TMP 18

Chemical Senses Vol.16 no.4 pp.
373-416, 1991
Ninth Congress of the European Chemoreception Research

Organisation (ECRO IX) held in The Netherlands,
September 2 - 7 , 1990
Abstracts
1. Multiple transduction pathways activate different ionic conductances in lobster
olfactory receptor cells
Downloaded from http://chemse.oxfordjournals.org/ by Sergey Novikov on February 16, 2014

Barry W.Ache and William C.Michel
Whitney Laboratory and Departments of Zoology and Neuroscience, University of Florida, St Augustine,
FL 32086. USA
Earlier, whole-cell current-clamp recordings revealed that components of odor mixtures could hyperpolarize
as well as depolarize the same lobster olfactory (aesthetasc) receptor cell, implying that odors were activating
two opposing transduction pathways [McClintock and Ache (1989) Chem. Senses, 14, 637].
We now show that odor-evoked hyperpolarizations suppress excitation in a dose-dependent manner to sub-
micTomolar concentrations and, like depolarizations, are associated with a specific conductance. Receptor
potentials of both polarities are associated with an increase in membrane conductance, as indicated by significant
decreases in RN and T o during odor stimulation. Altering the ionic environment of the cilia (outer dendritic
segments) reveals that either an Na + - or a Ca2+-selective or a non-selective cation conductance underlies
the depolarizing current. In contrast, similar ion substitution experiments as well as the sensitivity of odor-
evoked hyperpolarizations to 4-AP and Cs + blockade indicate that a K + conductance underlies the hyper-
polanzing current. Our evidence is consistent with the idea that odorants are not bidirectionally regulating
a single conductance, but rather are activating two separate types of conductances in the same cell. Experiments
in progress attempt to identify the second messenger^), if any, associated with these conductance pathways.
(Supported by NSF 88-10261 and ONR N00014-90-J-1566.)
2. Specificity, sensitivity and temporal response pattern of pheromone receptor

neurons in Heliothis tea
T.J.Almaas, T.A.Christensen1 and H.Mustaparta
Department of Zoology, University of Trondheim, AVH, 7055 Dragvoll, Norway and 'ARL, Division of
Neurobiology, 611 Gould-Simpson Science Building, University of Arizona, Tucson, AZ 85721, USA
In H.zjea the species-specific pheromone blend contain four components of which two (major components)
are necessary and sufficient for attraction and full sequence of sexual behaviour in males [Klun et al. (1980)
/. Chem. Ecol., 6, 165]. In principle, this species-specific pheromone composition might provide reproduc-
tive isolation between H.zea and the sympatric species H. virescens. However, an additional isolation mechanism
exists, where (Z)-9-tetradecenal, a major pheromone component in H. virescens, inhibits the attraction of
H.zea males [Shaver et al. (1982) /. Chem. Evol., 4, 755]. The aim of the present study was to characterize
responses of receptor neurons (in the male specific sensilla trichodea type 1) to pheromones and the in-
terspecific chemical cue. Spike activity was recorded extracellulariy by the use of tungsten microelectrodes.
The majority of receptor neurons were tuned to either (i) (Z)-l 1-hexadecenal (the major pheromone com-
ponent in all Heliothis species studied, including H.zea) or (ii) Z9-14:A1, the interspecific inhibitor. The
sensitivity and specificity (response to analogues) were similar within each group and to corresponding types
of receptor neurons in //. vir£jc«u [Almaas and Mustaparta( 1990)/. Chem. Ecol., 16, 1331]. Surprisingly,
no receptor neurons responded specifically to other pheromome components. The sensitivity of single recep-
tor neurons did not change with age from day 1 to day 3. However, at higher ages, the individual sensitivities
varied considerably.
In spite of similar specificity and sensitivity, receptor neurons were found to differ concerning the tem-
poral response pattern, and ability to follow pulsed stimuli, either showing a strong phasic component with
rapid decay and ability to follow pulses or having a stronger tonic component and lower ability to follow pulses.
In conclusion, the similarities of receptor neuron populations and response characteristics of the two types
of neurons imply an evolutionary conservation of membrane receptors in these species. The variation in
temporal characteristics might be of importance for orientation in the pheromone plume.
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ECRO IX: Abstracts
3. Pheromonal contraception in male laboratory mice: new evidence for the

existence of functional dualism in sexual primer pheromones?
Vardan V.Babalyan and Sergej N.Novilcov
I.P.Pavlov Institute of Physiology, USSR Academy of Sciences, Leningrad 199034, USSR
CBAB6FI male mice, 30 days old, were subjected for 2 h to constant airflow passed over the evaporating
cuvette with a urine sample from a 12-month-old CBA/LacSto or a 4-6-month-old C57BU6Sto male laboratory
mouse. After 17 days the recipients were sacrificed and spermatozoa counts in cauda epididymis were analysed.
• It was shown that the urine from old CBA and middle-aged C57BL/6 male mice contains volatile factor(s)
which significantly decrease the level of gametes with altered head shape from 6.1 ± 0.35% (basal level)
to 1.9 ± 0.23% and 3.2 ± 0.26% in CBA and C57BLJ6 variants respectively.
These data give experimental evidence for the existence in the voided urine from male mice of some
semiochemicals with a promotory as well with an inhibitor mode of action on spermatogenesis in recipients

and stress once more the importance of genotype and/or physiological state and age of the odor donors in
any primer pheromone investigations.
Taken together in the light of some data on the complex mode of action and chemical nature of pheromones
which modulate the maturation process in female mice the above new findings are strong evidence for the
existence of functional dualism at least in rodent sexual primer pheromones. We postulate here that the vec-
tor of action depends on a fine balance in the mixture of chemicals with opposite physiological effects and
reflects in complex donor-recipient communicative processes.
4. Odor response distributions and psychophysical functions generated

by free-number magnitude estimation
J.C.Baird1, B.Berglund and M.J.Olsson
Institute of Environmental Medicine, Kamlinska Institute and Department of Psychology, Stockholm University,
Sweden, 'Main affiliation: Departments of Psychology and Mathematical Social Sciences, Dartmouth College,
Hanover, NH, USA
The method of free-number magnitude estimation was used in order to explore the form of the response
distribution for perceived odor intensity. Pyridine, which is a malodorous garlic-like substance, was presented
at seven supra-threshold concentrations (339-2013 p.p.b.) in a random order together with a background
stimulus (charcoal-filtered air). Altogether, each concentration was presented 105 times to each of five sub-
jects, each participating during three days. The results show that (i) the response function for weaker odor
intensities is strongly positively skewed but it approaches normality for the stronger intensities, and (ii) that
the exponent of Stevens' psychophysical power function varies systematically but non-monotonically with
repeated presentations for all the subjects. The results have theoretical implications for (a) the assumed shape
of the perceived intensity distribution underlying the responses (magnitude estimates), (b) the stability and
validity of the psychophysical power function, and (c) the choice of an adequate statistical measure of central
tendency for the response distribution.
5. Cortical topographies and psychometric ratings of odours

John Behan, Steve Van Toller1, Martin Kendal-Reed1 and Pauline Howells
Quest International, Ashford, Kent and 'Department of Psychology, University of Warwick, Coventry, CV4
7AL, UK
Notwithstanding the noisy and apparently random electrical activity of the cortex, the objective of the study
was to statistically examine EEG activity taken from subjects given odours to smell, for: (i) intersubjective
trends in cortical activity; (ii) systematic changes in EEG activity; (iii) correlations between psychometric
ratings and EEG activity.
The EEG analysis involved using alpha frequency (8-14 Hz) taken from the first 2.56 s epoch after presen-
tation of an odour. Averaged EEG data from 28 electrodes for each odour were taken from 14 subjects.
Six odours, representing a wide range of odour types, were used. They were: linalyl acetate; White Sap-
phire; Chandanol; indole; 5-alpha-androstan-3-one; Mackenzie's smelling salts (eucalyptus oil/ammonia).
All odours were presented to the subjects via smelling strips and a blank strip was used as a control.
The results were analysed using a multivariate analysis involving principal components analysis (PCA).
Results will be presented from PCA analyses of EEG activity taken from the 28 recording sites on the scalp
and psychometric data derived from bipolar psychometric ratings for strength, pleasantness and familiarity.
The data will be discussed with respect to the cortical topographies obtained.
374
ECRO IX: Abstracts
6. Deficits in odor sensitivity to formaldehyde due to cigarette smoking

Birgitta Berglund and Steven Nordin1
Department of Psychology, Stockholm University and Institute of Environmental Medicine, Karolinska Institute,
Stockholm and 'Department of Psychology, Stockholm University, Sweden
Due to inconsistent empirical data on impaired odor sensitivity caused by tobacco smoking, the view that
smoking gives rise to changes in odor sensitivity in general, cannot be taken for granted. There are, however,
indications that the effect of tobacco smoking on olfactory perception might be selective, affecting only the
odors of substances contained in tobacco smoke. For this reason, odor detection sensitivity and perceived
odor intensity to formaldehyde (of which tobacco smoke has a relatively high content) among cigarette smokers
and a control group were studied. The interest in formaldehyde is accentuated by the fact that it is one of
the main causative agents in the 'sick building syndrome'. To investigate the odor sensitivity, 22 female
cigarette smokers and 22 aged-matched female non-smokers participated in an odor detection experiment
(experiment I) and in an odor magnitude estimation experiment (experiment II). The study was performed

with a controlled dynamic flow olfactometer with a dilution system of steel capillaries with different dimen-
sions. In both experiments, 18 concentrations of formaldehyde ranging from 7.5 to 1000 p.p.b. (0.125 log
unit steps) were presented 12 times each in a randomized order. In order to obtain detection data corrected
for false alarms, 108 inter-randomized clean-air stimuli were presented (for experiment II this number was
36). To correct the magnitude estimation data for individual scaling behavior, seven levels of pink noise
(42—96 dB[A]), each level repeated 12 times in a randomized order, were inter-randomized among the odor
stimuli in experiment II. This correction principle (a version of the Master Scale transformation) states that
the difference between a normal-hearing individual's psychophysical auditory noise function (in double
logarithmic coordinates), with respect to its slope and intercept, and the same type of function empirically
obtained from a group of normal-hearing observers (in a previous study), is due to that individual's scaling
behavior. The same individual's psychophysical odor function is then corrected by a transformation of the
slope and by an intercept transformation (to the same extent as that individual's noise function differs from
the empirically obtained noise function for the normal-hearing group). In this case, the transformation is
based on the assumption that the two groups of observers (smokers and non-smokers) do not differ with
respect to auditory sensitivity (controlled by audiograms). After correcting the odor detection data for false
alarms and the odor magnitude estimates for individual scaling behavior, the results demonstrated a signifi-
cant loss in odor detectability as well as in perceived odor intensity among the group of smokers.
7. The effects of odorants on olfactory mitochondria

Rosemary Bland, Stan Lovett and George H.Dodd
Olfaction Research Group, University of Warwick, Coventry CV4 7AL, UK
The effects of perfumes and some individual perfume components on liver, heart and brain mitochondrial
&\p have been described. This paper describes the effect of a number of odorants on rat hepatic and olfactory
mitochondrial membranes, using the fluorescence indicator method (PSI-SCREEN). The following odorants
were investigated; /J-ionone, isosafrole, safrole, eugenol, 3-trifluoromethylpyridine (TF), TFNO, menthone,
cineole, L-carvone, D-carvone, isophorone, linalool, citronellol, geraniol and pulegone.
8. Molecular reaction cascades in olfactory signal transduction

Ingrid Boekhoff, Sabine Gdggerle and Heinz Breer
University of Stuttgart-Hohenheim, Institute of Zoophysiology, 7000 Stuttgart 70, FRG
In contrast to the well understood reaction cascade underlying visual signal transduction, little is known about
the molecular basis of recognizing specific molecules and the primary events mediating the transduction of
odorous signals into electrical responses. Several lines of evidence suggest that olfactory transduction may
involve second messengers. However, it has never been demonstrated that the level of second messenger
actually changes according to odorant stimulation in a short time range relevant for the transduction process.
Using a rapid kinetic methodology and radiochemical binding assays the formation of second messenger
induced by pheromones and odorants in insect antennae and rat olfactory cilia was monitored in a subsecond
time range. The data indicate that olfactory stimuli induced a significant increase in second messenger con-
centration within 50 ms after odorant application; thereafter the signal rapidly attenuated to the basal level
within 100-200 ms. This rapid and transient molecular signal thus, clearly precedes the generator current
of the receptor cells. Whereas in insect antennae pheromones induced exclusively the formation of IP 3 , in
rat olfactory cilia a selective accumulation of cither cAMP or IP 3 was observed after stimulation with in-
375
ECRO DC: Abstracts
dividual odorams. However, the two different second-messenger pathways could not be correlated to classes
of odor quality. In all cases the odorant-induced formation of second messengers was found to be mediated
by G-proteins, suggesting the existence of specific odorant receptors. The rapid kinetic as well as the tran-
sient nature of the molecular signal favour the concept that receptor-activated chemosensory reaction cascades
are involved in the chemc-electrica! transduction of olfactory receptor cells.
9. The orientation of olfactory bulb mitral cell apical dendrites

