects of signal
contamination due to non-localized 31P-MRS spectra.
Regardless, the marked gains in both muscle strength and CSA of the plantar
flexors following 4 wks of BFRRE reported by Takada et al10 are noteworthy given the very
low exercise loads used in this study, in actuality probably closer to 10-15% of 1RM rather
than the reported 20% of 1RM. This raises interesting questions regarding the minimum load
in BFRRE that would be sufficient for inducing muscle hypertrophy and functional strength
gains.
6.6. Can muscle fibre recruitment order be changed during cuff occlusion?
Some authors have suggested that the normal order of activation of muscle fibres (type I 
type IIA  type IIX fibres according to the Henneman size principle of motor unit
recruitment) may be altered during exercise with occlusion, so that type II fibres could be
activated preferentially over type I fibres356,357. To the best of our knowledge, at least four
studies have reported indications of possible alterations in the recruitment order during
occluded blood flow, although being observed during relatively long-lasting ischemia (~20-
30 minutes) or with ischemia of unspecified durations108,191,343,358.

In their study on impulse transmission in the extensor digitorum communis
(EDC) muscle in humans with needle electromyography, Dahlbäck et al191 observed that
Accepted Article
”during ischemia the recruitment pattern of the motor units altered so that those formerly only
recruited at stronger contraction were now the first to be recruited”. Dahlbäck et al191 used a
blood pressure cuff (width not reported) on the upper arm inflated to 200 mm Hg to induce
ischemia, and instructed their subjects to maintain contractions with steady motor unit firing
rates of between 2-15 Hz. Probably because their main focus was neuromuscular
transmission, they did not further specify the strength of the contractions used and the
approximate level of effort that was required to recruit the higher-threshold units prior to
ischemia, nor did they report the time of ischemia when the reported alterations took place.
Nevertheless, it was stated that the experiments were performed on motor units which could
be activated even at the slightest contraction of the muscle. Furthermore, data in Monster &
Chan58 obtained in the EDC muscle show that the first recruited units can reach ~16 Hz
already at a few percent of the MVC. Taken together, this suggests that the contractions used
by Dahlbäck et al191 involved very low forces even when motor unit firing rates were 10-15
Hz.
Grimby and Hannerz studied motor unit recruitment order and firing rate ranges
during ischemia in the tibialis anterior and the extensor digitorum brevis muscles using fine
wire and needle electrode recording (Grimby & Hannerz358; Hannerz & Grimby108). A
pressure cuff (width not stated) was applied around the thigh and inflated to higher than the
systolic blood pressure, to ~140 mm Hg. The sensory and motor functions of the lower leg
successively disappeared over a time course of 20-40 minutes. It appears from the description
in Hannerz & Grimby108 that the occlusion was not quite complete, as they chose a pressure
which induced a partial nerve blockade in order to lengthen the period during which the
motor responses could be studied. In the first study, Grimby and Hannerz358 investigated

recruitment changes in the tibialis anterior during partial ischemic nerve blockade with
dorsiflexor test contractions at 10-20% of MVC for 20-30 minutes. At 15-20 minutes into the
Accepted Article
ischemic period, an altered order of motor unit recruitment was observed, in that increasingly
more other units became recruited before the unit which had the lowest threshold in free-flow
conditions. By 25 minutes, a few high-frequency units also had lower thresholds than this
unit. Presumably, these were high-threshold units, as they were not activated during test
contractions at 10-20% of MVC performed after the ischemic blockade had ended.
In their second study, Hannerz and Grimby108 focused their attention mostly on
motor unit firing rate changes during ischemia, but made some additional observations on
recruitment thresholds. While most intermittently firing units did not change their firing
properties during ischemia, three high-threshold units in the tibialis anterior which normally
were recruited above 75% of MVC became recruited at substantially lower absolute and
relative tensions upon ischemic blockade. In one dramatic example, a very high threshold
unit which was normally recruited at 88% of MVC became active already at 4% of the pre-
fatigue MVC during ischemic conditions, when the maximum tension had dropped to 32% of
the baseline MVC. The drastic decrease in recruitment threshold strongly suggests that for
this particular unit, an almost complete reversal in the normal recruitment order must have
taken place.
The findings of Magora et al343 (discussed previously in Section 6.4.2.) of
greater numbers of large EMG spike amplitudes and fewer low-amplitude spikes in the
beginning of an ischemic low-load endurance task may also be interpreted as evidence of
deviations in recruitment order. At a first glance, the increase in the average amplitude of the
motor units during the early part of the ischemic contractions seems quite large (72%).
However, the range of motor unit amplitudes that can be observed in the biceps brachii
during an MVC suggests on the order of ~30-fold differences between the smallest and the

largest amplitudes (see e.g. Moritani et al279). Data on motor unit amplitude in relation to
recruitment threshold in the biceps brachii during free-flow conditions in the paper of
Accepted Article
Gydikov & Kosarov109 suggests that a 72% increment in spike amplitude over those of motor
units recruited at 20% of MVC would correspond to units recruited at ~30% of MVC.
Furthermore, using multiple indwelling electrodes, Buchtal et al359 reported that during free-
flow conditions the amplitudes of successively recruited motor units in the biceps brachii
increased so that the secondly recruited units in their recordings on average demonstrated a
98% greater EMG amplitude than the firstly recruited units. The firstly recruited motor units
were active at very weak contraction efforts, less than required to support the weight of the
forearm.
Taken together, these observations indicate that if recruitment order changes
due to cuff occlusion occurred in the study of Magora et al343, they were still modest, possibly
on the same order as those observed in Masakado et al76 or less. It should also be noted that
the recruitment order of different motor units was not directly investigated in the study of
Magora et al343, and reversals in the recruitment order were therefore not definitely proved or
disproved in their study.
As discussed above, alterations in the motor unit recruitment order during cuff
ischemia might be caused by blockade of afferent inflow to the CNS358, intramuscular pain74,
and enhanced cutaneous inputs61,75; and ischemia can also impact on the activation of
individual myofibres in a given motor unit via effects on the motor end-plates and muscle
fibre excitability and/or motor nerve axons (Section 4.2.). In addition, the magnitude of
applied pressure can affect alpha-motorneuron excitability per se. Specifically, application of
limb pressure led to decreased Hoffman-reflex (H-reflex) responses both in a massage
setting360 and when administered as circumferential pressure via air splints or pressure
cuffs361,362. The applied pressures in these investigations were 19-28 mm Hg in the study of

Goldberg et al360 and 37-45 mm Hg in Robichaud et al361 and Agostinucci362. In contrast,
Agostinucci363 found no decrease in the soleus F-wave response with 45-50 mm of pressure.
Accepted Article
Agostinucci363 suggested that this difference reflected the different types of motor units
involved in the two responses, with the H-reflex primarily recruiting smalller motorneurons,
while the high-stimulus F-wave (supramaximal stimulation) may preferentially recruit larger
faster conducting motorneurons. If this scenario is true, it suggests the interesting possibilty
that alterations in recruitment order due to applied limb pressure may occur also in voluntary
muscle contractions.
If the occlusion is applied just before onset of contraction, as appears to have
been the case in the study of Magora et al343, partial reversals due to loss of some small motor
units seem unlikely, as the duration of fatigue would likely not have sufficient time to affect
the axonal branches, the motor end-plates and/or the excitabiliy of the slow fibres. In this
scenario, a facilitation of larger motor units arising from the cutaneous stimulation of the cuff
over the working muscle would seem a more logical explanation, in line with the study of
Garnett & Stephens61. An influence from group III/IV afferents would at a first glance seem
improbable to cause such rapid effects, as the firing of these afferents is known to be
stimulated by metabolite accumulation, which in turn requires a certain time to develop.
However, group III/IV afferents have also been shown to be very sensitive to pressure,
significantly more so than to stretch, and there is a linear relationship between the level of
pressure applied and the response of these afferents364. The alterations in motor unit activity
in the study of Magora et al343 might thus be explained by the placement of the cuff over the
biceps muscle belly and its effects on the inflow from cutaneous and/or group III & IV
afferents onto spinal alpha-motorneurons, with a consequent inhibition of smaller motor units
and a facilitation of larger units.

In contrast, the PCr and glycogen depletion data from the studies of Ingemann-
Hansen et al225, Krustrup et al20 and Cumming et al21 (discussed in detail above) are if
Accepted Article
anything consistent with an orderly recruitment of motor units. If any major departures from
the size principle occurred in these studies, this should have been reflected in greater
depletion of PCr and glycogen in type II fibres compared with type I fibres. Instead, an
overall tendency for the expected result can be seen, i.e. a greater use of type I fibres.
Altogether, the available evidence suggests that the size principle for motor unit recruitment
is largely intact during dynamic BFRRE and that if reversals occur, they are relatively minor
(only affecting a few units) and/or of temporary nature. The muscle activation patterns
observed by 31P-MRS analysis and EMG recordings are consistent with this scenario. If
systematic reversals or at least major deviations take place during dynamic BFRRE, these
would be expected to show up in the 31P-MRS spectra in the form of a lower pH Pi peak of
type II fibres appearing at the same time or even before the higher pH Pi peak of type I fibres,
due to the fast activation and rate of glycolysis in type II fibres. This has not been the case
(discussed in detail above). Moreover, the observation by Leonard et al187 of markedly
greater muscle fibre conduction velocites (signifying type II fibre recruitment) near the point
of exhaustion during low-force ischemic exercise in the soleus muscle is also consistent with
a largely intact orderly recruitment. Importantly, this was the case in spite of a probably
blocked proprioceptive inflow.
7. Conclusions and implications.
7.1. Order of muscle fibre recruitment in BFRRE
Of the various methods of investigating muscle fibre activation which have been reviewed
here, only glycogen and PCr depletion studies can provide essentially direct evidence of fibre

recruitment and usage during acute BFRRE, although these methods are not without caveats.
However, complementary information may be derived from EMG and 31P-MRS studies
Accepted Article
involving various BFRRE protocols. Studies with intramuscular EMG electrodes have
reported indications of alterations in the recruitment order during low-load contractions
combined with cuff occlusion, but these have generally been observed with isometric
contractions performed after many minutes of ischemia, with the possible exception of the
study of Magora et al343. Even under prolonged ischemic conditions, the extent of motor unit
recruitment alterations seems minor, and the relevance of these observations to BFRRE is
uncertain. The data from PCr and glycogen depletion studies involving BFRRE are consistent
with a largely intact order of motor unit recruitment, and indirect evidence from 31P-MRS and
EMG investigations further support this notion. In other words, there are no indications of
any major departures from the Henneman size principle for motor unit recruitment in low-
load dynamic resistance exercise using concurrent cuff occlusion.
7.2. Patterns of muscle fibre use and fatigue in BFRRE
The evidence reviewed strongly suggest that although type II fibres (including type IIX) can
be recruited with fatiguing low-load BFRRE, type I fibres are used to a greater extent, as
judged by both glycogen and PCr depletion data. In a previous paper, we suggested a
scenario where the type I fibres become highly fatigued during a first set of unilateral BFRRE
at 30% of 1RM, thus necessitating the recruitment of an increasing number of motor units
containing type II fibres as the exercise progresses. We here extend this model by proposing
that all subtypes of type II fibres (type IIA, type IIAX and type IIX) can be recruited during
low-load BFRRE even at 20-30% of 1RM if the work duration and degree of fatigue is close
to maximal. The suggested scheme for the recruitment of motor units during fatiguing
BFRRE is depicted in Figure 7.

In addition, we previously proposed that type II fibres will have to be recruited
earlier in a second and a third set than in the first set, presumably because of residual fatigue
Accepted Article
in type I fibres19. Importantly though, the typically short inter-set rest periods in BFRRE and
the differences in metabolite and force recovery between the fibre types (including subtypes)
will undoubtedly influence the specific fatigue patterns. Available evidence shows that if all
sets are performed to failure, the number of repetitions will drop for each set, but especially
between the first and second sets. For example, Wernbom et al178,324 reported that with 30%
of 1RM and 45 s rest periods, the number of repetitions performed before concentric torque
failure gradually decreased from 28 in the first set to 10 in the second set and to 6 ± 1 in the
fifth set, for a total of 57 repetitions. This clearly suggests a flatter slope in the decline of
contractile work output after the first sets, particularly from sets 3 to 5 where the slope is
approaching a plateau. Similar results can be found in yet other studies involving multiple
sets of BFRRE performed to failure such as Nielsen et al16 and Farup et al8.
Based on this observed plateau combined with the knowledge of the fatigue and
recovery of the differerent types of muscle fibres, the following modification to the scenario
of Wernbom et al19 is suggested: in a series of 4-5 sets of low-load BFRRE to failure, type I
fibres become highly fatigued during the first set, thus necessitating the recruitment of type II
fibres. However, type II fibres will inevitably also become fatigued. Because the plateau is
largely reached already after the second set, it is suggested that type II fibres also are severely
fatigued at this point. In result and since type I fibres have a superior rate of recovery in both
metabolites and force, they will produce the majority of the force output from set 3 and
onwards. The maintained partial occlusion pressure will if anything probably put the type II
fibres at a further disadvantage, and the type IIX and IIAX fibres will soon become
ineffective due to the lack of recovery and may even drop out when approaching contraction
failure.

A lower force production in the muscle fibres would be predicted to result in
reduced glycogen use, as glycogenolysis and glycolysis are related to the ATP turnover in the
Accepted Article
contracting muscle, which in turn is related to the force development166,365. The scenario of
markedly decreased forces and lower glycogenolytic rates in type II fibres after the first two
sets of BFRRE to failure, (and even a drop-out of some fibres) would thus largely explain the
findings of Cumming et al21 of less glycogen depletion in the type II fibres than in type I
fibres. Conversely, however, the ischemic conditions will accelerate the glycogenolysis in the
type I fibres as previously discussed, and the type I fibres will presumably recover their force
development sufficiently between exercise sets for a relatively high rate of glycogenolysis in
this fibre type to be maintained. Most importantly, the type I fibres will be exposed to a far
greater number of contractions in total.
In low-load BFRRE protocols of 4-5 sets with fixed number of repetitions (e.g.
30-15-15-15), motor unit recruitment typically will be submaximal in the first two sets,
whereas in the last sets the effort may well become near-maximal. Assuming a largely intact
size principle for motor unit recruitment, it is suggested that it is mainly in the later sets of
high efforts that type IIAX and type IIX fibres will be engaged to a significant degree. Still,
type I fibres will likely be recruited for a far greater number of contractions.
The scenarios outlined above mainly concern unilateral BFR resistance training,
engaging a relatively small total muscle mass (ranging from e.g. wrist and elbow flexors to
the quadriceps). As outlined in Section 3.7., performing bilateral exercises will increase the
magnitude of group III/IV afferent inflow to the CNS, and likely inhibit the maximun muscle
activation levels reached during severely fatiguing exercise. This will almost certainly apply
to BFRRE as well. Consequently, it may be hypothesised that performing bilateral knee
extensions with occlusion can result in lower peak muscle activation levels than unilateral
knee extensions with occlusion.

In our experience, it is very demanding even for well-trained athletes to perform
bilateral multijoint movements such as squats and leg presses at very low loads to failure with
Accepted Article
BFR. Accordingly, it is postulated that the voluntary drive during large muscle mass
exercises with BFR at ~20-30% of 1RM may rarely reach levels sufficiently high to recruit
the highest threshold motor units and their muscle fibres. Some support for this notion may
be found in the study of Bjørnsen et al23, who in high-level powerlifters observed markedly
lower EMG amplitudes during low-load bilateral front squats performed to failure with BFR
as compared with bilateral front squats with heavy resistance but without BFR.
However, as mentioned earlier, it is also possible that some fibres may
experience acute and severe reductions in excitability and force production and even drop out
during such protocols, along with decreased motor unit firing rates, all of which could
contribute to the attenuated EMG amplitudes observed in the low-load BFRRE protocol.
Such factors could conceivably act in concert with submaximal recruitment and too short
inter-set recovery periods to limit the training-induced hypertrophic effects in type II fibres,
as elaborated below.
7.3. Consequences for hypertrophic adaptations to BFRRE
As noted above, low-load BFRRE to failure will initially cause severe fatigue in the type I
fibres in the first 1-2 sets. Especially towards the end of the first sets, type II fibres will
probably be primarily responsible for the force production in the working muscle, since the
severely fatigued type I fibres by then will produce relatively little force. Consequently, the
type II fibres may be exposed to relatively high levels of tension development, as previously
suggested307. An illustrative analogy would be various models of compensatory overload,
where synergist muscles are rendered inactive by surgical procedures (tenotomy, synergist
removal or denervation) leaving the remaining muscle(s) having to compensate for the loss of

active muscle, which induces muscle hypertrophy366. BFRRE could thus be seen as a
temporary functional ablation of the type I fibres as they become fatigued, leaving type II
Accepted Article
fibres to be mainly responsible for generating contractile force.
After the first 1-2 sets, fatigue will result in markedly lower force development
in type II fibres. At the same time, the greater recovery capacity of the type I fibres and the
shorter duration of the subsequent sets will shift more and more of the relative load back to
the type I fibres. As a result, type I fibres likely will be exposed to a greater time-tension
integral, and perhaps experience an overall stronger stimulation of hypertrophy signaling
through mechanotransduction pathways, than type II fibres. The large HSP responses in type
I fibres observed in the study of Cumming et al21 support this scenario, although possibly also
resulting from synergistic interactions between mechanical and ischemic stresses.
The presence of additive or synergistic interaction(s) between the mechanical
and ischemic stimuli in BFRRE has been hypothesised previously5,367 and might in part
explain how loads as low as 20% of 1RM can result in marked muscle hypertrophy, despite
seemingly producing only moderate mechanical stress even in the ”functional ablation”
scenario outlined above. Although still somewhat speculative, there are good reasons to
postulate such interactions, given that heat stress alone and repeated brief ischemia-
reperfusion alone have been shown to induce muscle hypertrophy in some studies368,369 and
that concomitant blood flow restriction can potentiate the muscle hypertrophy370-372 and
strength gains370,373 resulting from electrically stimulated contractions. Furthermore, Goto et
al374 reported that unilateral low-intensity resistance exercise for the elbow flexors combined
with heat stress in the same arm resulted in increased elbow flexor muscle CSA, whereas the
same unilateral exercise protocol alone did not produce any hypertrophy in the contralateral
arm.

However, if mechanical stimuli are important for the effects of BFRRE, the
typical way of performing BFRRE, i.e. 3-5 sets of high-number repetitions with short inter-
Accepted Article
set rest periods and with maintained partial occlusion, may not necessarily represent the
optimal choice for eliciting type II fibre hypertrophy and type II fibre-related strength and
power gains. In the case of multiple sets performed to failure, the number of repetitions with
relatively high tension in these fibres will probably be limited for reasons discussed above. It
may be speculated that once active force generation is markedly reduced in the muscle fibres
due to severe fatigue, this will limit any further synergistic effects between
mechanotransduction and ischemic stresses on hypertrophy signaling, particularly in type II
fibres. In the case of multiple sets of standardized repetitions (e.g. 30-15-15-15), the last 1-2
sets are generally the ones with the highest muscle activation and likely favoring the most
extensive type II fibre recruitment. Here too, the number of repetitions performed with
relatively high tension development in type II fibres might be limited.
Nevertheless, a relatively low volume of repetitions with high tension
development in the type II fibres during BFRRE is not necessarily problematic, as already
~10 repetitions in total in conventional heavy resistance training can result in measurable
muscle hypertrophy (reviewed by Wernbom et al375) accompanied by both type I and type II
myofibre hypertrophy376. Furthermore, the results of Nielsen et al16 demonstrate that
substantial hypertrophy in both type I and type II fibres can be achieved in a short time period
with high-frequency (once to twice daily) unilateral BFRRE. In contrast, a recent study by
Bjørnsen et al24, who employed greater volumes of BFRRE per session than Nielsen et al16,
showed temporary decreases (particularly in type II fibres) and subsequent delayed increases
in myofibre areas after short-term high-frequency BFRRE, accompanied by delayed gains in
muscle strength. Interestingly, the temporary type II atrophy in the study of Bjørnsen et al24

was correlated with the increased mRNA expression of p21, which is upregulated in skeletal
muscle during various atrophic conditions (see Bjørnsen et al24 for discussion).
Accepted Article
Thus, there seems to be a relatively narrow window of overall training stress per
session that is well tolerated by the muscles, and especially so for high-frequency BFRRE,
and that exceeding a certain threshold can be detrimental for the resulting muscle adaptations.
This delicate balance may in part explain why exercising to volitional failure in low-load
BFRRE may not necessarily be more effective for inducing gains in muscle strength and size
than BFRRE with submaximal effort, even at more normal training frequencies377.
Furthermore, there is evidence that high volumes of BFR exercise may be
unproductive in terms of muscle strength even in the absence of measurable muscle damage
and stress. For example, Jakobsgaard et al22 did not find any increases in myofiber areas or
muscle strength after a 6-week training period with three weekly sessions of high-repetition
bodyweight squats with BFR, despite modest but statistically significant gains in quadriceps
CSA, myonuclear addition in type I fibres, and considerable increases in endurance capacity
(ending up at ~200-300 repetitions per session). We therefore propose that BFR exercise
protocols involving several hundreds of repetitions in total may simply be too endurance-
oriented to induce muscle adaptations that result in marked increases in maximal muscle
strength and power. In light of the pronounced upregulation of p21 mRNA that has been
found after strenuous BFRRE (discussed in Bjørnsen et al24), there may also have been
insufficient recovery between the high-volume training sessions in Jakobsgaard et al22 for
optimal muscle adaptations to take place.
Finally, differences in training frequency and the use of unilateral single joint
low-load BFRRE in Nielsen et al16 as opposed to bilateral multi-joint low-load BFRRE22,23
may also have contributed to the contrasting results of these studies with reference to
preferential type I vs type II fibre hypertrophy and myonuclear addition. Thus, submaximal

recruitment and firing rates of high-threshold motorneurons innervating type II fibers due to
the especially pronounced inhibitory influence of group III/IV afferents in large muscle mass
Accepted Article
exercises, and an overall too low force-time integral in these myofibres during the exercise
may in part explain the hypertrophy in type I myofibres but no change in type II fibre area in
powerlifters in response to a 7-wk training cycle that incorporated low-load BFRRE23.
7.4. Consequences for neural adaptations to BFRRE
It is suggested that because of the inhibitory influence of group III/IV afferent input to the
CNS and the pronounced spike-frequency adaptation during ischemic exercise, a limit might
be imposed on the firing rates in the highest-threshold units. It is thus possible that such units
will not reach near-maximal discharge frequencies during a typical high-repetition BFRRE
protocol even during the repetitions with the overall highest activation, much like the
situation in slowly ramped-up isometric MVCs (cf. Section 3.6). This may be particularly
true when large muscle mass exercises are combined with BFR. In turn, this would result in
less-than-optimal neural adaptations from a strength and power perspective.
In support of this notion, several studies have reported that low-load BFRRE induces
strength gains and muscle hypertrophy without any significant changes in measures of
voluntary activation and neural drive378-380. In addition, high load strength training appears to
induce greater neural adaptations than low-load BFRRE378,380. Combined, this information
indicates that strategies and training programs that are designed to enhance the degree of
neuromuscular activation during BFRRE are warranted if the goal is to optimize neural
adaptations.

7.5. Consequences for interpretations of surface EMG during BFRRE
To summarize from Section 6.4.1., it is hypothesised that the seemingly submaximal EMG
Accepted Article
levels during very low load BFRRE, which are seen even at very high levels of effort are in
large part due to the combined effects of decreases in motor unit firing rates and attenuated
muscle fibre excitability, potentially compounded by neuromuscular transmission failure in
some fibres. Since any combination of these three factors can result in decrements in EMG
activity, it may also be argued that using surface EMG recording to assess muscle activation
levels at the start and at peak exercise may underestimate the number of muscle fibres that
have been activated during a BFRRE protocol. For example, an EMG RMS amplitude during
low-load BFRRE which reaches only 60% of MVC even at concentric failure does not
necessarily exclude that the highest threshold units (innervating type IIX and/or IIAX fibres)
are recruited, as some of the motor units may demonstrate lowered firing rates and/or a
reduced number of active muscle fibres at this point.
As the muscle mass involved increases from unilateral to bilateral single joint to
bilateral multi-joint exercises, the influence of central fatigue (largely mediated by group
III/IV afferent input) also is expected to increase, which will probably affect both recruitment
thresholds and firing rates of motor units negatively during fatiguing low-load BFRRE.
Hypothetically, this will in turn be reflected in lower maximal EMG amplitudes during low-
load BFRRE with bilateral multi-joint exercise compared with unilateral single joint exercise.
On the other hand, since sEMG may not detect the deepest motor units and thus
is inherently limited to the more superficial parts of muscles, and because several muscles
and muscle groups display a more or less pronounced deep-to-superficial activation pattern, it
is suggested that fatigue and neuromuscular transmission failure can develop in the deeper
parts of the involved muscles during very strenuous low-load BFRRE without this being
readily detected in the sEMG recordings.

Given the similarities between BFRRE and low-load training to fatigue without
BFR8,19,314,333, and that some degree of intramuscular blood flow restriction can occur during
Accepted Article
free-flow (i.e. non-occluded ) resistance exercise already at relatively low loading intensities
(reviewed previously by Wernbom et al5), many points discussed in this review may to
varying extent apply also to low-to-moderate load resistance exercise performed to fatigue
without using concurrent external occlusion.
7.6. Consequences for the possible role of metabolic stress in BFRRE
Summarized from Section 6.5., it appears possible that in strenuous low-load ischemic
exercise, Pi splits may occur not only due to activation of type IIA fibres in addition to the
already active but fatiguing type I fibres, but also largely as a result of subsequent progressive
recruitment of type IIAX and/or type IIX fibres due to accumulating fatigue in type I and type
IIA fibres. In addition, non-localized 31P-MRS measurements must be viewed with caution,
as it cannot be ruled out that Pi splits are at least in part caused by signal contrasts between
active and less active or even inactive muscles.
Importantly, these and other uncertainties in the literature on 31P-MRS
measurements during BFRRE would seem to seriously limit the utility of 31P-MRS as a
stand-alone method to probe the mechanisms underpinning gains in muscle strength and
hypertrophy with this mode of exercise. Specifically, the strong correlations between acute
increases in Pi and training-induced gains in muscle area and strength that were reported by
Takada et al10 are open to question. Given that Pi increases concomittantly with decreases in
PCr158, and that gradually increasing fatigue is a major driver of the successive recruitment of
new motor units in low-load BFRRE, a greater elevation of Pi is likely to result not only from
more extensive depletion of PCr in each active myofibre (reflecting increased metabolic
stress) but also to stem from recruitment of a larger number of muscle fibres.