Nathalie Buonviso, Michel Chaput and John W.Scott1
Laboratoire de Physiologie Neurosensoriclle, University Claude Bernard, Lyon I, Vtlleurbanne 69622, France
and Department of Anatomy and Cell Biology, Emory University, Atlanta, GA 30322, USA
The mitral cells of the rodent olfactory bulb have a single apical (or primary) dendrite that enters a single
glomerulus where it receives synaptic contacts from olfactory nerve axons. Some investigators have sug-

gested that each glomerulus responds in a unique manner to odors. Therefore, it is of interest to know whether
the mitral cells innervating a particular glomerulus are contiguous. To study this question, we labeled mitral
cells of rat olfactory bulbs by small injections of horseradish peroxidase into the external plexiform layer.
This material is taken up and transported throughout the somata and dendrites of die cell. The brains were
processed with the cobalt-intensified diaminobenzidine reaction, which enabled reconstruction of the den-
drites. We reconstructed cells from this series of 12 brains with injections in the ventral part of the olfactory
bulb. The results were compared with new reconstructions of mitral cells from an older series of injections
into the lateral and medial faces of the olfactory bulb. Contrary to our original expectation, we found that
mitral cell apical dendrites are not generally perpendicular to the mitral cell layer. The apical dendrites of
cells caudal in the bulb are nearly perpendicular, but as one looks at more rostral cells one finds that the
apical dendrites enter glomeruli progressively more rostral to the cell body. We found a range of variation
in the contacts between mitral cells and their glomeruli. Often adjacent mitral cells innervated the same
glomerulus but in many other cases the apical tufts of adjacent mitral cells might be separated by two or
three glomeruli. Preliminary analyses suggest regional differences in the precision of these connections. These
relationships must be considered in interpreting the spatial organization of inputs to mitral cells.
10. Odor identification: filling in some blanks

William S.Cain, Franc T.Schiet, Christine A.Lulejian and Ren£ de Wijk
John B.Pierce Laboratory and Yale University, New Haven, CT 06519, USA
The task of odor identification holds interest in part because it figures prominently in clinical evaluation
of smell. For that reason, among others, we need to understand the basis for success or failure at the task.
Five issues received attention, each in a separate experiment. Two got attention (first and second experiments)
because of what people express when they fail to identify an odor: (i) that they could identify it on another
day, and (ii) that a recognition test would vindicate their level of certainty about whether they actually know
the odor. The data supported the first conclusion, i.e., that performance fluctuates enough that what might
prove unidentifiable one day may prove identifiable the next, but did not support the second conclusion of
an association between feeling of knowing and actual ability to recognize an unidentified odor. A control
task with trivia questions found an association between feeling of knowing and recognition. The third ex-
periment explored whether attribute ratings via a list of 146 odor descriptors would precipitate identification
of items a person had failed to identify. The ratings proved ineffective in that role, i.e. repeated smelling
and the availability of a rich vocabulary produced no gain in identification. The fourth experiment dealt with
whether training subjects to identify one set of odors would facilitate identification of another. The data im-
plied no transfer of training and suggested that the layman must learn to identify odors one at a time. The
fifth experiment dealt with the relation between the ability of a person to discriminate and his ability to iden-
tify odors. After reduction of initial problems with retrieval odor names, a strong association emerged bet-
ween identification and discriminating. In appropriate circumstances, failures of identification seem to reflect
failures of discrimination. (Supported by NIH grant DC00284).
11. Optical monitoring using a voltage-sensitive dye of the piriform cortex responses
to olfactory bulb stimulations in the in vivo rat
Martine Cattarelli, Philippe Litaudon and Michel Vigouroux
Physiologie Neurosensorielle, CNRS-Universitf Claude Bemard/Lyon, 69622 Villeurbanne cedex, France
In the rat, the piriform cortex receives direct projections from the olfactory bulb via the lateral olfactory
tract. As is well-known, these projections are largely divergent and convergent via numerous collaterals.
376
ECRO IX: Abstracts
Our understanding of the olfactory information processing at the piriform cortex level should be improved
if the spatial and the temporal dimensions of its activity could be studied simultaneously on the largest possible
area. Henceforth, optical monitoring of neuronal activity using voltage-sensitive dyes [Cohen et al. (1978)
Rev. Physiol. Biochem. Pharmacol., 83, 35; Grinvald et al. (1988) Physiol. Rev., 68, 1285] allows us to
record the spatio-temporal activity of a whole brain area of several millimeter square of surface. This method
is based on measurements of changes in fluorescence (or absorption) of a brain area stained with a voltage-
sensitive dye. In our device a photodiode array is used as detector and offers a fast time resolution ktlz.
(7 kHz) and a good enough spatial resolution (each photodiode records the activity of a square area of 280
micrometres on a side). As it is a fully parallel acquisition system a microcomputer (PC/AT) can be used
without sacrificing the performances and without important investments in terms of money and time for soft-
ware development. This device was brought into working order with a 4-photodiode array. The first ex-
periments performed on anaesthetized and curarized rats showed that the responses of the piriform cortex
were correlated with the intensity of the stimuli; their latencies increased along the antero-posterior axis

of the cortex. On a same cortical site signals which were different in their shape and their latencies were
recorded in response to different patterns of electrical stimulations delivered in the olfactory bulb. Thus,
information from the olfactory bulb does not seem uniformly distributed on the whole piriform area. In some
cases an increase in fluorescence appeared after the initial decrease (i.e. depolarization) usually observed.
In the near future a 144-photodiode array will be used. Thus, the activity of about 11 mm2 of piriform cor-
tex area will be recorded simultaneously and will allow us mapping, and comparison, of the spatio-temporal
distribution of the piriform cortex activity induced by different patterns of stimulation. The next step will
be to test the effect of biologically meaningful odorants on the piriform cortex activity. Acknowledgement.
The authors are grateful to A.Grinvald and R.Hildesheim for having provided them with the dye used in
these experiments.
12. Single cell recordings from tsetse antennae

C.J.Den Otter and W.M.Van der Goes van Naters
Sensory Physiology Group, Department of Animal Physiology, University of Croningen, The Netherlands
Action potentials from individual cells and receptor potentials were recorded from antennae of males and
females of Glossina m. morsitans and G. f. fuscipes using the 'surface-contact' technique [Den Otter et al.
(1980) J. Insect Physiol., 26, 465]. Stimuli were tsetse olfactory attractants identified in ox breath (carbon
dioxide, acetone, l-octen-3-ol) and in ox and buffalo urine (eight different phenolic compounds [Bursell
etal. (1988) Bull. Em. Res., 78, 281]). In addition, the effect of temperature changes on cell activities was
studied.
It was shown that all phenolic compounds stimulated one and the same type of receptor. In addition to
these cells, receptors were found which were selectively activated by either CO^, acetone, l-octen-3-ol or
a decrease in temperature. Not more than 10% of all cells tested appeared to be sensitive to two or more
of the olfactory stimuli. Some (20%) of the cells did not respond to any of the stimuli used.
Some antennal cells showed spontaneous action potential activity; others were silent when not stimulated.
Two types of olfactory cells were present, one type showing increased activity during stimulation only, the
other type firing spikes not only during, but also for several seconds after stimulation.
No indication was found that cells sensitive to the same stimulus were clustered in certain areas of the
antennae. Moreover, each cell type was found on both the inner and outer faces of the antennae.
13. The effect of self-, cross- and mixture adaptation on human olfactory reaction
times and perceived intensities
Ren* A.de Wijk1-2 and Ronald R.Nixdorf'
'University of Utrecht, The Netherlands and 2John B.Pierce Foundation Lab., New Haven, USA
Adaptation processes are investigated for four single odorants (1,8 cineole, n-hexane, n-butanol and geraniol)
and their binary mixtures. Olfactory reaction times and perceived intensities are recorded. Reaction times
become always longer after adaptation. Perceived intensities, however, are sometimes enhanced after adapta-
tion. Enhancement effects are frequently observed in case of different odorous qualities of adapting- and
test stimuli.
Furthermore, results for single test stimuli show that (i) effects of self-, cross- and mixture adaptation
are all highly correlated, i.e. an odorant that receives a relatively large self adapting effect, will also receive
relatively large cross- and mixture adapting effect; (ii) effects of self- and mixture adaptation are of similar
377
ECRO IX: Abstracts
magnitudes, and the magnitude of the effect of cross adaptation is smaller; (iii) effects of adaptation on reac-
tion times and intensities are, in general, positively correlated. Results for mixture test stimuli show that:
(i) effects of self adaptation are larger than effects of adaptation to one of the components of the mixture;
(ii) the adapting effect received by a mixture results probably from the action of both mixture components;
(iii) the quality of a binary mixture changes towards the quality of one component, after adaptation to the
other component. Our results indicate that reaction time measurements are well suited to the investigation
of olfactory processes.
14. Interpreting generalized Procrustes analysis 'analysis of variation' tables

Garmt Dijksterhuis and Pieter H.Punter
OP&P Sensory Research, PO Box 141 67, 3508 SG Utrecht, The Netherlands
The field of generalized Procrustes methods is still in development, and in 1988 a new method was published

by Peay. This method is implemented in the Procrustes-PC program. Because there are a number of GPA
software packages using different methods, it can be useful to be able to compare the results obtained by
different analysis and by different software packages. Most GPA software report the Procrustes analysis
of variance in absolute terms, using original or re-scaled units. This poster proposes measures relative to
the total amount of variation in the data, which are easier to interpret and are related to the VAF (variance
accounted for) measure and the percentage 'explained' variance. A relative measure makes it easier to com-
pare solutions from different Procrustes analyses made by different GPA software and makes the interpreta-
tion of the solutions more meaningful. The 'analysis of variance' tables produced by all GPA software can
be standardized using the relative measure proposed in this poster. These tables can be of great help in inter-
preting the solution of the analyses; they show the relation between the fit of the solution, the dimensionality,
the agreement between the judges and the agreement about the products.
15. Preimaglnal conditioning of a solitary bee to nest odors

Heidi Dobson and Lennart Agren
Ecological Research Station of Uppsala University, Olands Skogsby 6280, S-386 00 FOrjestaden, Sweden
Some solitary bee species show narrow host —plant associations, restricting their feeding to flowers of only
one or a few closely related plant species over successive yearly generations. The bee's initial recognition
of its host plant has been suggested to be partly due to an olfactory conditioning, in the preimaginal period,
to host—flower volatiles emanating from the larval food provisions (mainly pollen, mixed with nectar). As
a first step to test this hypothesis, we tried to condition Colletes fulgidus (Apoidea, Colletidae) to various
volatiles artificially added as environmental odors during the period from pupation to adult emergence. This
developmental stage was chosen because, in honey bees, it has been implicated by other workers to corres-
pond to a 'sensitive period' for chemical imprinting. Conditioning to the volatiles was evaluated both by
electroantennography and behaviorally using choice feeding tests. Results from EAG tests did not differ
significantly between control and experimental groups. The analyses thus show the added odors to have no
measurable effect on the ability of either male or female antennae to perceive the volatiles. In the behavioral
studies, evidence of preferences for conditioning volatiles was inconsistent and varied between chemicals
tested. In general, it suggests that bees do not become conditioned to nest volatiles during the pupal stage.
Currently under investigation is the question of whether bees undergo olfactory conditioning to host-plant
chemicals during the larval stage—when host-flower odors from the food provisions are detectable in the
nest by the human nose—and that the effects are carried over to the adult stage.
16. Spatial coding of olfactory information in fish

Kjell B.Duving
Department of Biology, University of Oslo, PO Box 1051, 0316 Oslo 3, Norway
Functional, morphological and behavioural studies of the olfactory system in teleost fish illustrate the spatial
concept of coding. Evidence for this concept can be demonstrated at different levels. In the epithelium, dif-
ferent types of receptor cells are inhomogeneously distributed and the relative size of EOG responses is depen-
dend upon electrode position [Thommesen (1983) Acta Physiol. Scand., 117, 241]. The spatial distribution
of the responses to odorants is evident in the olfactory bulb in salmonids [Deving el al. (1980) Acta Physiol.
Scand., 108, 123]. Here, bile salts induce responses in the medial and amino acids in the lateral part of
the olfactory bulb. In cod the spatial arrangement of the connections between the olfactory bulb and brain
378
ECRO IX: Abstracts
gives an excellent support for the spatial concept of quality coding. The olfactory tract bundlets mediate
different behaviour patterns [Deving and Selset (1980) Science, 207, 559], as stimulation of the lateral olfactory
tract evokes feeding behaviours while stimulation of the medial olfactory tract evokes courtship and alarm
reactions. The origin of these neurones in the bulb has a restricted distribution in the olfactory bulb and
connects to a restricted set of receptor cells. The bundlets of the lateral olfactory tract respond to the substances
glycine and taurocholate that evoke feeding behaviours. The tract bundlets terminate in different areas of
the telencephalon.
17. P300: a new technique of recording a cognitive component in the olfactory

evoked potential
M.Durand-Lagarde and G.Kobal
Institut fir Pharmakologie und Toxikologie, UniversitOtsstrafie 22, 8520 Erlangen, FRG

The cognitive component P300 was recorded in human event-related potentials (ERP) during a discrimina-
tion task where the subjects had to discriminate low-probability stimuli from high-probability stimuli. The
aim of the present study was to demonstrate that this endogenous cortical response can also be recorded
in an olfactory discrimination task, after the problem of producing the requisite short stimulus intervals had
been solved. Sixteen volunteers (eight male and eight female) between 20 and 36 years of age (mean 26.69;
SD: 3.84) participated in the experiments which consisted of four different randomized sessions. A mixture
of vanillin and H2S was used as standard stimulus (p = 0.85, 200 ms duration). Target stimuli (p = 0.15)
were either H2S or vanillin. Both types of targets were used for stimulation of the right and the left nostril.
Stimulus intervals could be reduced to 8 s. This still allowed the subjects to perceive stimulus onsets. EEG
was recorded from seven positions (10/20 classification) referenced to linked earlobes. Subjects were asked
to count the rare stimuli. A P300 component was recorded in responses to the rare stimuli, in all subjects
and with all type of targets. P300 amplitudes were significantly larger after H2S target stimuli. The recording
of P300 is regarded as a useful technique for testing olfactory discrimination capability, e.g. in Parkinson
or Alzheimer patients.
18. Projection of the primary olfactory neurons in cod