Therefore, the correlations between acute elevations in Pi and longer-term
increases in muscle CSA and strength in Takada et al10 may actually reflect the importance of
Accepted Article
activating a large number of muscle fibres (and thereby exposing them to significant
mechanical stress) to cause these gains, as opposed to a role of metabolic stress accumulation.
In other words, the relationships reported by Takada et al10 do not constitute firm evidence of
a role for metabolic stress per se in BFRRE, as the correlations may be partly or even fully
coincidental. However, we propose that the most likely scenario is a synergistic or additive
interaction between mechanical stimuli and metabolic stress and/or other types of ischemic
stimuli, as elaborated in detail above.
Acknowledgements
The authors gratefully thank Dr Kristoffer Cumming for valuable help with the pictures on
PAS staining of muscle fibres. Mathias Wernbom gratefully acknowledges the The Swedish
Research Council for Sport Science for post-doctoral fundings during the years 2013 to 2016,
and for supporting several research projects on BFRRE during the years 2004 to 2009.
Funding
No grants or funds were received to support this work.
Conflict of Interest
The authors declare no conflict of interest.

References
1. Takarada Y, Takazawa H, Sato Y, et al. Effects of resistance exercise combined with moderate
Accepted Article
vascular occlusion on muscular function in humans. J Appl Physiol. 2000: 88: 2097-2106.
2. Laurentino GC, Ugrinowitsch C, Roschel H, et al. Strength training with blood flow restriction
diminishes myostatin gene expression. Med Sci Sports Exerc. 2012; 44:406-412.
3. Ellefsen S, Hammarström D, Strand TA, et al. Blood flow-restricted strength training displays
high functional and biological efficacy in women: a within-subject comparison with high-load
strength training. Am J Physiol Regul Integr Comp Physiol. 2015;309:R767-779.
4. Lixandrão ME, Ugrinowitsch C, Berton R, et al. Magnitude of Muscle Strength and Mass
Adaptations Between High-Load Resistance Training Versus Low-Load Resistance Training
Associated with Blood-Flow Restriction: A Systematic Review and Meta-Analysis. Sports
Med. 2018;48:361-378.
5. Wernbom M, Augustsson J, Raastad T. Ischemic strength training: a low-load alternative to
heavy resistance exercise? Scand J Med Sci Sports. 2008; 18: 401-16.
6. Hughes L, Paton B, Rosenblatt B, et al. Blood flow restriction training in clinical
musculoskeletal rehabilitation: a systematic review and meta-analysis. Br J Sports Med.
2017;51:1003-1011.
7. Abe T, Loenneke JP, Fahs CA, et al. Exercise intensity and muscle hypertrophy in blood flow-
restricted limbs and non-restricted muscles: a brief review. Clin Physiol Funct Imaging.
2012;32:247-252.
8. Farup J, de Paoli F, Bjerg K, et al. Blood flow restricted and traditional resistance training
performed to fatigue produce equal muscle hypertrophy. Scand J Med Sci Sports.
2015;25:754-763.
9. Suga T, Okita K, Takada S, et al. Effect of multiple set on intramuscular metabolic stress
during low-intensity resistance exercise with blood flow restriction. Eur J Appl Physiol. 2012;
112: 3915-20.
10. Takada S, Okita K, Suga T, et al. Low-intensity exercise can increase muscle mass and
strength proportionally to enhanced metabolic stress under ischemic conditions. J Appl
Physiol. 2012;113:199-205.
11. Aagaard P, Andersen JL, Dyhre-Poulsen P, et al. A mechanism for increased contractile
strength of human pennate muscle in response to strength training: changes in muscle
architecture. J Physiol. 2001;534:613-623.
12. Kosek DJ, Kim JS, Petrella JK, et al. Efficacy of 3 days/wk resistance training on myofiber
hypertrophy and myogenic mechanisms in young vs. older adults. J Appl Physiol.
2006;101:531-544.
13. Suetta C, Andersen JL, Dalgas U, et al. Resistance training induces qualitative changes in
muscle morphology, muscle architecture, and muscle function in elderly postoperative
patients. J Appl Physiol. 2008;105:180-186.
14. Jakobsen MD, Sundstrup E, Randers MB, et al. The effect of strength training, recreational
soccer and running exercise on stretch-shortening cycle muscle performance during
countermovement jumping. Hum Mov Sci. 2012;31:970-986.
15. Faulkner JA, Claflin DR, McCully KK. Power output of fast and slow fibers from human
skeletal muscle. In: Human Muscle Power, edited by Jones NL, McCartney N and McComas
AJ, Champaign Illinois, Human Kinetics 1986, pp 81-94.
16. Nielsen JL, Aagaard P, Bech RD, et al. Proliferation of myogenic stem cells in human skeletal
muscle in response to low-load resistance training with blood flow restriction. J Physiol.
2012;590:4351-4361.
17. Moritani T, Sherman WM, Shibata M, et al. Oxygen availability and motor unit activity in
humans. Eur J Appl Physiol 1992;64:552-556.

18. Takarada Y, Nakamura Y, Aruga S, et al. Rapid increase in plasma growth hormone after low-
intensity resistance exercise with vascular occlusion. J Appl Physiol. 2000;88:61-65.
19. Wernbom M, Järrebring R, Andreasson MA, Augustsson J. Acute effects of blood flow
restriction on muscle activity and endurance during fatiguing dynamic knee extensions at
Accepted Article
low load. J Strength Cond Res. 2009;23:2389-2395.
20. Krustrup P, Söderlund K, Relu MU, et al. Heterogeneous recruitment of quadriceps muscle
portions and fibre types during moderate intensity knee-extensor exercise: effect of thigh
occlusion. Scand J Med Sci Sports. 2009;19:576-584.
21. Cumming KT, Paulsen G, Wernbom M, et al. Acute response and subcellular movement of
HSP27, αB-crystallin and HSP70 in human skeletal muscle after blood-flow-restricted low-
load resistance exercise. Acta Physiol. 2014; 211:634-646.
22. Jakobsgaard JE, Christiansen M, Sieljacks P, et al K. Impact of blood flow-restricted
bodyweight exercise on skeletal muscle adaptations. Clin Physiol Funct Imaging. 2018 Feb
15. doi: 10.1111/cpf.12509. [Epub ahead of print]
23. Bjørnsen T, Wernbom M, Kirketeig A, et al. Type 1 muscle fiber hypertrophy after blood
flow-restricted training in powerlifters. Med Sci Sports Exerc. 2019;51:288-298.
24. Bjørnsen T, Wernbom M, Løvstad AT, et al. Delayed myonuclear addition, myofiber
hypertrophy and increases in strength with high-frequency low-load blood flow restricted
training to volitional failure. J Appl Physiol. 2019;126:578-592.
25. Sumide T, Sakuraba K, Sawaki K, Ohmura H, Tamura Y. Effect of resistance exercise training
combined with relatively low vascular occlusion. J Sci Med Sport. 2009;12:107-112.
26. Shinohara M, Kouzaki M, Yoshihisa T, Fukunaga T. Efficacy of tourniquet ischemia for
strength training with low resistance. Eur J Appl Physiol. 1998;77:189-191.
27. Moritani T. Motor unit and motoneurone excitability during explosive movement. In:
Strength and Power in Sport, 2nd ed., edited by Paavo Komi, Oxford, Blackwell Science,
2003, pp 27-49.
28. Duchateau J, Semmler JG, Enoka RM. Training adaptations in the behavior of human motor
units. J Appl Physiol. 2006;101:1766-1775.
29. Chalmers GR. Can fast-twitch muscle fibres be selectively recruited during lengthening
contractions? Review and applications to sport movements. Sports Biomech. 2008;7:137-
157.
30. Heckman CJ, Enoka RM. Motor unit. Compr Physiol. 2012;2:2629-2682.
31. Burke RE, Levine DN, Zajac FE 3rd. Mammalian motor units: physiological-histochemical
correlation in three types in cat gastrocnemius. Science. 1971;174(4010):709-12.
32. Burke RE, Levine DN, Tsairis P, Zajac FE 3rd. Physiological types and histochemical profiles in
motor units of the cat gastrocnemius. J Physiol. 1973;234:723-748.
33. Edström L, Grimby L. Effect of exercise on the motor unit. Muscle Nerve. 1986;9:104-126.
34. McDonagh JC, Binder MD, Reinking RM, Stuart DG. Tetrapartite classification of motor units
of cat tibialis posterior. J Neurophysiol. 1980;44:696-712.
35. Burke RE. The structure and function of motor units. In: Disorders of Voluntary Muscle, 7th
ed., edited by Karpati G, Hilton D, Griggs RC, Cambridge University Press, 2001, pp 3-21.
36. Burke RE. Some unresolved issues in motor unit research. In: Sensorimotor Control of
Movement and Posture, edited by Gandevia S, Proske U, Stuart DG, Kluwer
Academic/Plenum Press, 2002, pp 171-178.
37. Garnett RA, O'Donovan MJ, Stephens JA, Taylor A. Motor unit organization of human medial
gastrocnemius. J Physiol. 1979;287:33-43.
38. Buchthal F, Schmalbruch H. Contraction times and fibre types in intact human muscle. Acta
Physiol Scand. 1970;79:435-452.
39. Ennion S, Sant'ana Pereira J, Sargeant AJ, et al. Characterization of human skeletal muscle
fibres according to the myosin heavy chains they express. J Muscle Res Cell Motil.
1995;16:35-43.

40. Sant'Ana Pereira JA, Sargeant AJ, Rademaker AC, et al. Myosin heavy chain isoform
expression and high energy phosphate content in human muscle fibres at rest and post-
exercise. J Physiol. 1996;496:583-588.
41. Staron RS, Herman JR, Schuenke MD. Misclassification of hybrid fast fibers in resistance-
Accepted Article
trained human skeletal muscle using histochemical and immunohistochemical methods. J
Strength Cond Res. 2012;26:2616-2622.
42. Sale DG. Influence of exercise and training on motor unit activation. Exerc Sport Sci Rev.
1987;15:95-151.
43. MacIntosh BR, Gardiner PF, McComas AJ. Skeletal Muscle: Form and Function, 2nd editon.
Champaign, Human Kinetics, 2006, pp 195-207.
44. Staron RS. Human skeletal muscle fiber types: delineation, development, and distribution.
Can J Appl Physiol. 1997;22:307-27.
45. Andersen JL, Klitgaard H, Saltin B. Myosin heavy chain isoforms in single fibres from m.
vastus lateralis of sprinters: influence of training. Acta Physiol Scand. 1994;151:135-142.
46. Andersen JL, Aagaard P. Myosin heavy chain IIX overshoot in human skeletal muscle. Muscle
Nerve. 2000;23:1095-1104.
47. Klitgaard H, Zhou M, Richter EA. Myosin heavy chain composition of single fibres from m.
biceps brachii of male body builders. Acta Physiol Scand. 1990;140:175-180.
48. Sant'ana Pereira JA, Wessels A, Nijtmans L, et al. New method for the accurate
characterization of single human skeletal muscle fibres demonstrates a relation between
mATPase and MyHC expression in pure and hybrid fibre types. J Muscle Res Cell Motil.
1995;16:21-34.
49. Karatzaferi C, de Haan A, Ferguson RA, et al. Phosphocreatine and ATP content in human
single muscle fibres before and after maximum dynamic exercise. Pflügers Arch.
2001;442:467-474.
50. Prats C, Gomez-Cabello A, Nordby P, et al. An optimized histochemical method to assess
skeletal muscle glycogen and lipid stores reveals two metabolically distinct populations of
type I muscle fibers. PLoS One. 2013 30;8(10):e77774.
51. Essén B, Jansson E, Henriksson J, et al. Metabolic characteristics of fibre types in human
skeletal muscle. Acta Physiol Scand. 1975;95:153-65.
52. Greenhaff PL, Söderlund K, Ren JM, Hultman E. Energy metabolism in single human muscle
fibres during intermittent contraction with occluded circulation. J Physiol. 1993;460:443-53.
53. Greenhaff PL, Nevill ME, Soderlund K, et al. The metabolic responses of human type I and II
muscle fibres during maximal treadmill sprinting. J Physiol. 1994;478:149-55.
54. Henneman E. Relation between size of neurons and their susceptibility to discharge. Science.
1957;126:1345-1347.
55. Henneman E, Somjen G, Carpenter DO. Functional significance of cell size in spinal
motoneurons. J Neurophysiol. 1965;28:560-580.
56. Milner-Brown HS, Stein RB, Yemm R. The orderly recruitment of human motor units during
voluntary isometric contractions. J Physiol. 1973; 230:359-370.
57. Freund HJ, Büdingen HJ, Dietz V. Activity of single motor units from human forearm muscles
during voluntary isometric contractions. J Neurophysiol. 1975;38:933-46.
58. Monster AW, Chan H. Isometric force production by motor units of extensor digitorum
communis muscle in man. J Neurophysiol. 1977;40:1432-43.
59. Stephens JA, Usherwood TP. The mechanical properties of human motor units with special
reference to their fatiguability and recruitment threshold. Brain Res. 1977;125:91-97.
60. Desmedt JE, Godaux E. Ballistic contractions in man: characteristic recruitment pattern of
single motor units of the tibialis anterior muscle. J Physiol. 1977;264:673-693.
61. Garnett R, Stephens JA. Changes in the recruitment threshold of motor units produced by
cutaneous stimulation in man. J Physiol. 1981;311:463-473.

62. Calancie B, Bawa P. Recruitment order of motor units during the stretch reflex in man. Brain
Res. 1984;292:176-178.
63. Bawa P, Lemon RN. Recruitment of motor units in response to transcranial magnetic
stimulation in man. J Physiol. 1993;471:445-464.
Accepted Article
64. Enoka RM, Fuglevand AJ. Motor unit physiology: some unresolved issues. Muscle Nerve.
2001;24:4-17.
65. Hannerz J. Discharge properties of motor units in relation to recruitment order in voluntary
contraction. Acta Physiol Scand. 1974;91:374-385.
66. Kukulka CG, Clamann HP. Comparison of the recruitment and discharge properties of motor
units in human brachial biceps and adductor pollicis during isometric contractions. Brain Res.
1981;219:45-55.
67. Christie A, Greig Inglis J, Kamen G, Gabriel DA. Relationships between surface EMG variables
and motor unit firing rates. Eur J Appl Physiol. 2009;107:177-185.
68. Oya T, Riek S, Cresswell AG. Recruitment and rate coding organisation for soleus motor units
across entire range of voluntary isometric plantar flexions. J Physiol. 2009;587:4737-4748.
69. De Luca CJ, Hostage EC. Relationship between firing rate and recruitment threshold of
motoneurons in voluntary isometric contractions. J Neurophysiol. 2010;104:1034-1046.
70. Bawa PN, Jones KE, Stein RB. Assessment of size ordered recruitment. Front Hum Neurosci.
2014;8:532.
71. Grimby L, Hannerz J. Firing rate and recruitment order of toe extensor motor units in
different modes of voluntary conraction. J Physiol. 1977;264:865-879.
72. Nardone A, Romanò C, Schieppati M. Selective recruitment of high-threshold human motor
units during voluntary isotonic lengthening of active muscles. J Physiol. 1989;409:451-471.
73. Grimby L. Single motor unit discharge during voluntary contraction and locomotion. In:
Human Muscle Power, edited by Jones NL, McCartney N and McComas AJ, Champaign
Illinois, Human Kinetics 1986, pp 111-130.
74. Hodges PW, Tucker K. Moving differently in pain: a new theory to explain the adaptation to
pain. Pain. 2011;152(3 Suppl):S90-8.
75. Kanda K, Burke RE, Walmsley B. Differential control of fast and slow twitch motor units in
the decerebrate cat. Exp Brain Res. 1977;29:57-74.
76. Masakado Y, Kamen G, De Luca CJ. Effects of percutaneous stimulation on motor unit firing
behavior in man. Exp Brain Res. 1991;86:426-32.
77. Binder MD, Heckman CJ, Powers RK. The physiological control of motoneuron activity. In:
Handbook of physiology. Exercise: regulation and integration of multiple systems, edited by
Rowell LB and Shepherd JT, American Physiological Society 1996, 12, pp 1-53.
78. Cope TC, Clark BD. Are there important exceptions to the size principle of α-motoneurone
recruitment? In: Alpha and gamma motor systems, edited by Taylor A, Gladden MH, and
Durban R, Springer, Boston, MA, 1995, pp. 71-78.
79. Bawa P, Jones KE. Do lengthening contractions represent a case of reversal in recruitment
order? Prog Brain Res. 1999;123:215-220.
80. Tucker K, Butler J, Graven-Nielsen T, et al. Motor unit recruitment strategies are altered
during deep-tissue pain. J Neurosci. 2009;29:10820-10826.
81. Tucker KJ, Hodges PW. Motoneurone recruitment is altered with pain induced in non-
muscular tissue. Pain. 2009;141:151-155.
82. Edwards RH, Young A, Hosking GP, Jones DA. Human skeletal muscle function: description of
tests and normal values. Clin Sci Mol Med. 1977;52:283-290.
83. Hultman E, Sjöholm H, Jäderholm-Ek I, Krynicki J. Evaluation of methods for electrical
stimulation of human skeletal muscle in situ. Pflugers Arch. 1983;398:139-141.
84. Gregory CM, Dixon W, Bickel CS. Impact of varying pulse frequency and duration on muscle
torque production and fatigue. Muscle Nerve. 2007;35:504-509.

85. Eberstein A, Goodgold J. Slow and fast twitch fibers in human skeletal muscle. Am J Physiol.
1968;215:535-541.
86. Moulds RF, Young A, Jones DA, Edwards RH. A study of the contractility, biochemistry and
morphology of an isolated preparation of human skeletal muscle. Clin Sci Mol Med.
Accepted Article
1977;52:291-297.
87. Bigland-Ritchie B, Rice CL, Garland SJ, Walsh ML. Task-dependent factors in fatigue of human
voluntary contractions. In: Fatigue. Edited by Gandevia SC, Enoka RM, McComas AJ et al.,
Springer, Boston, MA, 1995. pp. 361-380.
88. Desmedt JE, Godaux E. Ballistic contractions in fast or slow human muscles: discharge
patterns of single motor units. J Physiol. 1978;285:185-196.
89. Van Cutsem M, Duchateau J, Hainaut K. Changes in single motor unit behaviour contribute
to the increase in contraction speed after dynamic training in humans. J Physiol.
1998;513:295-305.
90. Abbate F, Bruton JD, De Haan A, Westerblad H. Prolonged force increase following a high-
frequency burst is not due to a sustained elevation of [Ca2+]i. Am J Physiol Cell Physiol.
2002;283:C42-47.
91. Cheng AJ, Place N, Bruton JD, et al. Doublet discharge stimulation increases sarcoplasmic
reticulum Ca2+ release and improves performance during fatiguing contractions in mouse
muscle fibres. J Physiol. 2013;591:3739-3748.
92. Kamen G, Knight CA. Training-related adaptations in motor unit discharge rate in young and
older adults. J Gerontol A Biol Sci Med Sci. 2004;59:1334-1338.
93. Christie A, Kamen G. Short-term training adaptations in maximal motor unit firing rates and
afterhyperpolarization duration.Muscle Nerve. 2010;41:651-660.
94. Knight CA, Kamen G. Relationships between voluntary activation and motor unit firing rate
during maximal voluntary contractions in young and older adults. Eur J Appl Physiol.
2008;103:625-630.
95. Woods JJ, Furbush F, Bigland-Ritchie B. Evidence for a fatigue-induced reflex inhibition of
motoneuron firing rates. J Neurophysiol. 1987;58:125-137.
96. Strojnik V. Muscle activation level during maximal voluntary effort. Eur J Appl Physiol Occup
Physiol. 1995;72:144-149.
97. Behm DG, Whittle J, Button D, Power K. Intermuscle differences in activation. Muscle Nerve.
2002;25:236-243.
98. Babault N, Pousson M, Michaut A, Ballay Y, Hoecke JV. EMG activity and voluntary activation
during knee-extensor concentric torque generation. Eur J Appl Physiol. 2002;86:541-547.
99. Beltman JG, Sargeant AJ, van Mechelen W, de Haan A. Voluntary activation level and muscle
fiber recruitment of human quadriceps during lengthening contractions. J Appl Physiol.
2004;97:619-626.
100. Place N, Casartelli N, Glatthorn JF, Maffiuletti NA. Comparison of quadriceps
inactivation between nerve and muscle stimulation. Muscle Nerve. 2010;42:894-900.
101. Van Leeuwen DM, De Ruiter CJ, De Haan A. Effect of stimulation intensity on
assessment of voluntary activation. Muscle Nerve. 2012;45:841-848.
102. Pucci AR, Griffin L, Cafarelli E. Maximal motor unit firing rates during isometric
resistance training in men. Exp Physiol. 2006;91:171-178.
103. Grimby L, Hannerz J, Hedman B. Contraction time and voluntary discharge properties
of individual short toe extensor motor units in man. J Physiol. 1979; 289:191-201.
104. Bellemare F, Woods JJ, Johansson R, Bigland-Ritchie B. Motor-unit discharge rates in
maximal voluntary contractions of three human muscles. J Neurophysiol. 1983;50:1380-
1392.
105. Grimby L, Hannerz J, Hedman B. The fatigue and voluntary discharge properties of
single motor units in man. J Physiol. 1981;316:545-554.

106. Kanosue K, Yoshida M, Akazawa K, Fujii K. The number of active motor units and their
firing rates in voluntary contraction of human brachialis muscle. Jpn J Physiol. 1979;29:427-
443.
107. Harwood B, Davidson AW, Rice CL. Motor unit discharge rates of the anconeus muscle
Accepted Article
during high-velocity elbow extensions. Exp Brain Res. 2011;208:103-113.
108. Hannerz J, Grimby L. The afferent influence on the voluntary firing range of individual
motor units in man. Muscle Nerve. 1979;2:414-422.
109. Gydikov A, Kosarov D. Some features of different motor units in human biceps brachii.
Pflugers Arch. 1974;347:75-88.
110. De Luca CJ, LeFever RS, McCue MP, Xenakis AP. Behaviour of human motor units in
different muscles during linearly varying contractions. J Physiol. 1982;329:113-128.
111. De Luca CJ, Erim Z. Common drive of motor units in regulation of muscle force. Trends
Neurosci. 1994;17:299-305.
112. Erim Z, De Luca CJ, Mineo K, Aoki T. Rank-ordered regulation of motor units. Muscle
Nerve. 1996;19:563-573.
113. De Luca CJ, Contessa P. Hierarchical control of motor units in voluntary contractions. J
Neurophysiol. 2012;107:178-195.
114. Person RS, Kudina LP. Discharge frequency and discharge pattern of human motor
units during voluntary contraction of muscle. Electroencephalogr Clin Neurophysiol.
1972;32:471-483.
115. Tanji J, Kato M. Recruitment of motor units in voluntary contraction of a finger muscle
in man. Exp Neurol. 1973;40:759-770.
116. Enoka RM, Duchateau J. Rate coding and the control of muscle force. Cold Spring Harb
Perspect Med. 2017;7(10): pii: a029702.
117. Piotrkiewicz M, Türker K. Onion skin or common drive? Front Cell Neurosci 2017;11:2.
118. Kosarov D, Gydikov A. Dependence of the discharge frequency of motor units in
different human muscles upon the level of the isometric muscle tension. Electromyogr Clin
Neurophysiol. 1976;16:293-306.
119. Moritz CT, Barry BK, Pascoe MA, Enoka RM. Discharge rate variability influences the
variation in force fluctuations across the working range of a hand muscle. J Neurophysiol.
2005;93:2449-2459.
120. Barry BK, Pascoe MA, Jesunathadas M, Enoka RM. Rate coding is compressed but
variability is unaltered for motor units in a hand muscle of old adults. J Neurophysiol.
2007;97:3206-3218.
121. Grimby L, Hannerz J. Recruitment order of motor units on voluntary contraction:
changes induced by proprioceptive afferent activity. J Neurol Neurosurg Psychiatry.
1968;31:565-573.
122. Gatev P, Ivanova T, Gantchev GN. Changes in the firing pattern of high-threshold
motor units due to fatigue. Electromyogr Clin Neurophysiol. 1986;26:83-93.
123. Warmolts JR, Engel WK. Open-biopsy electromyography. I. Correlation of motor unit
behavior with histochemical muscle fiber type in human limb muscle. Arch Neurol.
1972;27:512-517.
124. Warmolts JR, Engel WK. Correlation of motor unit behavior with histochemical
myofiber type in humans by open-biopsy electromyography. In: New Concepts of the Motor
Unit, Neuromuscular Disorders, Electromyographic Kinesiology. Karger Publishers, 1973. p.
35-40.
125. Grimby L. Firing properties of single human motor units during locomotion. J Physiol.
1984;346:195-202.
126. Søgaard K, Christensen H, Fallentin N, et al. Motor unit activation patterns during
concentric wrist flexion in humans with different muscle fibre composition. Eur J Appl Physiol
Occup Physiol. 1998;78:411-416.

127. Grimby L, Hannerz J, Borg J, Hedman B. Firing properties of single human motor units
on maintained maximal voluntary effort. In: Human Muscle Fatigue: Physiological
Mechanisms. Chichester, UK: John Wiley & Sons, Ltd., 1981. pp 157-177.
128. Van Cutsem M, Duchateau J. Preceding muscle activity influences motor unit
Accepted Article
discharge and rate of torque development during ballistic contractions in humans. J Physiol.
2005;562:635-644.
129. Harwood B, Choi I, Rice CL. Reduced motor unit discharge rates of maximal velocity
dynamic contractions in response to a submaximal dynamic fatigue protocol. J Appl Physiol.
2012;113:1821-1830.
130. Sawczuk A, Powers RK, Binder MD. Intrinsic properties of motoneurons. In: Fatigue.
Springer, Boston, MA, 1995. p. 123-134.
131. Kernell D, Eerbeek O, Verhey BA. Relation between isometric force and stimulus rate
in cat's hindlimb motor units of different twitch contraction time. Exp Brain Res.
1983;50:220-227.
132. Zengel JE, Reid SA, Sypert GW, Munson JB. Membrane electrical properties and
prediction of motor-unit type of medial gastrocnemius motoneurons in the cat. J
Neurophysiol. 1985;53:1323-1344.
133. Gardiner PF. Physiological properties of motoneurons innervating different muscle
unit types in rat gastrocnemius. J Neurophysiol. 1993;69:1160-1170.
134. Bigland-Ritchie BR, Dawson NJ, Johansson RS, Lippold OC. Reflex origin for the slowing
of motoneurone firing rates in fatigue of human voluntary contractions. J Physiol.
1986;379:451-9.
135. Taylor JL, Amann M, Duchateau J, et al. Neural contributions to muscle fatigue: from
the brain to the muscle and back Again. Med Sci Sports Exerc. 2016;48:2294-2306.
136. Amann M, Sidhu SK, Weavil JC, et al. Autonomic responses to exercise: group III/IV
muscle afferents and fatigue. Auton Neurosci. 2015;188:19-23.
137. Windhorst U, Kirmayer D, Soibelman F, et al. Effects of neurochemically excited group
III-IV muscle afferents on motoneuron afterhyperpolarization. Neuroscience. 1997;76:915-
929.
138. Martin PG, Weerakkody N, Gandevia SC, Taylor JL. Group III and IV muscle afferents
differentially affect the motor cortex and motoneurones in humans. J Physiol.
2008;586:1277-1289.
139. Garland SJ, Griffin L, Ivanova T. Motor unit discharge rate is not associated with
muscle relaxation time in sustained submaximal contractions in humans. Neurosci Lett.
1997;239:25-28.
140. Bonde-Petersen F, Mork AL, Nielsen E. Local muscle blood flow and sustained
contractions of human arm and back muscles. Eur J Appl Physiol Occup Physiol. 1975;34:43-
50.
141. Griffin L, Garland SJ, Ivanova T, Hughson RL. Blood flow in the triceps brachii muscle in
humans during sustained submaximal isometric contractions. Eur J Appl Physiol.
2001;84:432-437.
142. Freund PR, Hobbs SF, Rowell LB. Cardiovascular responses to muscle ischemia in man–
dependency on muscle mass. J Appl Physiol Respir Environ Exerc Physiol. 1978;45:762-767.
143. Mitchell JH, Payne FC, Saltin B, Schibye B. The role of muscle mass in the
cardiovascular response to static contractions. J Physiol. 1980;309:45-54.
144. Seals DR. Influence of muscle mass on sympathetic neural activation during isometric
exercise. J Appl Physiol. 1989;67:1801-1806.
145. Seals DR. Influence of active muscle size on sympathetic nerve discharge during
isometric contractions in humans. J Appl Physiol. 1993;75:1426-1431.