Anne Figenschou
Specific behaviour patterns can be evoked in cod by electrical stimulation of olfactory tract bundlets [Deving
and Seise (1980) Science, 207, 559]. This feature permits a description of the anatomical and functional
properties of the substrate that participates in the execution of each behaviour pattern.
In the present investigation the projection of the primary receptor cell onto the olfactory bulb was studied
by the use of the fluorescent carbocyanine dye 'Dil' (molecular probes). The dye was applied on limited
areas of the dorsal part of the olfactory bulb to see where on the epithelium the marked neurons originate.
The preparation was kept at 40°C in 4% paraformaldehyde. Eight weeks later the olfactory lamellae were
dissected free, separated in anterior and posterior surfaces, and mounted flat on glass slides. Inspections
of the lamellae in a fluorescent microscope showed single receptor cells as bright luminescent spots.
Fluorescent receptor cells were found in every lamella examined, both in anterior and in posterior sur-
faces. Application of 'dil' on the bulb also resulted in coloration of the secondary neurons. Local application
of the dorsal region gives fluorescence in the LOT only, demonstrating the specificity of connections between
this part of the olfactory bulb and the olfactory tract.
Systematic variation of the applications site on the bulb might reveal the topographical relationship between
the olfactory receptor cells and the olfactory bulb.
19. Odour processing in the bee brain studied at the level of identified antennal
lobe interneurones
Caroline Fonta
LNC1INRA-CNRS, 91440 Bures sur Yvette, France
Intending to analyze processing of odours in the CNS of the honeybee, intracellular recordings and stainings
have been applied to the antennal lobe interneurones in animals stimulated with pure odours (geraniol, hep-
tanone, isoamyle acetate) and their mixtures.
379
ECRO IX: Abstracts
The 200 neurones studied were classified into four main morphological groups: two types of local
intemeurones differing by their glomerular pattern of arborizations and two types of output neurones branching
either in one glomerulus or in many glomeruli. Such classification somehow matches with special characteristics
of olfactory responses among these antennal lobe neurones: the local intemeurones which have concentrated
arborizations mainly in one glomerulus and sparse arborizations in other invaded glomeruli and the
uniglomerular output neurones display stronger and more discriminating responses than the other neurones
(local and output neurones which arborize in many glomeruli in a more homogeneous pattern). Such results
suggest that the spatial dimension is important in odour coding assumed by the considered neuronal network
and they support the hypothesis of the glomerular functional unit [Masson and Mustaparta (1990) Physiol.
Rev., 70, 245.].
Besides, the processing of odour mixtures compared to single components coding appeared as a result
of more or less complex interactions between the antennal lobe neorone activities, with an important role
of inhibitory processes.

20. Stimulus-induced respiratory mucosal potentials are sensitive to capsaicin
pretreatment in rats
I.Friedel, N.Thurauf and G.Kobal
Instina fir Pharmakologie und Toxikologie, UniversiUUsstrafie 22, D-8520 Erlangen, FRG
In anesthetized rats electrical responses [Kobal (1985) Pain, 22, 151] were simultaneously recorded from
the nasal septum and the cerebral cortex after painful stimulation with carbon dioxide. Stimuli were presented
by aid of a device which allowed a precise definition of the stimulus parameters. The peripheral potential
obtained from the mucosa of the nasal septum (PMP) was easy to reproduce in rats and proved itself to
be a physiological signal, the presence of which is closely connected to functionally unimpaired C-fibres.
After local as well as systemic administration of capsaicin the PMPs were nearly extinct; this coincided with
a complete elimination of the evoked potentials recorded from the cerebral cortex. Thus, it can be assumed
that peripheral potentials from the nasal mucosa provide an excellent characterization of peripheral nocicep-
tion, and could well be employed to measure, in an objective and non-invasive way, the quantitative and
qualitative action of analgesics in man. (This work was supported by grant DFG Ko 82/1-4.)
21. Cell-cell interactions in developing bee antennal lobe

J.Gascuel, D.J.Beadle1 and C.Masson
Laboratoire de Ncurobiologie Comparie des Inventbris. INRA-CNRS URA, 1190, Bures sur Yvette 91440,
France and 'Laboratory of Molecular and Biological Sciences, Oxford Polytechnic, OX3 0BP, UK
The bee antennal lobe seems to be a particularly suitable model for exploring basic issues in developmental
neurobiology, especially studying the mechanisms by which specific cell-cell interactions lead to the for-
mation of appropriate synaptic connections and the establishment of the final neuronal network. Develop-
ment of the antennal lobe during the pupal stage, has been studied under normal conditions and compared
with development after early de-afferentation or during olfactory deprivation. The results demonstrate firstly
that sensory neurones trigger the morphogenesis of the glial cells which set up the glomeruli, and suggest
that contacts between glial cells and antenna] lobe neurones are determinant for the arborizations of the antennal
lobe neurones within the glomerulus. Secondly, the results show that a strong decrease in synapse frequency
occurs immediately after a burst of synaptogenesis. Moreover, an olfactory deprivation applied during the
same period results in a lower synapse frequency when compared to the controls. Together, the results strongly
suggest that the sensory afferents (i) could act in the setting up of the glomerulus mediated by cell—cell
contacts or by a release of diffusible molecular factors and (ii) control the development of the connectivity
through the neuronal activity. In the goal to analyse, at a molecular level, how the sensory neurones act
in the setting up of the glomerulus, primary cultures and co-cultures have been set up. Cellular and physiological
aspects of the differentiation of the different cell types have been studied. The results will be presented and
discussed in regard to in vivo data.
380
ECRO IX: Abstracts
22. Neurobiology of the olfactory memory: cellular and Integrative aspects

R.Gervais
Lab. Physiologic Neurosensorielle, URA CNRS 180, Universal Claude Bernard, Lyon-l, 69622 Villeurbanne,
France
Over the last decades an enormous number of studies have been devoted to the neurobiology of learning
and memory, but a very limited number of them have paid attention to the olfactory memory. This phenomenon
is ascribable to a relative poor understanding of coding in olfactory pathways and preferences of experimen-
tors for visual and auditory modalities, considered as being dominant in the human. However, for reasons
presented by the main speaker of the session, some recent data now allow us to consider the olfactory system
of lower mammals (rabbit, rat) as a suitable model for investigating mechanisms supporting plasticity and
learning.
The first part of the discussion points out functional similarities which are specific to peripherical and
central olfactory pathways and could present common mechanisms for the expression of neural plasticity.

These include long-term changes in synaptic transmission and cellular excitability as well as persistancy in
the adult animal of proteins (FOS, GAP-43 . . .) normally present in the neurones during development and
functional reorganization. Moreover, the involvement of the noradrenergic system in plasticity in telencephalic
structures including olfactory areas have to be underlined.
The second part of the discussion focuses attention on the anatomy of the olfactory memory, the aim being
the comprehension of the respective roles of olfactory bulb, piriform cortex and hippocampal formation in
acquisition and retention of the information. We can see that the present data lead to a puzzling view which
remains to be clarified. Because underlying mechanisms are based on complex interactions between large
neural assemblies and because different types of olfactory learning (one-trial learning; associative learning
. . .) involve different time scales, elucidation of this question represents a major challenge.
On the one hand, present data suggest a relative strong conservatism in mechanisms supporting plasticity
in olfactory structures with respect to other forebrain areas. On the other hand, the olfactory system is likely
to support an original network functioning for storage and retrieval of sensory information.
23. Gustatory responses in primates to the 'sweetener' aspartame and their

phylogenetic implications
D.Glaser, H.van der Wei, J.N. Brouwer, G.E.Dubois1 and G.Hellekant2
Anthropological Institute, University of TMrich-lrchel, CH-3057Zurich, Switzerland, 'NutraSweet Co., hit
Prospect, IL 60056 and 2Department of Veterinary Science, University of Wisconsin, Madison, Wl 53706,
USA
Electrophysiological and behavioural methods have been applied to more than 60 species of the ordo Primates
and, for comparison, to the tree shrew (Tupaia glis) to determine their responses to the first known sweet-
tasting dipeptide aspartame which has to man a sweetness intensity of about 200 times that of sucrose. However,
only the Cercopithecidae, the Hylobatidae and the Pongidae respond to this dipeptide as does man, and they
prefer aspartame to water. The other primates tested so far show no response to this 'sweetener'. From a
phylogenetic point of view we suggest that aspartame shows characteristics similar to thaumatin, which con-
stitutes a clear dichotomy within the ordo Primates. This similar effect occurs, although the molecules of
aspartame (mol. wt 294 daltons) and thaumatin (mol. wt 22 000 daltons) seem to be quite different. However,
maybe these two molecules have the same binding site(s) to the sweet receptor? The results here stress once
more the diversity among primates and can be a help in the discussion of the existence of single or multiple
sweet-taste receptors.
24. Specificities of pheromone receptor neurones in three species of Dendroctonus

S.Graabrack, H.D.Scharf1 and H.Mustaparta
Department of Zoology, University ofTrondheim, DragvoU, Norway and 'Lehrstuhl II, Organische Chemie,
RWTH, Aachen, FRG
Bark beetles of different genera live together in crowds on the same host tree or on neighbouring trees. In
field experiments interspecific interactions have been observed, both within a genus and between genera.
Furthermore, electrophysiological studies in species of Ips have demonstrated the presence of specific receptor
neurons for pheromones produced by Dendroctonus species.
381
ECRO IX: Abstracts
In the present investigation it has been recorded from single olfactory receptor neurones of three species
of the genus Dendroctonus, D.ponderosac, D.valens and D.terebrans, in order to determine the receptor
specificities for their pheromones and interspecific chemical signals. These compounds are enantiomers of
ketales and monoterpcnes.
In general, the results show, as in species of Ips, that the receptor neurones are tuned to a single compound
of which the optical configuration is important. Thus, the three species possess several types of receptor
cells, each tuned to e.g. (+)-ctt>-brevicomin, (-)-frontalin, (+)-ipsdienol and (-)-rranj-verbenol. Each
type of receptor neurone was similar in all tree species.
Thus, in both sexes of D.ponderosae the majority of receptor neurons responded specifically to (-)-trans-
verbenol which is important for the aggregation of this species. In addition, other types of receptor neurones
were tuned to (+ )-ipsdienol, amitinol and ( — H ro ntalin. In D.valens, the majority of receptor neurones in
males responded specifically to (—)-frontalin, whereas females possessed a majority of neurones tuned to
the Ips compound ( + )-ipsdienol. This suggests that (-)-frontalin may be an important pheromone com-

pound in D.valens, whereas (+ )-ipsdienol acts as an interspecific chemical cue between species of Ips and
D.valens. The pioneering D.valens females may be influenced by (+Hpsdienol from neighbouring Ips species
when orientating towards and selecting host trees. In D.terebrans no pronounced groups of receptor neurones
were found. In this species it was also found that cells tuned to (+)-ipsdienol.
In conclusion, the various bark beetle pheromone compounds are received by separate receptor neurones
in the three species of Dendroctonus, each neurone tuned to a particular optical configuration. Furthermore,
Dendroctonus species are able to receive information from signals that are characteristic for the genus Ips.
Thus the results suggest that the chemical interaction between Ips and Dendroctonus is mutual.
25. Inhibition of the mitochondrial respiratory chain by perfumes and perfume

components
D.E.Griffiths
Department of Chemistry, University of Warwick, Coventry CV4 7AL, UK
Using a cytocicity assay called the psi-screen, a simple fluorescence assay to measure mitochondrial mem-
brane potential, (A+ m ), a screening system has been developed for drugs, toxins, agrochemicals and poten-
tial neurotoxins.
Fifty-two perfumery preparations (perfume oil, perfume, eau dc parfum and eau de toilette) from British,
French, Japanese and US perfumers have been examined. All are active in the psi-screen assay and the associated
NADH-oxidasc assay at dilutions in the assay of 1:3000-1:30 000. The inhibitory effects on mitochon-
drial NADH oxidasc are equivalent to 0.05-0.5 mM rotenone, and the site of inhibition is similar to that
of potent lipophilic inhibitors of NADH —ubiquinone reductase, such as rotenone, piericidin, capsaicin and
the neurotoxin l-methyl-4-phenylpyridinium. These inhibitors interact with a hydrophobic pocket in the
NADH -ubiquinone reductase complex, which may serve as a receptor site for lipophilicodorant molecules,
thus regulating NADH-ubiquinone reductase activity. The key role of NADH -ubiquinone reductase com-
plex in cell respiration and the generation of cellular membrane potentials (A*^ and A*p), together with
this capacity to regulate cell calcium will be discussed as well as studies of the effects of individual perfume
components on liver, heart and brain preparations.
26. Olfactory receptors in Crustacea

Eric Hallberg, Kjell U.I.Johansson and Rolf Elofsson
Department of Zoology, University of Lund, Sweden
Traditionally the aesthetascs have been presumed to constitute the olfactory organs in crustaceans. The
aesthetascs have been described several times in higher crustaceans, the decapods, and exhibit here uniform
structural characteristics such as large numbers of receptor cells and enveloping cells. The receptor cell gives
rise to two cilia that spilt into small branches that fill the interior of the thin-walled hair. Pores are lacking,
but moulting pores can be present at the apex of the hair. Variations of this scheme occur in other crusta-
ceans and are expressed in the reduction of the number of sensory and enveloping cells, as well as variable
numbers of cilia and inconsistencies in their branching patterns. Due to this structural variation the defini-
tion of the aesthetascs has become less clear, and physiological investigations of a chemoreceptor seem to
be required to unequivocally determine its functional mode. A classification on physiological grounds can,
however, be difficult to achieve in many species.
382
ECRO IX: Abstracts
27. Morphological correlates of functionally identified antenna] lobe neurons in

adult male Manduca sexta
Bill S.Hansson1, Thomas A.Christensen and John G.Hildebrand
Arizona Research Laboratories, Division of Ncurobiology, University of Arizona, Tucson, AZ 85721, USA
'Present address: Department of Ecology, University of Lund, S-223 62, Lund, Sweden
The antennal lobes in the brains of male moths have an area that is specialized for the processing of informa-
tion about female sex pheromones, the macroglomerular complex (MGC). Olfactory interneurons with
arborizations in the MGC were examined in male Manduca sexta by tandem electrophysiological recording
and intracellular staining with Lucifer Yellow. Neurons that responded selectively to the major pheromone
component, (£,Z)-10,12-hexadecadienal (bombykal), had dendritic arborizations restricted to a toroid-shaped
subdivision of the MGC. Neurons that responded selectively to an established mimic [(6£,Z)-11,13-
pentadecadienal)] of another important pheromome component, (£,£,Z)-10,12,14-hexadecatrienal, had

arborizations confined to a different subdivision of the MGC, distinct from the toroid. One neuron that responded
to both pheromone components clearly had dendrites in both MGC subdivisions. The different morphological
subdivisions of the MGC appear, therefore, to be functionally distinct units as well, each specialized to process
information about one pheromone component. (Supported by NM grant AI-23253 and by postdoctoral
fellowships to BSH from the Swedish Agricultural and Forest Research Council and from the Fulbright
Program.)
28. Ion channels from insect olfactory receptor neurons