146. Matkowski B, Place N, Martin A, Lepers R. Neuromuscular fatigue differs following
unilateral vs bilateral sustained submaximal contractions. Scand J Med Sci Sports.
2011;21:268-276.
147. Rossman MJ, Venturelli M, McDaniel J, et al. Muscle mass and peripheral fatigue: a
Accepted Article
potential role for afferent feedback? Acta Physiol. 2012;206:242-250.
148. Rossman MJ, Garten RS, Venturelli M, et al. The role of active muscle mass in
determining the magnitude of peripheral fatigue during dynamic exercise. Am J Physiol
Regul Integr Comp Physiol. 2014;306:R934-340.
149. Dietz V. Analysis of the electrical muscle activity during maximal contraction and the
influence of ischaemia. J Neurol Sci. 1978;37:187-197.
150. Carvalho AJ, McKee NH. Active and passive forces during ischemia and reperfusion of
rat fast and slow twitch skeletal muscles. Adv Exp Med Biol. 1992;311:365-368.
151. Carvalho AJ, McKee NH, Green HJ. Metabolic and contractile responses of fast- and
slow-twitch rat skeletal muscles to ischemia. Can J Physiol Pharmacol. 1996;74:1333-1341.
152. Gossen ER, Ivanova TD, Garland SJ. Ischemia sensitivity and motoneuron
afterhyperpolarization in human motor units. Muscle Nerve. 2004;30:195-201.
153. Folkow B, Halicka H. A comparison between “red” and “white” muscle with respect to
blood supply, capillary surface area and oxygen uptake during rest and exercise. Microvasc
Res. 1968;1:1-14.
154. Sahlin K, Edström L, Sjöholm H. Force, relaxation and energy metabolism of rat soleus
muscle during anaerobic contraction. Acta Physiol Scand. 1987;129:1-7.
155. Gollnick PD, Karlsson J, Piehl K, Saltin B. Selective glycogen depletion in skeletal
muscle fibres of man following sustained contractions. J Physiol. 1974;241:59-67.
156. Murthy G, Hargens AR, Lehman S, Rempel DM. Ischemia causes muscle fatigue. J
Orthop Res. 2001; 19:436-440.
157. Bylund-Fellenius AC, Walker PM, Elander A, et al. Energy metabolism in relation to
oxygen partial pressure in human skeletal muscle during exercise. Biochem J. 1981;200:247-
255.
158. Allen DG, Lamb GD, Westerblad H. Skeletal muscle fatigue: cellular mechanisms.
Physiol Rev. 2008;88:287-332.
159. Allen DG, Trajanovska S. The multiple roles of phosphate in muscle fatigue. Front
Physiol. 2012;3:463.
160. Karatzaferi C, Franks-Skiba K, Cooke R. Inhibition of shortening velocity of skinned
skeletal muscle fibers in conditions that mimic fatigue. Am J Physiol Regul Integr Comp
Physiol. 2008;294:R948-955.
161. Nelson CR, Debold EP, Fitts RH. Phosphate and acidosis act synergistically to depress
peak power in rat muscle fibers. Am J Physiol Cell Physiol. 2014;307:C939-950.
162. Debold EP. Decreased myofilament calcium sensitivity plays a significant role in
muscle fatigue. Exerc Sport Sci Rev. 2016;44:144-149.
163. Hultman E, Del Canale S, Sjöholm H. Effect of induced metabolic acidosis on
intracellular pH, buffer capacity and contraction force of human skeletal muscle. Clin Sci.
1985;69:505-510.
164. Hultman E, Sjöhom H, Sahlin K, Edström L. Glycolytic and oxidative energy metabolism
and contraction characteristics of intact human muscle. In: Human Muscle Fatigue:
Physiological Mechanisms. Chichester, UK: John Wiley & Sons, Ltd., 1981. pp 19-40.
165. Hultman E, Sjöholm H. Electromyogram, force and relaxation time during and after
continuous electrical stimulation of human skeletal muscle in situ. J Physiol. 1983;339:33-40.
166. Hultman E, Sjöholm H. Biochemical causes of fatigue. In: Human Muscle Power. Jones
NL, McCartney N, McComas AJ (Editors). Champaign, Illinois: Human Kinetics Publishers
1986, pp 215-238.

167. Söderlund K, Hultman E. ATP content in single fibres from human skeletal muscle after
electrical stimulation and during recovery. Acta Physiol Scand. 1990;139:459-466.
168. Söderlund K, Greenhaff PL, Hultman E. Energy metabolism in type I and type II human
muscle fibres during short term electrical stimulation at different frequencies. Acta
Accepted Article
Physiologica Scandinavica. 1992;144:15-22.
169. Söderlund K, Hultman E. ATP and phosphocreatine changes in single human muscle
fibers after intense electrical stimulation. Am J Physiol 1991;261:E737-741.
170. Tonkonogi M, Walsh B, Tiivel T, et al. Mitochondrial function in human skeletal muscle
is not impaired by high intensity exercise. Pflugers Arch. 1999;437:562-568.
171. Tesch PA, Thorsson A, Fujitsuka N. Creatine phosphate in fiber types of skeletal
muscle before and after exhaustive exercise. J Appl Physiol. 1989;66:1756-1759.
172. Colliander EB, Dudley GA, Tesch PA. Skeletal muscle fiber type composition and
performance during repeated bouts of maximal, concentric contractions. Eur J Appl Physiol
Occup Physiol. 1988;58:81-86.
173. Tesch PA. Muscle fatigue in man. With special reference to lactate accumulation
during short term intense exercise. Acta Physiologica Scandinavica. Supplementum,
1980;480:1-40.
174. Sun QA, Hess DT, Nogueira L, et al. Oxygen-coupled redox regulation of the skeletal
muscle ryanodine receptor-Ca2+ release channel by NADPH oxidase 4. Proc Natl Acad Sci U S
A. 2011;108:16098-16103.
175. Loureiro AC, do Rêgo-Monteiro IC, Louzada RA, et al. Differential expression of NADPH
oxidases depends on skeletal muscle fiber type in rats. Oxid Med Cell Longev.
2016;2016:6738701.
176. Edwards RH, Toescu V, Gibson H. Historical perspective: a framework for interpreting
pathobiological ideas on human muscle fatigue. In: Fatigue. Edited by Gandevia SC, Enoka
RM, McComas AJ et al., Springer, Boston, MA, 1995. pp. 481-494.
177. Saugen E, Vøllestad NK, Gibson H, et al. Dissociation between metabolic and
contractile responses during intermittent isometric exercise in man. Exp Physiol.
1997;82:213-26.
178. Wernbom M, Paulsen G, Nilsen TS, et al. Contractile function and sarcolemmal
permeability after acute low-load resistance exercise with blood flow restriction. Eur J Appl
Physiol. 2012; 112: 2051-2063.
179. Sieljacks P, Matzon A, Wernbom M, et al. Muscle damage and repeated bout effect
following blood flow restricted exercise. Eur J Appl Physiol. 2016;116:513-525.
180. Yasuda T, Brechue WF, Fujita T, et al. Muscle activation during low-intensity muscle
contractions with varying levels of external limb compression. J Sports Sci Med. 2008;7:467-
474.
181. Yasuda T, Brechue WF, Fujita T, et al. Muscle activation during low-intensity muscle
contractions with restricted blood flow. J Sports Sci. 2009;27:479-489.
182. Moritani T, Muro M, Kijima A, et al. Electromechanical changes during electrically
induced and maximal voluntary contractions: surface and intramuscular EMG responses
during sustained maximal voluntary contraction. Exp Neurol. 1985;88:484-499.
183. Sadamoto T, Bonde-Petersen F, Suzuki Y. Skeletal muscle tension, flow, pressure, and
EMG during sustained isometric contractions in humans. Eur J Appl Physiol Occup Physiol.
1983;51:395-408.
184. Sjøgaard G, Savard G, Juel C. Muscle blood flow during isometric activity and its
relation to muscle fatigue. Eur J Appl Physiol Occup Physiol. 1988;57:327-335.
185. Cupido CM, Galea V, McComas AJ. Potentiation and depression of the M wave in
human biceps brachii. J Physiol. 1996;491:541-550.

186. Galea V. Electrical characteristics of human ankle dorsi- and plantar-flexor muscles.
Comparative responses during fatiguing stimulation and recovery. Eur J Appl Physiol.
2001;85:130-140.
187. Leonard CT, Kane J, Perdaems J, et al. Neural modulation of muscle contractile
Accepted Article
properties during fatigue: afferent feedback dependence. Electroencephalogr Clin
188. Hicks A, McComas AJ. Increased sodium pump activity following repetitive stimulation
of rat soleus muscles. J Physiol. 1989;414:337-349.
189. Rodriguez-Falces J, Place N. Determinants, analysis and interpretation of the muscle
compound action potential (M wave) in humans: implications for the study of muscle
fatigue. Eur J Appl Physiol. 2018;118:501-521.
190. Bigland-Ritchie B. EMG and fatigue of human voluntary and stimulated contractions.
In: Human Muscle Fatigue: Physiological Mechanisms. Chichester, UK: John Wiley & Sons,
Ltd., 1981. pp 130-156.
191. Dahlbäck LO, Ekstedt J, Stålberg E. Ischemic effects on impulse transmission to muscle
fibers in man. Electroencephalogr Clin Neurophysiol. 1970;29:579-591.
192. Sieck GC, Prakash YS. Fatigue at the neuromuscular junction. In: Fatigue. Edited by
Gandevia SC, Enoka RM, McComas AJ et al., Springer, Boston, MA, 1995. pp. 83-100.
193. Sandercock TG, Faulkner JA, Albers JW, Abbrecht PH. Single motor unit and fiber
action potentials during fatigue. J Appl Physiol. 1985;58:1073-1079.
194. Abramson DI, Rickert BL, Alexis JT, et al. Effect of repeated periods of ischemia on
motor nerve conduction velocity in forearm. J Appl Physiol. 1971;30:636-642.
195. Mittal P, Shenoy S, Sandhu JS. Effect of different cuff widths on the motor nerve
conduction of the median nerve: an experimental study. J Orthop Surg Res. 2008;3:1.
196. Desaulniers P, Lavoie PA, Gardiner PF. Incomplete recovery of endplate potential
amplitude while intermittently activating rat soleus neuromuscular junctions in situ. Muscle
Nerve. 2002;26:810-816.
197. Clausen T, Overgaard K, Nielsen OB. Evidence that the Na+-K+ leak/pump ratio
contributes to the difference in endurance between fast- and slow-twitch muscles. Acta
Physiol Scand. 2004;180:209-216.
198. Jansson E, Dudley GA, Norman B, Tesch PA. Relationship of recovery from intensive
exercise to the oxidative potential of skeletal muscle. Acta Physiol Scand. 1990;139:147-152.
199. Sjøgaard G, Houston ME, Nygaard E, Saltin B. Subgrouping of fast twitch fibres in
skeletal muscles of man. A critical appraisal. Histochemistry. 1978;58:79-87.
200. Staron RS, Hikida RS, Hagerman FC. Reevaluartion of human muscle fast-twitch
subtypes: evidence for a continuum. Histochemistry. 1983;78:33-39.
201. Staron RS, Hikida RS, Hagerman FC, et al. Human skeletal muscle fiber type
adaptability to various workloads. J Histochem Cytochem. 1984;32:146-152.
202. Essén-Gustavsson B, Henriksson J. Enzyme levels in pools of microdissected human
muscle fibres of identified type. Adaptive response to exercise. Acta Physiol Scand.
1984;120:505-515.
203. Sahlin K, Ren JM. Relationship of contraction capacity to metabolic changes during
recovery from a fatiguing contraction. J Appl Physiol. 1989;67:648-654.
204. Harris RC, Edwards RH, Hultman E, et al. The time course of phosphorylcreatine
resynthesis during recovery of the quadriceps muscle in man. Pflügers Arch. 1976;367:137-
142.
205. Umbel JD, Hoffman RL, Dearth DJ, et al. Delayed-onset muscle soreness induced by
low-load blood flow-restricted exercise. Eur J Appl Physiol. 2009 Dec; 107(6): 687-95.
206. Edström L, Kugelberg E. Histochemical composition, distribution of fibres and
fatiguability of single motor units. Anterior tibial muscle of the rat. J Neurol Neurosurg
Psychiatry. 1968;424-433.