H.Hatt and F.Zufall
Physiologisches Institot der Technischen UniversitOt MOnchen, Biedersteiner Strafie 29, 8000 MOnchen 40, FRG
Use of different modalities of the patch—clamp technique has made it possible to identify and characterize
the nature of the conductances present in somatic and outer-dendritic membranes of pheromone receptor
neurons of Antheraea polyphcmus. Records of single-channel currents have shown the presence of several
types of channels selective to potassium ions on the soma membrane. In addition, non-specific cation channel
could be identified that can be activated by pheromone or female gland extract The selectivity for cations
would be consistent with depolarization that might play an excitatory role. We have some indications that
second messengers (IP3) might be involved either directly or indirectly in the activation and/or desensitiza-
tion of this pheromone sensitive channel. (Supported by the DFG 1201/2-1.)
29. On the question of four basic tastes (qualities)

Goran Hellekant
University of Wisconsin, Department of Veterinary Science, and Wisconsin Regional Primate Center Madison,
Wl 53706, USA
The concept of taste basics, taste qualities or taste primaries (basic, quality or primary are here used as
synonyms) is an essential tool in taste research. It not only serves as the basis for many physiological and
biophysical investigations in the mechanisms of taste, but is also used for describing and understanding the
physical nature of flavor development of foods, the effects of aging, storage, and alternative processing.
Ail tastes are transmitted as impulses in nerve fibers. The question asked in this study; how well would
a number of single taste nerve fibres conform with the categories of sweet, sour, bitter and salty if the response
to a large array of compounds that on psychophysical bases fall into these categories is used as taste stimuli
in man's closest relative, the chimpanzee?
We have obtained approximately 25 single-fiber recordings in five chimpanzees to 11 different sweeteners,
three acids, three bitter compounds and five different salts, and have used the selective sweet taste blocker
gymnemk acid as well as the sweet taste inducer, miraculin, on some of these fibers. One of the most striking
finding is how well the single fibers fell into one or the other of the taste qualities (basics). For example,
it was very easy to find taste fibers that were highly specific to sweet compounds and responded to 100%
of the sweeteners used.
Although our material of single fiber recordings is limited it is evident that the psychophysical concepts
of taste qualities have their counterparts in the way the peripheral taste information is distributed among
individual taste fibers in the chimpanzee. In comparison with our experiences from other species, including
the rhesus monkey, these categories seem more pronounced and delineated.
383
ECRO IX: Abstracts
30. Single-point versus time—intensity ratings of sensory attributes in breads and

fat spreads
U.Hellemann, L.Vainio1, H.Tuorila, N.Barylko-Pikielna2 and I.Matuszewska2
University of Helsinki, Department of Food Chemistry and Technology, 'University of Helsinki, Department
of Nutrition and 2Polish Academy of Sciences, Sensory Analysis Laboratory
Single-point and time - intensity measurements were compared for the information yielded by both methods.
Three wheat and five sour rye breads with varying NaCl contents (from 0.25 to 1.55 %) were used as samples
in experiment I. Twelve fat spreads (80% fat) with different fat composition (dairy/vegetable) and with varying
NaCl contents (1, 1.5 or 2%) were used as samples in experiment n. Samples were rated for saltiness, sourness
and overall intensity of taste (breads) and for saltiness and melting rate (spreads).
Time—intensity maximum values were strongly correlated with single-point intensity ratings. The cor-
relation between areas under the response curves and single-point intensity values was in many cases lower.

This result supports an argument that in a single-point test, the assessors rate an attribute at the point of
maximum intensity. Generally, area under the curve was less discriminative than time—intensity maximum
and single-point intensity. This may be due to the variation in intensity perception and in time perception,
both contributing to the area under the curve.
Time-intensity measurements of saltiness in breads and fat spreads did not provide more information than
the single-point intensity measurement. On the other hand, area under the curve (time - intensity measure-
ment) gave valuable information about different melting patterns of spreads, which could not be obtained
by single-point intensity measurement.
31. Response properties of deutocerebral pberomone sensitive neurons in Periplaneta

americana
Manfred H6sl
Institut far Zoolgie, Universitat Regensburg, 8400 Regensburg, FRG
When the antenna of male Periplaneta americana is exposed to female sex attractant, deutocerebral pheromone-
sensirive projection neurons preferentially respond to one of the two components: periplanon A or periplanon
B. Of those respondng more strongly to periplanon B (the B neurons), many display a graded preference
for local stimulation. Responses are maximal at stimulation only of a short segment of the 60-mm antenna
and fall off to either side of it. The 50% boundaries include a segment between 15 and 25 mm long. A further
characteristic appears to be that the response of a given B neuron is diminished when equal stimuli are applied
to its central field and simultaneously to a segment some distance away from it. The effect is enhanced when
the stimulus outside the central field is stronger than inside it.
The above responses are all phasic-tonic. Short bursts are elicited exclusively, however, when octanol
and a number of other non-pheromone substances [Waldow (1978) J. Comp. Physiol., 116, 1] are directed
for 1 s at a region within the 50% boundaries. When picrotoxin is added to the Ringer solution bathing the
brain, the bursts become much longer, often several hundred microseconds. Furthermore, if periplanon B
is also directed at another section of the 50% field simultaneously with the octanol stimulus but in the absence
of picrotoxin, the response is also only a short burst without ensuing tonic excitation. In addition, octanol
presented in 50 ms pulses repeated at 200 ms intervals evokes bursts that grow progressively shorter and
soon disappear. This indicates that the inhibitory mechanism underlying the phasic response shows a land
of temporal summation. (Supported by the Deutsche Forschungsgemeinschaft SFB4/B3.)
32. Cortical activity and odours. A longitudinal study using brain electrical activity
mapping (BEAM) to investigate the cortical processing of odours
Sue Hotson and Steve Van Toller
Department of Psychology, University of Warwick, Coventry, England
The technique of monitoring real-time EEG activity in response to odours has become well established by
the Warwick Olfaction Research Group [Van Toller et al. (1989) Chem. Senses, 14, 1; Van Toller et al.
(1990) Psych. Res., 29,429]. This paper presents the work completed in the first year of a three-year longitudinal
study designed to systematically explore the cortical responses to odours of a large group of adult subjects.
Thirty-nine undergraduates form the subject group and have completed two of an anticipated six stages
of the study. All subjects have undergone a brief odour evaluation test to determine at the start of the study
384
ECRO EX: Abstracts
their odour acuity. The second stage of testing involved the use of a Neuroscience Series 111 Brain Imager.
This displays real-time cortical activity in the form of coloured topographical maps.
Subjects were fitted with a headcap containing 28 electrodes in a modification of the international 10/20
placement system with reference to linked earlobes. Testing took place in a low-odour chamber with the
subjects in perceptual isolation. Twelve odourants in total were presented to the subjects on perfumers' smelling
strips. Cortical activity was recorded for a total of 40 s per trial, the presentation of the odour occupying
10 of these. All subjects were tested in the morning and again in the afternoon of the same day, to observe
any diurnal EEG patterns. Additionally, all subjects completed psychometric rating scales and descriptive
questionnaires.
A smaller study of subjects' breathing patterns while being tested is planned but there are certain technical
difficulties to be overcome. One of the major problems encountered so far is handling the enormous amount
of data produced when using this technique. Results so far suggest that despite a high level of individual
variation it is possible to discern common patterns of cortical response to various odours in the 12 - 15 Hz

(alpha) waveband. Statistical analysis of the data is expected to add further support to this theory.
33. Relating structure to function in the peripheral olfactory system of the newborn
rabbit
Robyn Hudson
Institutfilr Medizinische Psychologic, UniversitOt MQnchen, Goethestrafle 31, D-8000 MOnchen, FRG
The relationship between structure and function is one of the enduring themes of biology, and one which
has greatly influenced efforts to understand the neural basis of olfactory function. Developmental change
and plasticity in the peripheral olfactory system provides a particularly good opportunity for a critical
examination of therelationshipbetween anatomical organization on the one hand and function at the whole
animal level on the other. For example, in the rabbit the number of olfactory receptor cells increases from
about 14 million at birth to about SO million in the adult, and is accompanied by an increase both in the
size and number of olfactory glomeruli. Although this growth presumably increases the animal's perceptual
capacities, it also raises the question as to how constancy in odour perception is achieved during such change
in afferent connectivity.
The modified glomerular complex, a well defined structure showing conspicuous 2-DG labelling both in
the newborn rabbit and rat, has traditionally been implicated in the mediation of suckling behaviour. However,
since lesioning this area does not impair suckling performance in either species, this functional interpreta-
tion must DOW be called into question.
Unilateral transsection of the olfactory nerves in newborn rabbits and subsequent re-innervation of the
de-afferented bulb has demonstrated few and very small glomeruli to be sufficient for the recovery of olfac-
tory function. As the location of new connections appears to be rather arbitrary, the re-innervation of par-
ticular glomeruli or there-establishmentof a particular map does not seem to be prerequisite for function. \
Although hand-raising pups following de-afferentation does not affect the occurrence of re-innervation,
at least one suckling experience is necessary for these animals to respond to the maternal pheromome using
there-innervatedbulb alone. Thus, learning and experience may play equally important parts in the process-
ing of odour signals as specific patterns of primary connections. (Supported by the Deutsche Forschungsge-
meinschaft, Po 121/13.)
34. Olfactory evoked potentials in clinical diagnosis

Th.Hummel and O.Kobal
Institutfilr Pharmakologie und Toxikologie, UniversiUUssirqfie 22, D-8520 Erlangen, FRG
One of the major problems in olfactometry was how to obtain data which exceed subjective reports—comparable
to near-field and far-field potentials routinely recorded in audiology. This would be of greatest importance
in cases when patients (e.g. with Alzheimer's disease) are not fully able to report their sensory deficits.
In our laboratory a method was developed by the aid of which it became possible to obtain olfactory evoked
potentials (OEP). After we had overcome the technical problems of how to present suitable olfactory stimuli
another problem arose—namely, to distinguish between responses elicited by chemical stimulation of the
trigeminal nerve and those evoked by chemical stimulation of the olfactory nerve. Only by employing multichan-
nel recordings and magneto-encephalographic methods were we finally able to distinguish between the specific
385
ECRO IX: Abstracts
patterns of those responses. Olfactory evoked potentials tend to be distributed at parietal central sites of the
skull, whereas trigeminal responses (chemosomatosensory evoked potentials CSSEP) can be found at cen-
tral frontal positions. Recent experiments revealed that a classification of odorants relative to their property
to activate one or the other sensory system can be successfully attempted by the subjects' ability to localize
the stimulated nostril. Patients with olfactory deficits of different degrees and causes were investigated in
this study: patients with M.Alzheimer, M.Parkison, Kallman's syndrome, influenza, other types of infec-
tions, and head injury It will be demonstrated that olfactory deficits can be more precisely diagnosed by
employing the technique of olfactory evoked potentials.
35. Model of the rat nasal cavity anatomy by a CAD code

E.Jaillardon1-3, M. Cattarelli2, B.Gay u L.Astic2
'Laboratoire de Mtcanique des fluides et acoustique UA CNRS 263 ECL 69131 Ecully, 2Laboratoire de
Physiologic Neuroscnsoriclle UA CNRS 180 UCBL and 3Centre de Mtcanique UCBL, 69622 Villeurbanne,

France
This work follows the study presented in ECRO 88 on the follow patterns in a rat nasal model. The purpose
of the present summary is to obtain a geometric modeling of the rat nasal cavity in order to generate a finite
element net of the cavity volume. To obtain a geometric modeling of the rat nasal cavity, the CAD code
CATIA was employed in order to obtain a representation of the cavity surface. This representation was obtained
as follows: acquiring data in frontal sections, smoothing the points characterizing every sections, and
constructing the surfaces supported by smoothed curves.
Two kinds of image design are possible with CATIA: facetted surfaces and realistic display of images.
36. Very short-term recognition memory for odors

C.Jehl, J.P.Royet and A.Holley
Physiologie Neurosensorielle, Universilt Claude Bernard, CNRS URA 180, F-69622 Villeurbanne Cidex,
France
As compared with memory for visual and auditory events, olfactory memory is characterized by a slow rate
of forgetting. However, memory for odors has rarely been explored using very short retention intervals,
i.e. the first 60 s following initial odor presentation.
The present experiments aimed at investigating olfactory recognition memory in conditions analogous to
those classically used for making similarity judgements on odors presented in pairs. The question was whether
memories for the first odor of a pair faded from memory store with time elapsed between presentation of
each of the two odors. During memory testing, subjects had to decide whether the second odor of a pair
was identical to that presented first, or different. Eight groups of 12 subjects performed on this memory
task, each group ascribed to one of the eight retention intervals tested (2, 10, 20, 30, 40, 50, 60 and 300 s).
Subjects were presented with two sets of 12 pairs. Each set included six pairs of chemically identical odorous
substances and six pairs of different ones. Among these chemically different pairs, substances were either
closely similar in terms of quality (SI) or highly dissimilar (S2).
For pairs of identical odors, results showed that the number of correct responses (i.e. pairs judged to be
similar when odors were identical) did Dot vary with time elapsed from 2 to 300 s. For chemically different
odor pairs, the number of false alarms (i.e. pairs judged to be similar when substances were dissimilar)
depended significantly on test intervals for closely similar odors (SI). However, this number was lower for
pairs of highly dissimilar odors (S2) than for pairs of closely similar odors (SI): the more odors were dissimilar,
the lower the number of false alarms. In conclusion, odor recognition seems to be more sensitive to qualitative
similarity (SI vs S2) than to time effect. Nevertheless, the temporal factor seems to display an effect on
very-short-term odor recognition memory when odors of chemically different pairs are highly similar in
quality: more false alarms are produced for longer intervals.
Finally, it was shown in a second experiment that retention scores are influenced by subjects' familiariza-
tion with tested odors. For a fixed retention interval (20 s), the number of false alarms rapidly decreased
as a function of the number of presentations (1 to 3) of the first odor of each pair, during a familiarization
session, the day before memory testing. The recognition score is therefore directly related to the level of
odor familiarity.
37. Peripheral and central structures in the olfactory system in mysids (Crustacea)
Kjell U.I.Johansson, Eric Hallberg
Department of Zoology, University of Lund, Sweden
The olfactory system in mysids (Mysidacea, Crustacea) has shown some remarkable features that can be
compared with the olfactory system in both decapod crustaceans and insects. Both decapods and mysids
386
ECRO IX: Abstracts
have chemosensory sensilla on the first antenna, the aesthetascs, which have a similar structure. They are
innervated by biopolar sensory cells whose axons terminate in the olfactory lobes in the brain. The male
mysids have a structure at the peduncle on the first antenna called lobus masculinus, that carries long slender
aesthctasc like sensilla. Each sensillum is innervated by one sensory cell, and the dendrite have two unbranched
cilia with 9 + 0 tubular arrangement.
In mysids, as in some insects, the sexual dimorphism of the first antenna reflects a dimorphism in the
brain displayed by male specific neuropil areas. These neuropil areas arc, in mysids, located anterior to
the olfactory lobes and in silver-stained sections the neuropil appears as a rounded, unstructured region.
It is believed that sensory afferents from lobus masculinus and fibers from intemeurons establish contact
in this neuropil. The paired olfactory lobes in mysids consists of dense neuropil arranged in spherical struc-
tures, glomeruli, which lie scattered in the lobes. The glomeruli resemble similar structures in insects in
having a central core of fibers from intemeurons which is surrounded by a cap of afferent terminals. In
decapods the neuropil of the olfactory lobe consists of tapered columns into which sensory afferents converge.

38. Effects of the odour source distribution on the odour plume structure
G.Karg, A.E.Sauer and U.T.Koch
University of Kaiserslautem, 6750 Kaiserslautern, FRG
The concentration of pheromone and the structure of the odour plume can be considered important parameters
for a successful application of the mating disruption method in pest control. With the help of electroanten-
nogTamms (EAGs) we have investigated the dispersion of the pheromone of the grape wine moths (Lobesia
botrana) in the field. The registered EAG signal generated by a single odour source (dispenser) was found
to depend on wind direction and distance to the detector. Besides changes in the average DC components,
strong fluctuations were detected lying well within the measuring range of our system. In contrast, these
fluctuations were not detected in vineyards treated with pheromone for mating disruption. These results sup-
port the idea that the pheromone plume inside a treated vineyard is a rather homogenous cloud.
39. Pheromonal steroids of male yeUowfin Baikal sculpin Cottocomephorus grewingki

(Dyb.)
P.L.Katsel, T.M.Dmitrieva, R.B.Valeyev, V.A.Ostroumov and Yu.P.Kozlov
Institute of Biology, Irkutsk University, Irkutsk, USSR
It was found that mature males of yeUowfin Baikal sculpin released sex pheromones with the urine during
the spawning period. Three components of male pheromonal signal have been isolated from urine by diethyl
ether extraction, TLC and HPLC. Using mass spectrometry, chromatographic methods and UV spectroscopy
of the sulfuric chromogenes of steroids, we have identified two of them as testosterone (T) and II/3-hydroxy-
testosterone (HT). The concentrations of T and HT in male urine are 1 x 10~9 M and 9 x 10~ 8 M respec-
tively. These steroids are synthesized in testes during full spermatogenesis and penetrate in male urine with
sperm. The adding of water solution of HT (1 x 10~" M) in aquarium with non-mature females in four
hours during 24 h results in about 100% maturing of female oocytes. Water solutions of T
(I x 10""—1 x 10~' 6 M) and HT (1 x 10""—1 x 1 0 " " M) induce the spawning behaviour of mature
females. Other A 4 - 3 keto C19 steroids, such as androstendione and andrenosterone, also induce spawning
behaviour of mature females.
40. The relationship between the detection thresholds and proton spin-spin
relaxation times of the amino acids
Sarah E.Kemp, Gordon G.Birch1 and John M.Grigor1
Monell Chemical Senses Center, 3500 Market Street, Philadelphia, PA 19104, USA ancf' Department of
Food Science and Technology, University of Reading, Whiteknights, Reading, Berks, RG6 2AP, UK
When a taste receptor binds to a sapid substance it exchanges hydrogen bonding with water for hydrogen
bonding with the sapid molecule. This process involves proton exchange. Nuclear magnetic resonance relaxation
rates are due to proton exchange and NMR spectroscopy is, therefore, capable of studying processes on
the level at which they occur at the receptor. NMR has been used to study taste. A trend has been found
between the proton spin—spin relaxation times (T2 values) of sixteen L-amino acids and their detection
thresholds. This indicates that those amino acids which perturb water structure the most are the most easily
detected. It is postulated that at low concentrations, below those at which taste quality perception occurs,
taste receptors may respond to the perturbation of water structure caused by the sapid molecule.
387
ECRO K: Abstracts
41. Olfaction in infancy. A study of human infant olfactory processes by brain

electrical activity mapping (BEAM)
Martin Kendal-Reed and Steve Van Toller
Department of Psychology, University of Warwick, Coventry, UK
The development of the human infant has been studied intensively for many years. Most workers have con-
centrated on vision and audition to the virtual exclusion of the chemoreceptive senses. Hence, relatively
little is known about how the human sense of smell relates to infant behaviour. This paper will briefly outline
some of the early work in this area before attempting to delineate the current state of knowledge. Work
on human infant responses to food odours will be reported.
The Warwick Olfaction Research Group [Van Toller et al. (1989) Chem. Senses, 14, 1; Van Toller et al.
(1990) Psych. Res., 29, 429] has developed an existing psychophysiological method to examine cortical events,
as they occur, in response to various odours. This computerized BEAM technique is a development of tradi-

tional EEG measurement, which displays cortical activity in real-time as colour topographic maps. All our
subjects are tested using a Neuroscience Series III Brain Imager. A 28-electrode modification of the interna-
tional 10/20 EEG system is used, mounted in fabric headcaps. Systems have been developed for handling
the large data sets produced by this method of testing.
In a recent study, three-month-old infants were tested in a low-ambient-odour environment to a small range
of common baby-food odours. The testing chamber was perceptually impoverished and all subjects were
tested whilst seated on the mother's knee. Odours were remotely presented without cuing. Despite wide
individual variations, cortical maps tended to show a fairly coherent response in the low-frequency (1 —4 Hz)
delta waveband. This was interpreted as indicating an 'orienting' response to the odorants.
42. Suppression of responses to sweet stimuli by the gymnemic acids, ziziphins and
hodulcln
Linda M.Kennedy
Department of Biology, Clark University, Worcester, MA 01610, USA
The gymnemic acids, ziziphins and hodulcin (GA, Z and H) (from Gymnema sylvestre, Ziziphus jujuba,
and Hovenia dulcis leaves) selectively suppress human perception of sweet stimuli and fly behavioral and
receptor cell responses to sucrose. We have been studying the chemistry and physiological actions of these
"taste modifiers' in order to elucidate the affected sweet taste mechanisms.
GA, Z and H are triterpene saponin glycosides, amphipathic molecules with surfactant properties. They
produce biphask effects on receptor cell responses to sweet stimuli: firing is first suppressed and then becomes
increased and irregular. For Z, and probably also GA and H, the onset of the increased firing depends upon
a mechanism involved in firing, such as the opening of ion channels. In the case of the initial suppression,
concentration-effects (C-E) curves for the three modifiers are bell-shaped, with similar effective concentra-
tion ranges, slopes, and peaks. The critical micelle concentration for each modifier corresponds to the rising
phase of the C-E curve; thisrelationshipsuggests a role for physkochemical mechanisms. Also, the amphipathic
nature of the modifiers, and evidence of hemolysis by GA and Z, suggest that the modifiers penetrate into
receptor cell plasma membranes. Receptor cell and human perceptual data for H and GA show a concentration-
dependent pattern of mixed kinetics that is characteristic of an effect on a later step in a multistep process.
Moreover, the kinetics are similar for a variety of sweeteners.
Overall, modifier action on a single sweet receptor seems unlikely. We propose that the modifiers penetrate
into the receptor cell membrane and affect a membrane molecule involved in a multistep transduction pro-
cess which is either coupled to multiple sweet receptors or functions without a classical protein receptor
molecule. The modifier could act (i) directly on the transduction molecule or (ii) on membrane lipids adja-
cent to, and thus indirectly affect the transduction molecule. Physicochemical factors, e.g. hydrophobic and
dielectric effects, are presumed to drive the membrane penetration and interactions with membrane molecules.
43. Judging the appropriateness of a generalized procrustes analysis on data from

a sensory panel
Bonnie M.King, Paul Arents and Garmt Dijksterhuis1
Quest International, PO Box 2, 1400 CA Bussum and 'OP&P Sensory Research, PO Box 14167, 3508 SG
Utrecht, The Netherlands
In sensory research it is often desirable to represent, in a two-dimensional 'map', a set of samples so that
those having similar sensory qualities are grouped together and those having different qualities are spread
388
ECRO IX: Abstracts
apart. The procedure requires a form of multivariate data analysis because the n samples are usually evaluated
by a panel trained to recognize k specific sensory qualities (descriptors) and to measure the intensities of
these qualities, i.e. each panelist produces a data matrix (nxk) of intensity scores. The experimentalist wishes
to obtain from these matrices the best 'average relationship' among the samples without losing the qualitative
information that defines their similarities/differences.
Generalized Procrustes analysis (GPA) has been proposed as a means of matrix-matching which would
allow the averaging of 'maps' produced by individual panelists even if there were no prior agreement as
to the number or nature of descriptors. Several computer programs are available for carrying out GPA. While
most of these programs also provide an analysis of variance which enables one to judge the relative agree-
ment among panelists as to the consensus 'map', they offer no absolute criterion to warn the experimentalist
that he may have created a consensus sample grouping where the original data do not support (or even hint
at) such a trend. In fact, GPA on random data has been shown to create groups of 'samples' in a consensus
plot for which the residual variances (samples as well as panelists) were extremely low.

We propose a guideline for acceptance of GPA solutions based on the examination of variance partitioned
over each dimension in a full-dimensional solution. The guideline also makes use of results from cluster
analysis. The procedure is illustrated on data sets showing varying degrees of agreement among panelists.
44. The pyranose site in Drosophila is a mosaic

G.Wolff, A.Klarenberg and H.Wieczorek
Zoologisches Institut der UniversitOI, Luisenstrafie 14, D-8000 MOnchen 2, FRG
Previous electrophysiological experiments with the sugar-sensitive receptor cell of Drosophila melanogastcr
had led to the hypothesis that chemospecificity is the result of a mosaic of many different, independently
expressed receptor sites [Wieczorek and WolfT(1989)/. Comp. Physiol. A, 164, 825; Wolff and Wieczorek
(1989) Verh. Dtsch. Zool. Ges., 82, 285].
Among the various different receptor sites investigated in the fleshfly or the blowfly, the pyranose site
appears to be rather homogeneous. Investigating the pyranose site of Drosophila, we compared dose-response
curves of sixteen different Drosophila species for three structurally very similar sugars: sucrose, maltose
and D-glucose. The species were chosen with regard to their relationship (five different systematic groups)
and to their nutrition habit (fruit, sap, decaying vegetables and mushrooms). All sixteen species showed
a similar sensitivity to sucrose, but differences with regard to maltose and to D-glucose. The sensitivity spectra
were correlated to the kind of feeding source but not to the degree of relationship. Regarding the sensitivity
to maltose, it was the same or better than that to sucrose in flies living on fruits or feeding sap and four-fold
lower in those feeding on cactus rots. The spectrum for D-glucose ranged from an identical down to a 10-fold
lower sensitivity compared to that to sucrose. The biological relevance of the electrophysiological results
could be ascertained by behavioural assays [tests according to Tanimura et al. (1982) / . Comp. Physiol.,
147, 433].
Phloridzin, a competitive inhibitor of the glucose transporter, exerted different effects on the reception
of the three sugars. Concerning the reception of D-glucose, the sensitivity in the presence of 10 mM phlorid-
zin was depressed to nearly zero. In contast there was nearly no influence on the reception of maltose and
of sucrose. We observed this effect in Drosophila melanogaster, a species with a high sensitivity to all three
sugars, as well as in Drosophila mojavensis which has a sugar receptor with low sensitivity to maltose and
an even lower one to D-glucose.
Our results are in line with the hypothesis that the pyranose site is not a homogeneous population; sucrose,
maltose and D-glucose may bind to different receptor sites. We propose that the pattern of the different sen-
sitivities in different species may be the result of a species-specific expression of more than one receptor
site. [Supported by the DFG (Wi 698/2-2).]
45. A vacuolar-type H + -ATPase energizes electrogenic K + -transport in insects—