207. Gollnick PD, Armstrong RB, Sembrowich WL, et al. Glycogen depletion pattern in
human skeletal muscle fibers after heavy exercise. J Appl Physiol. 1973;34:615-618.
208. Gollnick PD, Armstrong RB, Saubert CW 4th, et al. Glycogen depletion patterns in
human skeletal muscle fibers during prolonged work. Pflugers Arch. 1973;344:1-12.
Accepted Article
209. Gollnick PD, Piehl K, Saltin B. Selective glycogen depletion pattern in human muscle
fibres after exercise of varying intensity and at varying pedalling rates. J Physiol. 1974: 241:
45-57.
210. Vøllestad NK, Vaage O, Hermansen L. Muscle glycogen depletion patterns in type I and
subgroups of type II fibres during prolonged severe exercise in man.
Acta Physiol Scand, 1984: 122: 433-441.
211. Vøllestad NK, Blom PC. Effect of varying exercise intensity on glycogen depletion in
human muscle fibres. Acta Physiol Scand, 1985: 125: 395-405.
212. Vøllestad NK, Tabata I, Medbø JI. Glycogen breakdown in different human muscle
fibre types during exhaustive exercise of short duration. Acta Physiol Scand. 1992;144:135-
141.
213. Andersen JL, Gruschy-Knudsen T. Rapid switch-off of the human myosin heavy chain
IIX gene after heavy load muscle contractions is sustained for at least four days. Scand J Med
Sci Sports. 2018;28:371-380.
214. Robergs RA, Pearson DR, Costill DL, et al. Muscle glycogenolysis during differing
intensities of weight-resistance exercise. J Appl Physiol. 1991;70:1700-1706.
215. Pascoe DD, Gladden LB. Muscle glycogen resynthesis after short term, high intensity
exercise and resistance exercise. Sports Med. 1996;21:98-118.
216. Tesch PA, Ploutz-Snyder L., Yström L, et al. Skeletal muscle glycogen loss evoked by
resistance exercise. J Strength Cond Res. 1998:12:67-73.
217. Schmitz JP, Groenendaal W, Wessels B, et al. Combined in vivo and in silico
investigations of activation of glycolysis in contracting skeletal muscle. Am J Physiol Cell
Physiol. 2013;304:C180-193.
218. Jin ES, Sherry AD, Malloy CR. Evidence for reverse flux through pyruvate kinase in
skeletal muscle. Am J Physiol Endocrinol Metab. 2009;296:E748-757.
219. Jin ES, Sherry AD, Malloy CR. Lactate Contributes to Glyceroneogenesis and
Glyconeogenesis in Skeletal Muscle by Reversal of Pyruvate Kinase. J Biol Chem.
2015;290:30486-30497.
220. Lund J, Aas V, Tingstad RH, et al. Utilization of lactic acid in human myotubes and
interplay with glucose and fatty acid metabolism. Sci Rep. 2018;8:9814.
221. McLane JA, Holloszy JO. Glycogen synthesis from lactate in the three types of skeletal
muscle. J Biol Chem. 1979;254:6548-6553.
222. Beltman JG, Sargeant AJ, Haan H, et al. Changes in PCr/Cr ratio in single characterized
muscle fibre fragments after only a few maximal voluntary contractions in humans. Acta
Physiol Scand. 2004;180:187-193.
223. Beltman JG, de Haan A, Haan H, et al. Metabolically assessed muscle fibre recruitment
in brief isometric contractions at different intensities. Eur J Appl Physiol. 2004;92:485-492.
224. Tupling AR, Bombardier E, Stewart RD, et al. Muscle fiber type-specific response of
Hsp70 expression in human quadriceps following acute isometric exercise. J Appl Physiol.
2007;103:2105-2111.
225. Ingemann-Hansen T, Halkjaer-Kristensen J, Halskov O. Skeletal muscle phosphagen
and lactate concentrations in ischaemic dynamic exercise. Eur J Appl Physiol. 1981; 46: 261–
270.
226. Tesch P, Karlsson J. Lactate in fast and slow twitch skeletal muscle fibres of man
during isometric contraction. Acta Physiol Scand. 1977;99:230-236.
227. Johnson MA, Polgar J, Weightman D, Appleton D. Data on the distribution of fibre
types in thirty-six human muscles. An autopsy study. J Neurol Sci. 1973;18:111-129.

228. Elder GC, Bradbury K, Roberts R. Variability of fiber type distributions within human
muscles. J Appl Physiol. 1982;53:1473-1480.
229. Lexell J, Henriksson-Larsén K, Sjöström M. Distribution of different fibre types in
human skeletal muscles. 2. A study of cross-sections of whole m. vastus lateralis. Acta
Accepted Article
Physiol Scand. 1983;117:115-122.
230. Lexell J, Taylor CC. Variability in muscle fibre areas in whole human quadriceps
muscle. How much and why? Acta Physiol Scand. 1989;136:561-568.
231. Hultman E, Bergström J, Anderson NM. Breakdown and resynthesis of
phosphorylcreatine and adenosine triphosphate in connection with muscular work in man.
Scand J Clin Lab Invest. 1967;19:56-66.
232. Karlsson J, Saltin B. Lactate, ATP, and CP in working muscles during exhaustive
exercise in man. J Appl Physiol. 1970;29:596-602.
233. Karlsson J, Diamant B, Saltin B. Muscle metabolites during submaximal and maximal
exercise in man. Scand J Clin Lab Invest. 1970;26:385-394.
234. Rehunen S, Näveri H, Kuoppasalmi K, Härkönen M. High-energy phosphate
compounds during exercise in human slow-twitch and fast-twitch muscle fibres. Scand J Clin
Lab Invest. 1982;42:499-506.
235. Madapallimattam AG, Cross A, Nishio ML, Jeejeebhoy KN. Stability of high-energy
substrates in fast- and slow-twitch muscle: comparison of enzymatic assay of biopsy with in
vivo 31P nuclear magnetic resonance spectroscopy. Anal Biochem. 1994;217:103-139.
236. Farina D, Holobar A, Merletti R, Enoka RM. Decoding the neural drive to muscles from
the surface electromyogram. Clin Neurophysiol. 2010;121:1616-623.
237. Vigotsky AD, Halperin I, Lehman GJ, et al. Interpreting signal amplitudes in surface
electromyography studies in sport and rehabilitation sciences. Front Physiol. 2018;8:985.
238. Moritani T, Stegeman D, Merletti R. Basic physiology and biophysics of EMG signal
generation. In: Electromyography: physiology, engineering and noninvasive applications.
Edited by Merletti, R. and Parker, P. USA. IEEE Press. Wiley-lnterscience, 2004, pp. 1-25.
239. De Luca CJ, Chang SS, Roy SH, et al. Decomposition of surface EMG signals from cyclic
dynamic contractions. J Neurophysiol. 2015;113:1941-1951.
240. Farina D, Enoka RM. Surface EMG decomposition requires an appropriate validation. J
Neurophysiol. 2011;105:981-982.
241. De Luca CJ, Nawab H. Reply to Farina and Enoka: The reconstruct-and-test approach is
the most appropriate validation for surface EMG signal decomposition to date. J
Neurophysiol. 2011;105:983-984.
242. Farina D, Merletti R, Enoka RM. The extraction of neural strategies from the surface
EMG: an update. J Appl Physiol. 2014;117:1215-1230.
243. Farina D, Merletti R, Enoka RM. Reply to De Luca, Nawab, and Kline: The proposed
method to validate surface EMG signal decomposition remains problematic. J Appl Physiol.
2015;118:1085.
244. De Luca CJ, Nawab SH, Kline JC. Clarification of methods used to validate surface EMG
decomposition algorithms as described by Farina et al. (2014). J Appl Physiol. 2015;118:1084.
245. Arabadzhiev TI, Dimitrov VG, Dimitrova NA, Dimitrov GV. Influence of motor unit
synchronization on amplitude characteristics of surface and intramuscularly recorded EMG
signals. Eur J Appl Physiol. 2010;108:227-237.
246. Arabadzhiev TI, Dimitrov VG, Dimitrov GV. The increase in surface EMG could be a
misleading measure of neural adaptation during the early gains in strength. Eur J Appl
Physiol. 2014;114:1645-1655.
247. Dimitrova NA, Dimitrov GV. Interpretation of EMG changes with fatigue: facts, pitfalls,
and fallacies. J Electromyogr Kinesiol. 2003;13:13-36.
248. Hanson J, Persson A. Changes in the action potential and contraction of isolated frog
muscle after repetitive stimulation. Acta Physiol Scand. 1971;81:340-348.

249. Hanson J. The effects of repetitive stimulation on the action potential and the twitch
of rat muscle. Acta Physiol Scand. 1974;90:387-400.
250. Hanson J. Effects of repetitive stimulation on membrane potentials and twitch in
human and rat intercostal muscle fibres. Acta Physiol Scand. 1974;92:238-248.
Accepted Article
251. Keenan KG, Farina D, Maluf KS, et al. Influence of amplitude cancellation on the
simulated surface electromyogram. J Appl Physiol. 2005;98:120-131.
252. Dideriksen JL, Enoka RM, Farina D. Neuromuscular adjustments that constrain
submaximal EMG amplitude at task failure of sustained isometric contractions. J Appl
Physiol. 2011;111:485-494.
253. Farina D, Merletti R, Enoka RM. The extraction of neural strategies from the surface
EMG. J Appl Physiol. 2004;96:1486-1495.
254. Neyroud D, Kayser B, Place N. The need to normalize surface EMG. J Appl Physiol.
2015;119:1519.
255. Martinez-Valdes E, Negro F, Falla D, et al. Surface electromyographic amplitude does
not identify differences in neural drive to synergistic muscles. J Appl Physiol. 2018;124:1071-
1079.
256. Moritani T, Muro M. Motor unit activity and surface electromyogram power spectrum
during increasing force of contraction. Eur J Appl Physiol Occup Physiol. 1987;56:260-265.
257. Clamann HP. Activity of single motor units during isometric tension. Neurology.
1970;20:254-260.
258. Knight CA, Kamen G. Superficial motor units are larger than deeper motor units in
human vastus lateralis muscle. Muscle Nerve. 2005;31:475-480.
259. Edgerton VR, Smith JL, Simpson DR. Muscle fibre type populations of human leg
muscles. Histochem J. 1975;7:259-266.
260. Ray CA, Dudley GA. Muscle use during dynamic knee extension: implication for
perfusion and metabolism. J Appl Physiol. 1998;85:1194-1197.
261. Enocson AG, Berg HE, Vargas R, et al. Signal intensity of MR-images of thigh muscles
following acute open- and closed chain kinetic knee extensor exercise - index of muscle use.
Eur J Appl Physiol. 2005;94:357-363.
262. Kalliokoski KK, Boushel R, Langberg H, et al. Differential glucose uptake in quadriceps
and other leg muscles during one-legged dynamic submaximal knee-extension exercise.
Front Physiol. 2011;2:75.
263. Richardson RS, Frank LR, Haseler LJ. Dynamic knee-extensor and cycle exercise:
functional MRI of muscular activity. Int J Sports Med. 1998;19:182-187.
264. Watanabe K, Akima H. Normalized EMG to normalized torque relationship of vastus
intermedius muscle during isometric knee extension. Eur J Appl Physiol. 2009;106:665-673.
265. Thorstensson A, Karlsson J, Viitasalo JH, et al. Effect of strength training on EMG of
human skeletal muscle. Acta Physiol Scand. 1976;98:232-236.
266. Häkkinen K, Komi PV. Electromyographic changes during strength training and
detraining. Med. Sci. Sports Exerc. 1983;15:455-460.
267. Eloranta V. Patterning of muscle activity in static knee extension. Electromyogr. Clin.
268. Alkner BA, Tesch PA, Berg HE. Quadriceps EMG/force relationship in knee extension
and leg press. Med Sci Sports Exerc. 2000;32:459-463.
269. Rabita G, Pérot C, Lensel-Corbeil G. Differential effect of knee extension isometric
training on the different muscles of the quadriceps femoris in humans. Eur J Appl Physiol.
2000;83:531-538.
270. Amiridis IG, Martin A, Morlon B, et al. Co-activation and tension-regulating
phenomena during isokinetic knee extension in sedentary and highly skilled humans. Eur J
Appl Physiol Occup Physiol. 1996;73:149-156.