immunologicaJ evidence in sensory epithelia
UUa Klein, GCnter Loffelmann, Bernhard Zimmermann1 and Helmut Wieczorek
Zoologisches Institut der Universitat, Luiscnstrafie 14, D-8000 MOnchen 2 and 'institut fur Zoologie der
UniversiUU, Universitatsstrafie 31, D-8400 Regensburg, FRG
Electrogenic K+-transport in insects serves as energy source for generation of receptor current in sensilla
and for secretion of absorption of nutrients and ions in gastrointestinal epithelia like midgut, Malpighian
tubules and salivary glands. In insect sensilla, the apical membrane of the auxiliary cells is the putative site
of the K+-transport. So far, an ouabain-insensitive K+-ATPase activity could be demonstrated in membrane
389
ECRO IX: Abstracts
fractions of sensilla-rich labella of flies [Wieczorek (1982) J. Comp. Physiol. A, 148, 303]. In the midgut
of Manduca sexta, the goblet cell apical membrane is the site of active K+-transport. From this membrane,
a vacuolar-type H + -ATPase has been purified [Schweikl et at. (1989) J. Biol. Chem, 264, 11136]. The
H + -pump drives an H + /K + -antiport, resulting in the net active and electrogenic transport of K + [Wieczorek
el at. (1989) J. Biol. Chem., 264, 11143].
These findings put forward the question of whether a similar vacuolar-type H+-ATPase acts as primary
active ion pump in other K+-transporting insect epithelia, too. Therefore, SDS-treated crude extracts of anten-
nal sensory epithelium and Malpighian tubules of Manduca sexta were probed by immunobiochemical
investigations. Polyclonal antibodies raised against the purified midgut ATPase exhibited cross reactivity
in both epithelial extracts with protein bands corresponding to the five major subunits of the midgut ATPase.
For immunocytochemical studies at the light and electron microscopical level, monoclonal antibodies directed
against single subunits of the midgut ATPase were used. In the midgut, they specifically bound to the goblet
cell apical membrane, the site of active K+-transport. In the antennal sensory epithelium of Antheraea pernyi,

strong cross-reactivity was found in the apical membrane of the auxiliary cells. These results support our
suggestion that a vacuolar-type H+-ATPase is a basic constituent of insect K+-transporting membranes.
(Supported by the Deutsche Forschungsgemeinschaft, Wi 698/2 and SFB 4.)
46. Olfactory P300: lateralization of hedonics and discrimination performances

G.Kobal, M.Durand-Lagarde and E.Pauli
Instilul fir Pharmakologie und Toxikologie, UniversitOatsstrafie 22, 8520 Erlangen, FRC
P3 is the third positive-going wave of an olfactory-evoked potential occurring after a surprising, or low-
probability stimulus. The aim of the present study was to investigate whether this endogenous cortical response
is related to the emotion induced by the stimulus. Sixteen volunteers (eight male and eight female) participated
in the experiments which consisted of four different randomized sessions. The low-probability stimuli
(P = 0.05) called target were either H2S or vanillin. The standard stimuli (high-probability stimuli:
P = 0.85) consisted of a mixture of vanillin and H 2 S. Both nostrils were stimulated with all combinations
of standard and target stimuli. Stimulus interval was 8 s. EEG was recorded from seven positions (10/20
classification) referenced to linked earlobes. Subjects ranked vanillin as pleasant and H2S as unpleasant.
Pleasant stimuli were identified better when delivered to the right nostril. As already observed [Kobal el at.
(1989) Chem. Senses, 14, 5] pleasant stimuli also led to smaller amplitudes and shorter latencies in the early
components when delivered to the right nostril. On the contrary, unpleasant stimuli provoked smaller amplitudes
and shorter latencies when presented to the left nostril where they were also identified better. No correlation
between the hedonic estimates and the P300 latencies was displayed. However, in accordance with the literature,
the P300 component revealed significant greater amplitudes with unpleasant stimulation. It can then be assumed
that the recognition of an odorant seems to be related to the emotion it induces, and that this phenomenon
is determined by the specialization of the corresponding hemisphere.
47. Molecular cloning of pheromone-binding proteins in insect antennae

Jflrgen Krieger, Klaus Raming and Heinz Breer
Pheromone-binding proteins (PBP), characteristic proteins of the sensillum lymph, are supposed to enhance
the capture rate of pheromone molecules by solubilizing these odor molecules, enhancing their partitioning
into the sensillum fluid surrounding the dendrites and transferring the pheromones to receptor proteins located
in the dendrite membrane. Pheromone binding proteins have been isolated and identified as 15 kd polypep-
tides. For a more precise molecular characterization, which may provide some clues for elucidating the
structure-function relationship, pheromone-binding proteins from two species of the giant silk moth (Antheraea
polyphemus, Antheraea pernyi) have been cloned and sequenced using recombinam DNA techniques. It was
found, that PBPs are expressed as precursor proteins with a signal peptide of 21 amino acids. The deduced
structure of the mature protein consists of 142 amino acid residues giving a calculated molecular mass of
15 783 daltons. Analysis of secondary structural features as well as the hydrophobity profile revealed that
the polypeptide can be arranged to display a hydrophilic surface as well as hydrophobic pockets. Thus, the
primary structure allows the protein to meet its functional requirements, i.e. it is soluble in aqueous media
and binds hydrophobic molecules. Northern blot analysis indicated that the PBP is selectively expressed in
male antennae and is encoded by a 1.4 kb mRNA. In order to find out whether a proposed diversity of PBPs
may be accomplished by alternative splicing of the primary transcription products, the structural organization
390
ECRO IX: Abstracts
of the gene encoding the PBP was explored. Sequence analysis of several genomic clones revealed that a
PBP gene is composed of several exons separated by large introns.
48. Olfactory recognition of androgens in scent marks of male saddle back tamarins
(Saguinus fuscicollis, Callitrichidae)—a preliminary study
I.Kflderling, G.Epple and A.Belcher
German Primate Center, Gottingen, FRG and Monell Chemical Senses Center, Philadelphia, PA, USA
Information transfer by scent is one of the major means of social and sexual communication in the saddle
back tamarin, a South American primate. Both sexes of this species possess a large complex of specialized
skin glands in the suprapubic and circumgenital region, which serves as a scenting organ. The scent marks

are deposited on items in the environment by distinct ritualized behaviors. They are composed mainly of
secretions from the scent gland and urine. Behavioral studies on the communicatory content of these scent
marks have demonstrated that they convey a number of messages characteristic of the donor individual.
Recent studies have shown that die organic solvent fraction of scent marks from intact males is discriminated
from that from females and castrated males [Epple et al., Olfaction and Taste, 10, in press]. This indicates
that the olfactory information about maleness is testosterone-dependent. Since a variety of steroids including
androgens may have semiochemical potential and because testosterone occurs naturally in the scent marks
of male Saguinus fuscicollis, we tested whether the presence of the hormone in the marks contributes to
the recognition of maleness. Pooled scent marks from castrated males were spiked with testosterone (60 ng/
sample) and offered in a two-sample choice test together with non-spiked aliquots from the same pool.
Interestingly, the test individuals (males and females) scent marked over the testosterone-spiked samples
significantly more frequently than over the control preparation (n = 26, P < 0.008, Wilcoxon), while they
made no clear difference in sniffing at the samples.
This preliminary study indicates that the sensory quality of the scent marks is altered when they contain
testosterone. In further studies we examine the extent to which the presence of the hormone adds to the biological
activity of fractionated scent material.
49. Biological determination of the solubility properties of the frog olfactory mucus
H.J.Kuiper1, P.Marfaing, E.P.Koster1 and P.Laffort
Laboratoire de Physiologie de la Chimiortception (URA 1190), CNRS, 91190 Gif-sur-Yvette, France and
'Psychological Laboratory, University of Ultrechl, Sorbonnelaan 16, 3584 CA Utrecht, Netherlands
In a previous study of electroantennogram stimulus —response curves in the honey-bee, it has been shown
that the three parameters of the Hill equation are strongly interdependent, provided odorant concentrations
are expressed in terms of parital saturated vapor pressures [Patte et al. (1989) J. Insect Physiol., 9, 667].
That the saturated vapor pressure plays such an important role in the EOG of the honey-bee and not in the
EOG of vertebrates may be due to the way in which the molecules reach the dendrites.
In order to verify this hypothesis and to specify the solubility properties of the frog olfactory mucus, EOG
responses in the frog Rana esculata were recorded for six different concentrations of 11 odorants in Utrecht
and of 10 odorants in Gif-sur-Yvette, with two overlapping odorants. In addition, a 'biologically determined
filter effect' (BDFE) was defined by grouping the three parameters of the Hill equation. As an illustration,
the BDFE values in the honey-bee for 59 odorants, calculated from the [Patte et al. (1989)] study, is cor-
related with the saturated vapor pressure with r «* 0.93.
In the present study on the frog, an acceptable agreement between the BDFE values obtained in Utrecht
and in Gif for the two overlapping odorants (1-butanol and benzene) has been observed: the difference in
value equals 16% of the range for the 19 odorants studied.
A multiple regression analysis was applied to the frog BDFE values versus serveral physicochemical pro-
perties including the saturated vapor pressure, the air-water partition coefficient, and the 'solubility parameters'
determined by Patte et al., Patte et al. (1982) Anal. Chem., 54, 2239]. The best fitting is obtained by com-
bining the five solubility parameters in a regression equation in which those involved in London forces (alpha
and epsilon) are prominent (r = 0.89; F = 11). Similar results are obtained with the two subsets of data
taken separately or together.
391
ECRO DC: Abstracts
50. The human sense of smell has a limited capacity for identifying odors in mixtures
D.G.Laing, B.A.Livermore and G.W.Francis
CS1RO Food Research Laboratory and Macquarie University, Australia
Since most odors in our environment, whether at home, work or at play, are complex and often consist of
dozens—even hundreds—of odorants, a question of considerable significance concerns the number of odorants
humans are capable of perceiving in mixtures. The present study addresses this question using untrained
and trained panels of laypersons, and an expert panel of perfumers and flavorists. A computer controlled
air dilution olfactometer was used to deliver stimuli consisting of between one and five odorants, from a
pool of seven odorants, to subjects. The stimuli were common, dissimilar odorants of equal moderate inten-
sity and each was a single chemical. The results were unexpected and dramatic and showed that (i) regardless
of training and experience, less than 5% of judgements of mixtures containing more than three odorants
were correct and (ii) regardless of whether judgements were correct or incorrect, subjects rarely selected
more than three odorants, indicating that perceived complexity did not increase with mixtures containing

more than three odorants. Overall, the results indicate that the human sense of smell is very limited in its
capacity to discriminate and identify components of mixtures and this capacity is limited to three or four
odorants. It is proposed that identification of the constituents of mixtures is limited by the convergence of
multiple neural inputs at higher centres, e.g. olfactory cortex, but is aided to some extent by temporal separation
of input from each odorant.
51. Threshold perception of banana odour by a frugivorous bat: a case of

hyperadditivity?
Matthias Laska1-2, Robyn Hudson2 and Hans Distel2
'Zoology Institute of the University at Bonn and 2Institute of Medical Psychology, University ofMQnchen,
Goethestrasse 31, D-8O00, Munchen, FRG
Naturally occurring odours generally consist of highly complex mixtures of volatiles. With regard to odour
intensity, components may theoretically exhibit either complete addition, hyperaddition, or hypoaddition.
While the latter effect is well established, complete addition or hyperaddition are less common and have
even been called into question. However, such effects have generally been investigated using simple mix-
tures at above-threshold concentrations, with relatively little attention having been given to threshold
concentrations.
Here we present evidence from threshold measurements in a fruit-eating bat (n = 4) that the sensitivity
to a complex, biologically relevant food odour may represent an example of hyperadditivity, with sensitivity
to the whole mixture substantially exceeding the sum of the component thresholds.
Table: Concentrations of major components of banana odour (molecules/cm3) estimated to be present in
threshold concentrations of the whole odour (A) and when tested individually at threshold (B)
A B
Ethanol 1.8 X 10* 7.2 X 10"

Ethyl acetate 3.7 X 106 3.6 X 10 12
Iso-butanol 4.8 X 107 1.2 X 10 12
n-Pentanol 1.0 X 107 4.3 X 10"
Iso-butyl acetate 1.3 X 107 8.7 X 1010"
n-Butylacetate 9.1 X 106 6.5 X 10 10
Ethyl butyrate 9.5 X 106 2.7 X 10 10
Penryl acetates 1.2 X 107 not tested
Iso-pentyl acetate 2.5 X 107 7.4 X 10 10
n-Pentyl acetate 3.5 X 106 2.8 X 10 10
Hexyl acetate 8.4 X 106 not tested
E major components 1.4 X 108
£ minor components 2.1 X 107
Whole odour 1.6 X 10*
We would therefore like to suggest that hyperadditive effects can occur, and under natural conditions, would
serve to enhance the detection of biologically relevant odours. (Supported by the Deutsche Forschungsge-
meinschaft, Po 121/13.)
392
ECRO DC: Abstracts
52. Spatial organization of the bilateral bulbar projection of the primary olfactory'
neurons in the frog
J.Lcvcteau, I.Andriason and P.Maclcod1
Departement de Neurophysiologie Comparie, IDN, University Pierre et Marie Curie, 75230 Paris and
'Laboratoire de Neurvbiologie Sensorielle, EPHE, ENSIA, 91315 Massy, France
Bilateral topographical projections of both intact olfactory receptor mucosae onto the olfactory bulbs were
investigated by means of both electrophysiological and neuroanatomical methods.
Each of the olfactory nerve fascicles subserving either the dorsal mucosa or the ventral one was electrically
stimulated and averaged evoked potentials were recorded from the olfactory bulbs. Isopotential maps were
drawn in the three orthogonal planes at critical post stimulus major peaks in such a way that regional organization
could be worked out. In order to trace the pathways, horseradish peroxidase was iontophoretically injected
in the dorsal nerve endings and processed according to the TMB method.

The results indicated: (i) the neuroreceptor axons projected bilaterally to the medial olfactory bulb, and
(ii) the existence of a transposition of the medial-lateral and the dorso-ventral axes of the receptor sheet into
a medially-laterally and a dorsally-ventrally oriented respective axes into the bulb.
We think that the present data prove that some fibers of the olfactory nerve do have contralateral projec-
tions in the glomerular layer. The question of what can be the physiological and behavioral implication of
anatomical evidence remains fully open.
53. The relationship between the nerve fibers in the lateral olfactory tract and the
olfactory bulb in cod
Birgitte Lindeman
Department of Biology, University of Oslo, PO Box 1051, Blindem, 0316 Oslo 3, Norway
In cod specific behaviour patterns can be evoked by electrical stimulation of olfactory tract bundlets [Dewing
and Selset (1980) Science, 207, 559]. This feature permits a description of the anatomical and functional
properties of the nervous substrate that participates in the execution of each behaviour pattern. In the present
investigation the relationship between the nerve fibers in the lateral olfactory tract (LOT) and the olfactory
bulb in cod was studied. A fluorescent carbocyano dye, Dil or DiO, was applied at well defined parts of
the LOT in paraformaldehyde-fixed preparations. These dyes will colour the nerve fibers in question from
the application site to their termination in the bulb by simple diffusion in the plasma cell membrane [Godement
et al (1987) Development, 101, 697].
The results show that the fibers from the LOT extend into limited areas of the dorsal, dorso-lateral and
dorso-medial parts of the bulb. This finding points to the importance of spatial information processing in
the olfactory system.
The first workers to describe ruffed cells in the olfactory bulb of fish [Kosaka and Hama (1979) Brain
Res., 163, 151; Kosaka and Hama (1979)7. Comp. Neurol., 186, 301; Kosaka (1980) J. Comp. Neurol.,
193, 119] used methods which did not permit the determination of the axonal projection of these cells. In
my preparations, cells with morphological characteristics similar to those of ruffed cells, also took on colour.
This implies that these cells have axons in the LOT. The somas of the ruffed cells identified were situated
in the internal cell layer of the central part of the bulb.
54. Taste thresholds, detection probabilities and stimulated area

Miriam R.I.Linschoten and Jan H.A.Kroeze
Laboratory of Psychology, Utrecht University, The Netherlands
In two experiments, the effect of areal size and location of stimulation on the NaCl-threshold was measured.
In a temporal 2-AFC procedure five concentrations around the individual threshold were presented 32 times
each under two conditions. A filter paper stimulation method was used in order to control the location and
the areas compared. In the first experiment 12 subjects took part in two conditions: a circular area of
63.62 mm2 (A) and half a circle with an area B = 0.5A. The halves were presented in four orientations
on the same location as the full circle, a lateral and a medial orientation parallel to the midline of the tongue
and a superior and an inferior orientation perpendicular to this line. The location of stimulation was on the
anterior dorsal surface of one tongue side.
The results showed that all subjects had a lower threshold for the full circle than for the averaged half
393
ECRO IX: Abstracts
circle. The mean ratio between the thresholds was 1.46. Thus, summation in taste is partial and the relation-
ship between threshold intensity and area obeys Piper's law.
Comparing the threshold function for the two conditions leads to tho conclusion that doubling the area
increases the detection probability for each of the five concentrations according to chance. The observed
values of the threshold function for the full circle did not deviate from the predicted values on the basis
of the independent probability summation models [Pirenne (1943) Nature, 152,698; Erikson (1966)7. Exp.
Psychol., 72, 26].
There was no difference in sensitivity between the medial and lateral half circles. However, the sensitivity
to the inferior half circle was significantly greater than to the superior half circle.
In the second experiment 13 subjects took part and three conditions were compared, a full circle (A), half
a circle on either of two tongue sides (B) and half a circle on both tongue sides simultaneously (C). For
the half circles only the lateral and the medial orientations were used. The results obtained on the separate
tongue sides matched those of the first experiment and no difference between the tongue sides was found.
The thresholds obtained with the half circles on both tongue halves (C) were equal to the ones obtained with

the full circles (A), and both were lower than the ones obtained with the single half circles (B).
In the unilingual conditions the threshold functions for the full circle (A) could be predicted on the basis
of the functions for the single half circles (B), using the independent probability summation theorem. However,
in the biolingual condition a difference was found between the lateral and the medial orientations. Location
seems to be important when two tongue sides are stimulated simultaneously. In the bilateral orientation sum-
mation is better than chance for the three lowest concentrations, while in the bimedial orientation detection
can be predicted according to chance. These results seem to suggest that probability summation fits most
of the data; central enhancement may contribute if low stimulus concentrations are presented near the lateral
edges of the tongue.
55. Recognition threshold concentrations of migrants from food packages: styrene

and ethylbenzene
J.P.H. Linssen, A.Legger-Huysman and J.P.Roozen
Agricultural University, Department of Food Science, PO Box 8129, 6700 EV Wageningen, The Netherlands
Polystyrene, which is used as a food packaging material, contains residual styrene monomer and ethylbenzene
in detectable amounts. These components are able to migrate into the packed food in amounts depending
on the properties of both packaging material and packed substance.
Ethylbenzene was added to water, and styrene was added to water, to an oil-in-water emulsion and to
a water-in-oil emulsion (both 30% oil) in different concentrations. The taste recognition threshold concen-
trations were determined by an aselect panel of 53 persons. Fifty per cent of the assessors (z-score = 0)
recognized styrene in water at a concentration of 22 p.p.b. and ethylbenzene in water at a concentration
of 16 p.p.b. In both emulsions styrene was recognized by 50% of the assessors at much higher concentra-
tions: 2.8 p.p.m. in the oil-in-water emulsion and 1.7 p.p.m. in the water-in-oil emulsion. These values
were about 100 times higher than the values of styrene in water.
The continuous phase of the emulsion type does not seem to be important for the recognition threshold
concentration of styrene. Paired student (-test shows no significant difference for these thresholds (P > 0.2).
Linear regression of the combined z-scores versus log concentration styrene results in a value of 2.1 p.p.m.
at z-score = 0.
56. Influence of androstenon, androstenol, menstrual cycle and oral contracep-

tives on the attractivity ratings of female probands
Regina E.Maiworm and Werner U.Langthaler
It was the aim of the study to evaluate the influence of the putative pheromones 5a-androst-16en-3a-ol and
5a-androst-16en-3a-on, menstrual cycle and oral contraceptives on the rating of photographs of men by female
probands. The evaluation was done with defined adjective scales expressing the attractivity of the demonstrated
standardized photographs. The observation of the double-blind study was performed by female experimen-
tors who were not aware of the aim of the survey. Seventy-one female students of psychology took part
in the program. Testing probands were randomized according to testing with androstenon, androstenol and
solvent (ethanol without pheromone). Test substances were offered at a concentration of 1 mg/ml. Ten minutes
after the first application on the upper lip, five photographs had to be rated immediately after repetitive
394
ECRO IX: Abstracts
application of the same test substances. The rating was done according to an earlier method [Kirk-Smith
(1978) Res. Com. Psycho. Psych. Behav.]. A scale of attractivity from 0 (unattractive) to 100 (attractive)
was added. Three-factorial analysis (MANOVA) showed a significant influence of the menstrual cycle on
the rating of masculinity (P = 0.01) and a significant influence of the substances on the rating of intelligence
(P = 0.05). There was a significant interaction of the menstrua] cycle phase (preovulatory/postovulatory)
and of oral contraception for the ratings of intelligence, 'interesting', 'sexy' (P = 0.01); 'attractivity', 'emo-
tional', 'active' (P = 0.05) and the attraction value. The ratings 'warm' and 'intelligent' were significantly
(P = 0.05) influenced by the test substances and the menstrual cycle. The mid-cyclic phase (days 10—19),
the substances and oral contraception showed a significant interaction for the rating 'warmness'. The mid-
cyclic phase and oral contraception showed a significant (P = 0.05) interaction on the evaluation of 'intelligent',
'interesting', 'emotional' and 'black'. The evaluation of emotional (P = 0.01) and black (P = 0.05) was
influenced by the test substances. Factorial analysis discriminated four factors. The first parameter group
consisted of the items attractive, intelligent, interesting, sexy and the attraction value. This factor was signi-

ficantly influenced (P = 0.01) by the phase of the menstrual cycle and oral contraception. The second factor
(f2) is subsummanzing the items 'emotional', 'warm' and 'sensitive'. The ratings of f2 are significantly
(P = 0.005) influenced by the phase of the menstrua] cycle (9,9,9) and oral contraceptives. In summary,
the experiment revealed that the attractivity rating of presented photographs of men by female probands was
significantly influenced by the phase of the menstrual cycle and the application of oral contraceptives. The
presentation of putative pheromones was not a significant influence on the evaluated items with the exception
of the rating of 'intelligence', 'emotional' and 'black'.
57. On the synaptology of glomerular projection neurons in the antennal lobe of

Periplaneta americana
Dagmar Malun
Institute of Zoology, University of Regensburg, UniversitOustrqfie 31 D-8400 Regensburg, FRG
The uniglomerular olfactory projection neurons (PNs) in the antennal lobe of the American cockroach repre-
sent the major link between the first and the second central stage of the insect central olfactory pathway.
Each of them has dense dendritic arborizations in a single glomerulus within the lobe. The axons project
via the tractus olfactorio-globularis to the calyces of the mushroom bodies and the lobus lateralis protocerebralis
of the protocerebrum respectively. For a morphological study of details of their synaptology inside their
glomeruli, PNs were stained by intracellular injection of (i) Lucifer Yellow or (ii) Biocytin. Both labels
were prepared for an investigation of the PN processes in the electron microscope; Lucifer Yellow-filled
neurons were incubated with a primary antiserum against the fluorescent dye and visualized by the indirect
peroxidase-antiperoxidase technique (the Lucifer Yellow-antiserum was kindly provided by Dr P.H.Taghert).
Biocytin was labelled with avidinylated peroxidase. Series of thin sections of portions of severfal PNs were
reconstructed to reveal the distribution of their synapses.
Inputs and outputs are concentrated in different regions of PN processes. Output synapses are located on
the large-diameter fibres entering the glomerulus. Thinner and more ramified branches are predominantly
presynaptic, input synapses are rare. These branches represent the transition zone to the terminals which
bear input synapses only. The many input synapses in these sections may reflect the high degree of con-
vergence of receptor input on PNs and may result in the high sensitivity of the PNs. PNs with especially
rich arborizations appear to respond to odor stimulation of the antenna more sensitively than do PNs with
less well developed arborizations. The high number of output synapses indicates that PNs have a more com-
plex role than merely relaying olfactory signals to the protocerebrum and that they are involved in glomerular
integration. Unlike the vertebrate bulb, no reciprocal synapses have yet been found. (Supported by the Deutsche
Forschungsgemeinschaft SFB4/B3.)
58. Sensory thresholds evaluated from dose-response curves

Anna B.Marin, J.Barnard1, Richard B.Darlington2 and Terry E.Acree3
Department of Food Science and Technology, 'Department of Computer Services and ^Department
Psychology, Cornell University, New York, USA
3
To whom correspondence should be addressed at Department of Food Science & Technology, Cornell
University, NYSAES, Geneva, NY 14456, USA
Three methods of threshold estimation are compared for the detection of 1,8-cineole added to Concord grape
juice presented in a semi-ascending paired difference test. Two methods consider dose—response behavior
395
ECRO IX: Abstracts
to be a discontinuous function with the threshold at the point of discontinuity. Method A tests the significance
of obtaining a non-zero response, and method B determines the transition point of a two-phase linear model
fitted to the data. An alternative method fits a logistic model to the data and assumes that dose-response
behavior is a continuous function. Method C approximates a threshold at the point of maximum curvature,
Cmar Method A yields thresholds which are inversely related to the error in the data and systematically
decrease with increasing sample size. Methods B and C yield thresholds which are independent of sample
size, are a property of the dose - response data, and are similar in value.
59. Plasticity in the olfactory system: general concept or specific problem?