271. Komi PV, Linnamo V, Silventoinen P, Sillanpää M. Force and EMG power spectrum
during eccentric and concentric actions. Med Sci Sports Exerc. 2000;32:1757-1762.
272. Ekstrom RA, Osborn RW, Goehner HM, et al. Electromyographic normalization
procedures for determining exercise intensity of closed chain exercises for strengthening the
Accepted Article
quadriceps femoris muscles. J Strength Cond Res. 2012;26:766-771.
273. Beutler AI, Cooper LW, Kirkendall DT, Garrett WE Jr. Electromyographic analysis of
single-leg, closed chain exercises: implications for rehabilitation after anterior cruciate
ligament reconstruction. J Athl Train. 2002;37:13-18.
274. Westing SH, Cresswell AG, Thorstensson A. Muscle activation during maximal
voluntary eccentric and concentric knee extension. Eur J Appl Physiol Occup Physiol.
1991;62:104-108.
275. Aagaard P, Simonsen EB, Andersen JL, et al. Neural inhibition during maximal
eccentric and concentric quadriceps contraction: effects of resistance training. J Appl Physiol.
2000;89:2249-2257.
276. Tesch PA, Dudley GA, Duvoisin MR, et al. Force and EMG signal patterns during
repeated bouts of concentric or eccentric muscle actions. Acta Physiol Scand. 1990;138:263-
721.
277. Duchateau J, Enoka RM. Human motor unit recordings: origins and insight into the
integrated motor system. Brain Res. 2011;1409:42-61.
278. Basmajian JV, De Luca CJ. Muscles Alive. Baltimore: Williams & Wilkins, 1985.
279. Moritani T, Muro M, Nagata A. Intramuscular and surface electromyogram changes
during muscle fatigue. J Appl Physiol. 1986;60:1179-1185.
280. Trontelj JV, Jabre J, Mihelin M. Needle and wire detection techniques. In:
Electromyography: physiology, engineering and noninvasive applications. Edited by Merletti,
R. and Parker, P. USA. IEEE Press. Wiley-lnterscience, 2004, 27-46.
281. Kent-Braun JA, Miller RG, Weiner MW. Human skeletal muscle metabolism in health
and disease: utility of magnetic resonance spectroscopy. Exerc Sport Sci Rev. 1995;23:305-
347.
282. Sullivan MJ, Saltin B, Negro-Vilar R, et al. Skeletal muscle pH assessed by biochemical
and 31P-MRS methods during exercise and recovery in men. J Appl Physiol. 1994;77:2194-
3200.
283. Constantin-Teodosiu D, Greenhaff PL, et al. Anaerobic energy production in human
skeletal muscle in intense contraction: a comparison of 31P magnetic resonance
spectroscopy and biochemical techniques. Exp Physiol. 1997;82:593-601.
284. Vandenborne K, McCully K, Kakihira H, et al. Metabolic heterogeneity in human calf
muscle during maximal exercise. Proc Natl Acad Sci U S A. 1991;88:5714-5718.
285. Park JH, Brown RL, Park CR, et al. Functional pools of oxidative and glycolytic fibers in
human muscle observed by 31P magnetic resonance spectroscopy during exercise. Proc Natl
Acad Sci U S A. 1987;84:8976-8980.
286. Achten E, Van Cauteren M, Willem R, et al. 31P-NMR spectroscopy and the metabolic
properties of different muscle fibers. J Appl Physiol. 1990;68:644-659.
287. Houtman CJ, Heerschap A, Zwarts MJ, Stegeman DF. pH heterogeneity in tibial
anterior muscle during isometric activity studied by (31)P-NMR spectroscopy. J Appl Physiol.
2001;91:191-200.
288. Harris RC, Essén B, Hultman E. Glycogen phosphorylase activity in biopsy samples and
single muscle fibres of musculus quadriceps femoris of man at rest. Scand J Clin Lab Invest.
1976;36:512-516.
289. Mizuno M, Secher NH, Quistorff B. 31P-NMR spectroscopy, rsEMG, and histochemical
fiber types of human wrist flexor muscles. J Appl Physiol. 1994;76:531-538.

290. Sahlin K, Harris RC, Hultman E. Creatine kinase equilibrium and lactate content
compared with muscle pH in tissue samples obtained after isometric exercise. Biochem J.
1975;152:173-180.
291. Meyer RA, Slade JM, Towse TF, et al. Phosphocreatine resynthesis during recovery
Accepted Article
after exercise with blood flow occlusion. Med Sci Sports Exerc 2008;40.5: S349.
292. Taylor DJ, Bore PJ, Styles P, et al. Bioenergetics of intact human muscle. A 31P nuclear
magnetic resonance study. Mol Biol Med. 1983;1:77-94.
293. Jeneson JA, Wesselink MW, de Boer RW, Amelink HG. Peak-splitting of inorganic
phosphate during exercise: anatomy or physiology? A MRI-guided 31P MRS study of human
forearm muscle. In: Proc. 8th A. Meet. Soc. Magn. Reson. Med. 1989. p. 1030.
294. Johnson R, Arnold D, Leger G. Splitting of the phosphate peak in exercising muscle is
due to macroscopic not microscopic heterogeneity (Abstract). Magn. Reson. Med.
1990;Suppl. 2: 866.
295. Vandenborne K, Walter G, Leigh JS, Goelman G. pH heterogeneity during exercise in
localized spectra from single human muscles. Am J Physiol. 1993;265:C1332-1339.
296. Mizuno M, Justesen LO, Bedolla J, et al. Partial curarization abolishes splitting of the
inorganic phosphate peak in 31P-NMR spectroscopy during intense forearm exercise in man.
Acta Physiol Scand. 1990;139:611-2.
297. Mizuno M, Horn A, Secher NH, Quistorff B. Exercise-induced 31P-NMR metabolic
response of human wrist flexor muscles during partial neuromuscular blockade. Am J
Physiol. 1994;267:R408-414.
298. Yanagisawa O, Niitsu M, Yoshioka H, et al. MRI determination of muscle recruitment
variations in dynamic ankle plantar flexion exercise. Am J Phys Med Rehabil. 2003;10:760-
765.
299. Segal RL, Song AW. Nonuniform activity of human calf muscles during an exercise
task. Arch Phys Med Rehabil. 2005;86:2013-2017.
300. Meyerspeer M, Robinson S, Nabuurs CI, et al. Comparing localized and nonlocalized
dynamic 31P magnetic resonance spectroscopy in exercising muscle at 7 T. Magn Reson
Med. 2012;68:1713-1723.
301. Valkovič L, Chmelík M, Just Kukurová I, et al. Depth-resolved surface coil MRS (DRESS)-
localized dynamic (31) P-MRS of the exercising human gastrocnemius muscle at 7 T. NMR
Biomed. 2014;27:1346-1352.
302. Rossiter HB, Ward SA, Howe FA, et al. Dynamics of intramuscular 31P-MRS P(i) peak
splitting and the slow components of PCr and O2 uptake during exercise. J Appl Physiol.
2002;93:2059-2069.
303. Tesch PA, Karlsson J. Effects of exhaustive, isometric training on lactate accumulation
in different muscle fiber types. Int J Sports Med. 1984;5:89-91.
304. Houtman CJ, Heerschap A, Zwarts MJ, Stegeman DF. An additional phase in PCr use
during sustained isometric exercise at 30% MVC in the tibialis anterior muscle. NMR Biomed.
2002;15:270-277.
305. Yoshida T, Watari H. Exercise-induced splitting of the inorganic phosphate peak:
investigation by time-resolved 31P-nuclear magnetic resonance spectroscopy. Eur J Appl
Physiol Occup Physiol. 1994;69:465-473.
306. Miller RG, Kent-Braun JA, Sharma KR, Weiner MW. Mechanisms of human muscle
fatigue. Quantitating the contribution of metabolic factors and activation impairment. In:
Fatigue. Edited by Gandevia SC, Enoka RM, McComas AJ et al., Springer, Boston, MA, 1995.
pp. 195-201.
307. Miller RG, Giannini D, Milner-Brown HS, et al. Effects of fatiguing exercise on high-
energy phosphates, force, and EMG: evidence for three phases of recovery. Muscle Nerve.
1987;10:810-821.

308. Vestergaard-Poulsen P, Thomsen C, Sinkjaer T, et al. Simultaneous electromyography
and 31P nuclear magnetic resonance spectroscopy--with application to muscle fatigue.
Electroencephalogr Clin Neurophysiol. 1992;85:402-411.
309. Vestergaard-Poulsen P, Thomsen C, et al. Simultaneous 31P-NMR spectroscopy and
Accepted Article
EMG in exercising and recovering human skeletal muscle: a correlation study. J Appl Physiol.
1995;79:1469-1478.
310. Bendahan D, Jammes Y, Salvan AM. Combined electromyography--31P-magnetic
resonance spectroscopy study of human muscle fatigue during static contraction. Muscle
Nerve. 1996;19:715-721.
311. Houtman CJ, Stegeman DF, Van Dijk JP, Zwarts MJ. Changes in muscle fiber
conduction velocity indicate recruitment of distinct motor unit populations. J Appl Physiol.
2003;95:1045-1054.
312. Giannesini B, Cozzone PJ, Bendahan D. Non-invasive investigations of muscular
fatigue: metabolic and electromyographic components. Biochimie. 2003;85:873-883.
313. Crenshaw AG, Hargens AR, Gershuni DH, Rydevik B. Wide tourniquet cuffs more
effective at lower inflation pressures. Acta Orthop Scand 1988: 59: 447-451.
314. Wernbom M, Augustsson J, Thomeé R. Effects of vascular occlusion on muscular
endurance in dynamic knee extension exercise at different submaximal loads. J Strength
Cond Res. 2006;20:372-377.
315. Brooke MH, Kaiser KK. Muscle fiber types: how many and what kind? Arch Neurol.
1970;23:369-739.
316. Green H, Goreham C, Ouyang J, et al. Regulation of fiber size, oxidative potential, and
capillarization in human muscle by resistance exercise. Am J Physiol. 1999;276:R591-596.
317. Billeter R, Heizmann CW, Howald H, Jenny E. Analysis of myosin light and heavy chain
types in single human skeletal muscle fibers. Eur J Biochem. 1981;116:389-395.
318. Brooke MH, Kaiser KK. Three "myosin adenosine triphosphatase" systems: the nature
of their pH lability and sulfhydryl dependence. J Histochem Cytochem. 1970;18:670-672.
319. Dubowitz V, Sewry CA, Oldfors A. Muscle biopsy: a practical approach. 4th Edition.
Elsevier Health Sciences, 2013.
320. Fridén J, Seger J, Ekblom B. Sublethal muscle fibre injuries after high-tension
anaerobic exercise. Eur J Appl Physiol Occup Physiol. 1988;57:360-368.
321. Han YS, Proctor DN, Geiger PC, Sieck GC. Reserve capacity for ATP consumption during
isometric contraction in human skeletal muscle fibers. J Appl Physiol. 2001;90:657-664.
322. Piehl K. Time course for refilling of glycogen stores in human muscle fibres following
exercise-induced glycogen depletion. Acta Physiol Scand. 1974;90:297-302.
323. Fairchild TJ, Armstrong AA, Rao A. Glycogen synthesis in muscle fibers during active
recovery from intense exercise. Med Sci Sports Exerc. 2003;35:595-602.
324. Wernbom M, Apro W, Paulsen G, et al. Acute low-load resistance exercise with and
without blood flow restriction increased protein signalling and number of satellite cells in
human skeletal muscle. Eur J Appl Physiol. 2013;113:2953-2965.
325. Zehnder M, Muelli M, Buchli R, et al. Further glycogen decrease during early recovery
after eccentric exercise despite a high carbohydrate intake. Eur J Nutr. 2004;43:148-159.
326. Andersen P, Saltin B. Maximal perfusion of skeletal muscle in man. J Physiol.
1985;366:233-249.
327. Person RS, Golubovich K. Electromyographic study of fatigue in man in conditions of
artificial ischemia in the working muscle. Fiziologicheskii zhurnal SSSR imeni IM Sechenova,
1960;46:1181-1187.
328. Arendt-Nielsen L, Mills KR. The relationship between mean power frequency of the
EMG spectrum and muscle fibre conduction velocity. Electroencephalogr Clin Neurophysiol.
1985;60:130-134.

329. Myers SJ, Sullivan WP. Effect of crculatory occlusion on time to muscular fatigue. J
Appl Physiol. 1968;24:54-59.
330. Sullivan WP. Muscle fatigue and electromyographic changes during isometric exercise
in presence of vascular occlusion. 1968. PhD Thesis. The Ohio State University.
Accepted Article
331. Edgerton VR, Roy RR, Apor P. Specific tension of human elbow flexor muscles. In:
Saltin B (ed) Biochemistry of exercise VI. Human Kinetics, Champaign, Ill., 1986, pp 487-500.
332. Signorile JF, Digel S, Moccia G, et al. Effects of partial occlusion of circulation on
frequency and amplitude of surface electromyography. J Electromyogr Kinesiol. 1991;1:124-
129.
333. Yasuda T, Fukumura K, Iida H, Nakajima T. Effect of low-load resistance exercise with
and without blood flow restriction to volitional fatigue on muscle swelling. Eur J Appl Physiol.
2015;115:919-926.
334. Alway SE, MacDougall JD, Sale DG. Contractile adaptations in the human triceps surae
after isometric exercise. J Appl Physiol. 1989;66:2725-2732.
335. Alway SE, Sale DG, MacDougall JD. Twitch contractile adaptations are not dependent
on the intensity of isometric exercise in the human triceps surae. Eur J Appl Physiol Occup
Physiol. 1990;60:346-352.
336. Moore DR, Burgomaster KA, Schofield LM, et al. Neuromuscular adaptations in human
muscle following low intensity resistance training with vascular occlusion. Eur J Appl Physiol.
2004;92:399-406.
337. Borg G. Borg's perceived exertion and pain scales. Human Kinetics, Champaign, Ill.,
1998.
338. Kooistra RD, de Ruiter CJ, de Haan A. Muscle activation and blood flow do not explain
the muscle length-dependent variation in quadriceps isometric endurance. J Appl Physiol.
2005;98:810-816.
339. Cook SB, Murphy BG, Labarbera KE. Neuromuscular function after a bout of low-load
blood flow-restricted exercise. Med Sci Sports Exerc. 2013;45:67-74.
340. Wernbom M. (2011). Effects of an acute bout of low-load resistance training with
blood flow restriction - with special reference to muscle damage, hypertrophic signaling and
satellite cells (Doctoral dissertation, Norwegian School of Sport Sciences, Oslo, Norway).
Retrieved from Google Scholar.
341. Loenneke JP, Kim D, Fahs CA, et al. Effects of exercise with and without different
degrees of blood flow restriction on torque and muscle activation. Muscle Nerve 2015;713-
721.
342. Fatela P, Reis JF, Mendonca GV, et al. Acute neuromuscular adaptations in response
to low-intensity blood-flow restricted exercise and high-intensity resistance exercise: are
there any differences? J Strength Cond Res. 2018;32:902-910.
343. Magora A, Blank A, Gonen B. Effects of artificially induced ischemia (AII) on the
electrophysiological pattern of muscular fatigue in healthy humans. Electromyogr Clin
344. Yoshida T, Watari H. Effect of circulatory occlusion on human muscle metabolism
during exercise and recovery. Eur J Appl Physiol Occup Physiol. 1997;75:200-205.
345. Suga T, Okita K, Morita N, et al. Intramuscular metabolism during low-intensity
resistance exercise with blood flow restriction. J Appl Physiol. 2009;106:1119-1124.
346. Suga T, Okita K, Morita N, et al. Dose effect on intramuscular metabolic stress during
low-intensity resistance exercise with blood flow restriction. J Appl Physiol . 2010;108:1563-
1567.
347. Trappe TA, Raue U, Tesch PA. Human soleus muscle protein synthesis following
resistance exercise. Acta Physiol Scand. 2004;182:189-196.
348. Gravel D, Richards CL, Filion M. Angle dependency in strength measurements of the
ankle plantar flexors. Eur J Appl Physiol Occup Physiol. 1990;61:182-187.

349. Peeters M, Svantesson U, Grimby G. Effect of prior isometric muscle action on
concentric torque output during plantar flexion. Eur J Appl Physiol Occup Physiol.
1995;71:272-275.
350. Nadeau S, Gravel D, Arsenault AB, Goyette M. Preloading and range of motion effect
Accepted Article
on plantarflexor muscle performance. Arch Phys Med Rehabil. 1996;77:1000-1004.
351. Svantesson U, Grimby G, Thomeé R. Potentiation of concentric plantar flexion torque
following eccentric and isometric muscle actions. Acta Physiol Scand. 1994;152:287-293.
352. Yanagisawa O, Sanomura M. Effects of low-load resistance exercise with blood flow
restriction on high-energy phosphate metabolism and oxygenation level in skeletal muscle.
Interv Med Appl Sci. 2017;9:67-75.
353. Saltin B, Gollnick PD. Skeletal muscle adaptability: significance for metabolism and
performance. Handbook of Physiology. Skeletal Muscle, 1983,10:555-631.
354. Bangsbo J, Johansen L, Quistorff B, Saltin B. NMR and analytic biochemical evaluation
of CrP and nucleotides in the human calf during muscle contraction. J Appl Physiol.
1993;74:2034-2039.
355. Vandenborne K, Walter G, Ploutz-Snyder L, et al. Energy-rich phosphates in slow and
fast human skeletal muscle. Am J Physiol. 1995;268:C869-876.
356. Takarada Y, Sato Y, Ishii N. Effects of resistance exercise combined with vascular
occlusion on muscle function in athletes. Eur J Appl Physiol 2002: 86: 308-314.
357. Kawada S. What phenomena do occur in blood flow-restricted muscle? Int J Kaatsu
Training Res 2005: 1: 37-44.
358. Grimby L, Hannerz J. Disturbances in voluntary recruitment order of low and high
frequency motor units on blockades of proprioceptive afferent activity. Acta Physiol Scand.
1976;96:207-216.
359. Buchthal F, Pinell P, Rosenfalck P. Action potential parameters in normal human
muscle and their physiological determinants. Acta Physiol Scand. 1954;32:219-229.
360. Goldberg J, Sullivan SJ, Seaborne DE. The effect of two intensities of massage on H-
reflex amplitude. Phys Ther. 1992;72:449-457.
361. Robichaud JA, Agostinucci J, Vander Linden DW. Effect of air-splint application on
soleus muscle motoneuron reflex excitability in nondisabled subjects and subjects with
cerebrovascular accidents. Phys Ther. 1992;72:176-183.
362. Agostinucci J. Circumferential pressure’s inhibitory effects on soleus H-reflex. Transl
Neurosci 2013;4:251-259.
363. Agostinucci J. The effects of circumferential pressure on the soleus muscle F-wave in
healthy subjects. J Electromyogr Kinesiol. 2012;22:223-227.
364. Ge W, Khalsa PS. Encoding of compressive stress during indentation by group III and
IV muscle mechano-nociceptors in rat gracilis muscle. J Neurophysiol. 2003;89:785-792.
365. Ørtenblad N, Macdonald WA, Sahlin K. Glycolysis in contracting rat skeletal muscle is
controlled by factors related to energy state. Biochem J. 2009;420:161-168.
366. Lowe DA, Alway SE. Animal models for inducing muscle hypertrophy: are they
relevant for clinical applications in humans? J Orthop Sports Phys Ther. 2002;32:36-43.
367. Goto K, Okuyama R, Sugiyama H, et al. Effects of heat stress and mechanical stretch
on protein expression in cultured skeletal muscle cells. Pflügers Arch. 2003:447:247-253.
368. Goto K, Oda H, Kondo H, et al. Responses of muscle mass, strength and gene
transcripts to long-term heat stress in healthy human subjects. Eur J Appl Physiol.
2011;111:17-27.
369. Sudo M, Ando S, Kano Y. Repeated blood flow restriction induces muscle fiber
hypertrophy. Muscle Nerve. 2017;55:274-276.
370. Natsume T, Ozaki H, Saito AI, et al. Effects of electrostimulation with blood flow
restriction on muscle size and strength. Med Sci Sports Exerc. 2015;47:2621-2627.

371. Gorgey AS, Timmons MK, Dolbow DR, et al. Electrical stimulation and blood flow
restriction increase wrist extensor cross-sectional area and flow meditated dilatation
following spinal cord injury. Eur J Appl Physiol. 2016;116:1231-1244.
372. Nakajima T, Koide S, Yasuda T, et al. Muscle hypertrophy following blood flow-
Accepted Article
restricted, low-force isometric electrical stimulation in rat tibialis anterior: role for muscle
hypoxia. J Appl Physiol. 2018;125:134-145.
373. Slysz JT, Burr JF. The effects of blood flow restricted electrostimulation on strength
and hypertrophy. J Sport Rehabil. 2018;27:257-262.
374. Goto K, Oda H, Morioka S, et al. Skeletal muscle hypertrophy induced by low-intensity
exercise with heat stress in healthy human subjects. Jpn J Aerosp Environ Med. 2007;44:13-
18.
375. Wernbom M, Augustsson J, Thomeé R. The influence of frequency, intensity, volume
and mode of strength training on whole muscle cross-sectional area in humans. Sports Med.
2007; 37: 225-264.
376. Mitchell CJ, Churchward-Venne TA, West DW, et al. Resistance exercise load does not
determine training-mediated hypertrophic gains in young men. J Appl Physiol. 2012;113:71-
77.
377. Sieljacks P, Degn R, Hollaender K, et al. Non-failure blood flow restricted exercise
induces similar muscle adaptations and less discomfort than failure protocols. Scand J Med
Sci Sports. 2019;29:336-347.
378. Kubo K, Komuro T, Ishiguro N, et al. Effects of low-load resistance training with
vascular occlusion on the mechanical properties of muscle and tendon. J Appl Biomech.
2006;22:112-119.
379. Colomer-Poveda D, Romero-Arenas S, Vera-Ibáñez A, et al. Effects of 4 weeks of low-
load unilateral resistance training, with and without blood flow restriction, on strength,
thickness, V wave, and H reflex of the soleus muscle in men. Eur J Appl Physiol.
2017;117:1339-1347.
380. Cook SB, Scott BR, Hayes KL, Murphy BG. Neuromuscular Adaptations to low-load
blood flow restricted resistance training. J Sports Sci Med. 2018;17:66-73.

Figure legends.
Accepted Article
Figure 1. Suggested scheme of the interplay between the recruitment of motor units
innervating type I, IIA, IIAX and IIX muscle fibres and the accompanying increases in firing
rates with increases in contractile force production. The scheme depicts short (~0.5-1 s) but
non-ballistic contractions, as opposed to slow ramp (~10 s) contractions (see text). Modified
from Sale42 based on data discussed in this review.
Figure 2. Some of the possible pathways and processes of fatigue in strenuous BFRRE. See
text for details. Thicker lines indicate a relatively more substantial influence.
Figure 3. Neighbouring muscle cross sections stained with the PAS staining method for
glycogen (left) and for MHC I (right, bright red staining) to display type I fibres, showing
lower glycogen levels in type I fibres at 1 hour after an acute bout of BFRRE in one subject.
Figure courtesy of Kristoffer Cumming, with permission.
Figure 4. Nonconsecutive muscle cross sections stained with the PAS staining method for
glycogen (left) and for MHC I (right, green staining) to display type I fibres, showing lower
glycogen levels in type I fibres at 24 hours after an acute bout of BFRRE in one subject.
Figure courtesy of Kristoffer Cumming and Mathias Wernbom.
Figures 5a-d. Muscle activity measured by EMG during an acute bout of BFRRE exercise
consisting of 4 sets to volitional failure of low-load dynamic knee extensions (20-25% of
1RM) with continuous partial BFR (~50% of seated resting arterial occlusion pressure). Rest
between the sets: 45 seconds. Upper left figure: 1st set, 61 repetitions. Lower right figure: 4th
set, 8 repetitions. EMG normalised to MVC (100%). Red = vastus lateralis, left leg. Blue =
vastus medialis, left leg. Figures adapted from Wernbom340.
Figure 6. Muscle EMG activity during heavy resistance exercise bout with dynamic knee
extensions (80-85% of 1RM), a single set to failure (7 repetitions). Same subject as in Figure
6. EMG normalised to isometric MVC (100%). Red = vastus lateralis, right leg. Blue = vastus
medialis, right leg. Adapted from Wernbom340.
Figure 7. Suggested scheme of the successive and cumulative activation of type I, IIA, IIAX
and IIX muscle fibres in low-load BFRRE with increasing fatigue and duration. The scheme
depicts BFRRE performed at ~20-30% of 1RM with a hypothetical representative limb
muscle (e.g. the vastus lateralis or the biceps brachii) consisting of 50% type I, 30% type IIA,
10% type IIAX and 10% type IIX muscle fibres. The scheme reflects the proposed largely
intact orderly recruitment of motor units in BFRRE, but also allows for slight alterations in
recruitment thresholds that may occur in some motor units (see text for details).

Accepted Article

Accepted Article

Accepted Article

Accepted Article

Accepted Article

Accepted Article

Accepted Article

Apha 13302

Transféré par

Informations du document

Titre original

Copyright

Formats disponibles

Partager ce document

Partager ou intégrer le document

Options de partage

Avez-vous trouvé ce document utile ?

Ce contenu est-il inapproprié ?

Droits d'auteur :

Formats disponibles

Apha 13302

Transféré par

Droits d'auteur :

Formats disponibles

DR MATHIAS WERNBOM (Orcid ID : 0000-0003-0411-3611)

Muscle fibre activation and fatigue with low-load blood

Short title: Muscle fibre use in ischemic exercise

reached a plateau in muscle mass and strength.

fiber activation during BFRRE.

evidence on muscle fibre recruitment during BFRRE as revealed by various methods

Keywords: Occlusion, ischemia, muscle hypertrophy, motor units, fatigue

1. Introduction, background and purpose

2. Definitions and limitations

3. Neural control of muscle force development

This article is protected by copyright. All rights reserved.

5. Methods to investigate muscle fibre activation and use

6. Studies on muscle fibre activation and use in BFRRE

7. Conclusions and implications

conventional heavy resistance exercise in untrained individuals1-3, although a recent meta-

exercise (BFRRE). Nevertheless, the marked neuromuscular adaptations observed with

This article is protected by copyright. All rights reserved.

in conventional strength training (reviewed by Wernbom et al5 and Hughes et al6).

This article is protected by copyright. All rights reserved.

primarily in the type I myofibres with low-load BFRRE22-24.

some of the advantages and limitations of these methods.

necessary to provide an introductory background of the physiology of these processes

(Sections 3 and 4) as well as a discussion of different methods to investigate motor unit

neuromuscular adaptations to systematic training periods of BFRRE.

2. Definitions and limitations

This article is protected by copyright. All rights reserved.

and the load range referred to here is 20-50% of MVC.

physiological range are included and discussed in this review.

3. Neural control of muscle force development

Recent studies using sophisticated decomposition of surface electromyographic signals have

advantages and disadvantages with these methods.

This article is protected by copyright. All rights reserved.

Furthermore, it is worth mentioning that the human studies which failed to

medial gastrocnemius muscle using intramuscular electrical microstimulation and reported

This article is protected by copyright. All rights reserved.

whereas many other muscles display a more continuous distribution.

three subtypes are considered together (see Staron41).

in some types of athletes. As an extreme example, Andersen et al45, using electrophoretic

immunohistochemistry methods also express varying amounts of type IIA myosin41,45,47. In

This article is protected by copyright. All rights reserved.

myofibres, as indicated by the recent discovery of two metabolically distinct populations,

types in the fibre type continuum as proposed by Prats et al50.

The size principle of motor unit recruitment was first demonstrated by

contractions and with transcranial stimulation of the motor cortex62,63.

at ~50-60% of MVC27,28,64. However, accumulating evidence suggests recruitment of new

This article is protected by copyright. All rights reserved.

categories of very rapid (ballistic) contractions, lengthening (eccentric) contractions and

voluntary movements, including ballistic and eccentric contractions.

However, an unequal balance of excitatory and inhibitory inputs onto motor

electrical stimulation of cutaneous afferents, resulting in elevated recruitment thresholds in

This article is protected by copyright. All rights reserved.

these high-threshold units. Furthermore, electrical stimulation of cutaneous afferents might be

considered a non-physiological condition compared with normal tactile stimulation78,79. Still,

Masakado et al76 being a notable exception.

3.3. Rate coding of motor units in voluntary movements

be accomplished entirely by rate coding (reviewed by Duchateau et al28). A generalized

This article is protected by copyright. All rights reserved.

increasing stimulation frequency and reaches a plateau of maximum torque at ~50-60 Hz

Interestingly, even at 100% of MVC, the average maximum firing rates

when averaged across af few seconds.

This article is protected by copyright. All rights reserved.

lateralis94. Moreover, although earlier studies reported near-maximal activation of the

The study of Pucci et al102 is of particular interest, as it demonstrated not only