Claudine Masson
LNC1 INRA-CNftS (URA 1190) F-91440 Bures sur Yvette, France
All animals have to adapt to changing environmental conditions. In most species—ranging from lower
invertebrates to mammals—this results in lasting changes in behaviour as a consequence of a defined interac-

tion with their environment. This suggests that the underlaying nervous systems responsible for such widespread
behavioural plasticity might share common neural rules and mechanisms. The search for such bask and common
rules represents one of the most fascinating fields in neurobiology.
A large body of experimental evidence indicates that the interaction with the outside world regulates, or
is even required, for the full development of the connectivity and behavioural performance of the adult brain.
Consequently, epigenetic mechanisms associated with an interaction with the outside world appear to be
partly responsible for the establishment of the neuronal network during embryonic and postnatal develop-
ment leading to the adult pattern of connectivity and behaviour.
Adaptative changes occur at all levels of neuronal complexity. Because the flexible recruitment of many
neural circuits into assemblies might represent the most relevant unit in terms of behavioural structure rather
than individual neurones or local circuits, to have some chances of identifying the rules of neural plasticity
it is necessary to select a number of adequate biological model systems. Such models need to be suitable
for complementary approaches at different levels of organization, taking into account assemblies of neurones
and individual nerve cells in a close and permanent relationship with the whole organism and its behaviour.
Moreover, it is necessary to consider that each level may possess its own rules which in turn depend on
those of its constituent elements, and so to have a careful and critical attitude with respect to interpreting
correlations between data collected at different levels or from different model systems in terms of causality.
Based on these general assumptions the talk will be oriented to show that, compared to other sensory systems,
the olfactory system provides particularly interesting possibilities for studying developing neural systems,
especially with respect to neurobehavioural plasticity during development and the neurobiology of learning
and memory. For example, olfactory information is processed through a well defined, simple pathway, the
anatomical and physiological details of which have been precisely analysed. Moreover, olfactory processing
is very similar in phylogenetically distant species (e.g. insects/mammals). Similarly, olfactory preference
learning in early life has been related to clear plasticity in the developing brain areas that could underlie
life-long individual differences in behaviour, and this in different animal species. Thus, one is justified in
using 'simple' model preparations allowing the useful comparison between truly comparable organizational
levels but of rather different complexity. In addition, as the olfactory receptor cells are but a single neurone
removed from the olfactory cortex, the olfactory system, unlike other sensory systems, has direct access
to higher brain structures involved in cognitive processes such as learning and memory.
As most of the statements are based on results from different and complementary approaches, including
anatomy, physiology and behaviour, it is proposed to concentrate the talks of the two discussants on the
validity of the relationship between structure and function on which most of our knowledge of the neurobiology
of olfactory function has been based. For the debate it is suggested to consider as a fruitful model the olfac-
tory preference learning in young mammals (rat, rabbit) because such learning might share mechanisms both
with other forms of developmental plasticity and with adult olfactory learning (e.g. associative processes).
60. Microvilli of rat olfactory supporting cells contain ample amiloride-sensitive

sodium channels. A rapid-freeze, freeze-substitution Lowicryl K11M embedding
immunocytochemical study
Bert Ph. M.Menco and Dale J.Benos1
Department of Neurobiology and Physiology, O. T. Hogan Hall, Northwestern University, Evanston, IL 60208
and 'Department of Physiology and Biophysics, University of Alabama, Birmingham, AL 35294, USA
In this study electron-microscopic immunocytochemistry was used to localize amiloride-sensitive sodium
channels in olfactory epithelia. Rat olfactory epithelial samples were rapidly frozen, freeze-substituted with
3%
ECRO IX: Abstracts
acetone, embedded at low temperatures with Lowicryl Kl 1M and labeled on the sections with polyclonal
antibodies raised against the amiloride-sensitive sodium channel from bovine renal papilla [Sorscher et al.
(1988) Am. J. Physiol., 255, C835; Tousson et al. (1989) J. Cell Sci., 93, 349] and with protein G con-
jugated to colloidal gold. Apart from the aforementioned use of acetone, substitution was carried out in the
complete absence of chemical fixation, i.e. neither aldehydes nor OsO4 were used. This procedure resulted
in localization concurrent with a good ultrastructural preservation. The epithelial sodium channel blocker
amiloride [Kleyman and Cragoe (1988) / . Membrane Biol., 105, 1] inhibits partly and reversibly electrical
responses to odorants [Frings and Lindemann (1988) /. Membrane Biol., 105, 233; Persaud et al. (1987)
Biochim. Biophys. Acta, 902, 65]. However, the antibody bound to supporting cell microvilli instead of
olfactory receptor cells. Hence, physiological inhibition might be caused by action of amiloride on olfactory
supporting instead of on receptor cells. Negative controls were carried out with normal rabbit serum. The
cytochemistry supplied for the first time evidence about a possible role for supporting cell microvilli in ol faction,
namely to take part in electrolyte and water balance regulation of the mucus around supporting cell apices

and more important, around apical parts of the receptor cells and thus, odorant receptor and signal transduc-
tion sites. Structures of olfactory supporting cell apices share the presence of these channels, of binding
sites for the lectin Dolichos biflorus agglutinin and of typical intercalated dumbbell-shaped freeze-fracture
particles with cells of kidney collecting tubules [Brown et al. (1985) Am. J. Physiol., 248, C348; Brown
(1989) Am. J. Physiol., 25, Fl; Brown et al. (1989) Am. J. Physiol., 25, F366; Menco (1980) Cell Tissue
Res., 211, 5, and in press; Weyer et al. (1988) Am. J. Physiol., 254, C450] suggesting similarities in func-
tion at various levels. (Supported by grants from NSF (BNS-809839) and the Erna and Victor Hasselblad
Foundation.)
61. Differential binding of lectins to olfactory and respiratory cilia and microvilli
in freeze-substituted or deep-etched rat samples
Bert Ph.M.Menco
Department of Neurobiology and Physiology, O. T. Hogan Hall, Northwestern University, Evanston, IL 60208,
USA
Biochemical and electrophysiological studies on olfactory epithelia with lectins have suggested that a major
part of the ciliary cell-surface proteins involved in odorant reception and signal transduction are glycopro-
teins. Therefore, binding of colloidal gold-conjugated lectins was studied in cilia and microvilli of rat olfac-
tory with cilia and microvilli of nasal respiratory epithelial surfaces as comparison. This was done in sections
of rapidly-frozen, freeze-substituted specimens embedded in Lowicryl Kl 1M or, for wheat germ agglutinin
(WGA) alone, in deep-etched replicas. Olfactory dendritic endings and cilia labeled with WGA and, faintly,
with soybean agglutinin (SBA); olfactory supporting-cell microvilli bound only Dolichos biflorus agglutinin
(DBA). Microvilli of an infrequent cell bound peanut agglutinin (PNA), SBA, and WGA. Relative to olfactory
cilia these microvilli labeled more strongly with the last two lectins. The cell resembles a presumptive sensory
one [Rowley et al. (1989) Brain Res., 502, 387]. Cilia of nasal respiratory cells bound WGA and, somewhat
more weakly, PNA; microvilli of respiratory cells bound all four lectins. In respiratory ciliated cells cilia
and microvilli of the same cell type showed a quite different pattern of labeling. Visualization of specific
labeling improved after pre-incubation of sections with neuraminidase, except for DBA where lectin binding
was abolished. PNA labeling was seen only after neuraminidase pre-incubation. The densities of membrane
surface particles labeled with WGA corresponded with those of fracture plane particles in a quantitative freeze-
fracture, deep-etch analysis. Hence, most of the lectin-bound particles could reflect transmembrane proteins
in olfactory dendritic endings and cilia and in respiratory cilia. So, lectins were useful markers for cilia,
including olfactory ones, and microvilli of nasal epithelial cells: cilia and microvilli in olfactory and nasal
respiratory epithelial surfaces could be distinguished by membrane-associated glycoconjugates using electron-
microscopic cytochemistry. Thus the new cytochemical data extended freeze-fracture and freeze-etch data
on the special morphology of cilia and microvilli of olfactory and nasal respiratory epithelia. (Supported
by grants from NSF (BNS-809839) and the Erna and Victor Hasselblad Foundation.)
62. Lectin-binding studies on the main olfactory epithelium and vomeronasal

neuroepithelium of the rat and golden hamster
Andres S.Mendoza and Wolfgang Kflhnel
Institute far Anatomie, Medizinische Universitat Lubeck, Ratzeburger Atte 160, D-2400 Labeck 1, FRG
It is known that air-borne compounds are more readily registered by the main olfactory epithelium (MOE),
whereas liquid-bound compounds are most readily registered by the vomeronasal neuroepithelium (VNO-NE).
397
ECRO IX: Abstracts
Lectins, which are sugar-binding proteins or glycoproteins that agglutinate cells or precipitate glycocon-
jugates having saccharides of appropriate complementarity, were used to detect some of the sugar residues
(lectin-binding sites) present in the apical border of the MOE and VNO-NE of the rat and golden hamster
in order to determine whether chemosensory epithelia with supposedly different odorant affinities also vary
with respect to their lectin binding properties. The following lectins were used: wheat germ agglutinin (WGA),
horseshoe crab agglutinin (LPA), gorse agglutinin (UEA I), peanut agglutinin (PNA), soybean agglutinin
(SBA) and horse gram agglutinin (DBA). Lectin-binding procedure was performed on paraffin sections of
the MOE and VNO-NE using the pcroxidase-antiperoxidase method. Comparisons of the lectin-binding
properties of the MOE surface with that of the VNO-NE as estimated by the intensity of staining are depicted
for both species in the following table.
Rat Golden hamster

MOE VNO-NE MOE VNO-NE
WGA +++
LPA (-)
UEA I +++
PNA + +
SBA +++
DBA (±)
( - ) = no reaction; ( ± ) = weak reaction; (+ + +) •= strong reaction.
Our results demonstrate that in both species differences exist between the lectin-binding properties of the
MOE and VNO-NE. The LPA reaction of the VNO-NE differs from species to species, as does the DBA
reaction of the MOE.
63. Extreme olfactory sensitivity of precocious male Atlantic salmon parr (Salmo
salar L.) to the steroid testosterone
A.Moore and A.P.Scott
Ministry of Agriculture, Fisheries and Food, Directorate of Fisheries Research, Pakefield Road, Lowestofi,
Suffolk NR33 OHT, UK
Electrical responses (electro-olfactogram: EOG) were recorded from the olfactory epithelium of precocious
male Atlantic salmon (Salmo salar L.) parr exposed to gonada) steroids (thought to function as pheromones),
several L-amino acids and a bile acid. Responses of non-precocious parr and female parr were also com-
pared. Testosterone was a potent odorant in male precocious parr with a threshold of 10" l 4 M. The respon-
siveness of groups of fish to testosterone was limited however to a short time period of approximately 4
weeks during October. The olfactory epithelium of non-precocious and female parr did not respond to
testosterone at any time. No responses were recorded from the precocious male parr to the pheromone 17a,
20jS-dihydroxy-4-pregnen-3-one, (17.20P), which has been demonstrated to be a potent odorant in the goldfish,
or to any of the other steroids. The olfactory thresholds or precocious male parr to L-amino acids and taurocholic
acid were similar to other teleost species: 10~ 8 M for L-cerine and 1 0 " ' M for L-cysteine and taurocholic
acid.
64. Innervation of taste buds and adjacent epithelia of the goldfish as studied in
methylene blue supra vitally stained and paraffin-embedded tissues
T.Muller and K.Reutter
Anatomical Institute, University of Tubingen, FRG
The aim of the study was to adapt the classical supravital methylene blue staining of nerve fibres [Ehrlich
(1886) Dtsch. med. Wschr., 12, 49; Dogiel (1897) Arch. mikr. Anat., 49, 769] to paraffin-embedded tissues
in order to investigate the fine innervation patterns of taste buds (TB) and the adjacent epithelia. The investigation
was done on juvenile goldfish (Carassius auratus), 80—100 mm in length. Into the lip's regions of each
398
ECRO IX: Abstracts
MS 222-anaesthetized animal 1 ml of 0.25% methylene blue (in 0.9% NaCl) was subcutaneously injected.
Then the heads were removed and exposed to the air in a moist chamber for 25 min. The first fixation was
done in 10% ammonium heptamolybdate (in 0.9% NaCl) for 30 min at room temperature. After a short
wash the second fixation step took place for 2 h at 4°C in a mixture of 2% paraformaldehyde and 2.5%
glutaraldehyde in 0.1 M phosphate buffer (pH 7.4) which contained 1.5% phosphomolybdic acid (final pH
5.5). After washing in aqua dem. and dehydration in tert-butanol (melting point: 25°C) for 2 days, the heads
were transferred through methyl benzoate and Rotihistol into Paraplast. Sections (15 /im) were cut,
deparaffiriized in xylene and cover-slipped with DePeX [MQller (1989) Stain Tectwol., 64, 93]. The investigation
shows that in the lip's epidermis and in the TBs of the goldfish distinct variations of innervation patterns
occur. In taste buds the TB nerve fibres ascending to the TB's nerve fibre plexus vary; the nerve fibre plexus
itself forms delicate varicosities (synapses) to the distinctly visible and differently arranged basal processes
of sensory cells and to the (unstained) basal cells, too. Outside the TBs, within the adjacent epithelia, there
exist occasionally solitary perigemmal nerve fibres which may arise along the lateral side of a TB up to

the apical cell layer; some intraepithelial nerve fibres derive from the TB's nerve and may vary too; numerous
intraepithelial nerve fibres may be directly in connection to the corium's nerve fibre bundles. In future, the
fact of an intense variability of innervation patterns in TBs and their adjacent epithelia should also be con-
sidered in studies done by the use of very specific methods, as usual in histochemistry and electron microscopy.
In this respect, our variation of the old methylene blue staining method might be a useful tool.
65. Sensory and semantic factors in recognition memory for odors and graphic
stimuli: elderly vs young persons
Claire Murphy, William S.Cain1, Magdalena M.Gilmore and R.Blair Skinner
San Diego Stale University and 'John B. Pierce Laboratory and Yale University, USA
Four experiments explored episodic recognition memory for both graphic stimuli (faces of American presidents
and vice-presidents, engineering symbols, and free-forms) and everyday odors in young and elderly people.
The elderly consistently performed about equal to the young on the graphic stimuli, but worse than the young
on the odors. Although apparent for retention intervals as short as 26 s, the poorer performance of the elderly
on odors became truly marked only after an hour or more. Somewhere between 1 h and two weeks, odor
memory performance of the elderly fell to chance, whereas their graphic memory performance remained
well above chance. Although the young showed progressive forgetting over a six-month period, their perfor-
mance remained above chance for both graphic stimuli and odors. When measured (experiments 1 and 2),
the absolute threshold of the elderly fell about an order of magnitude below that of the young. Sensitivity
may have governed part of the poorer memory performance of the elderly, but seemed not to govern it entirely.
Knowledge that the subjects brought into the experiment, e.g. degree of familiarity with and ability to name
the odors and faces, played a positive role in recognition memory. That role, and the ability of verbal distraction
to disrupt odor memory, implied that odor recognition depends somewhat, and perhaps heavily, on semantic
processing. Even the negative influence of olfactory sensitivity on odor memory might have been mediated
via impaired opportunities for semantic processing of odors that seemed desaturated, or pastel, to the elderly.
(Supported by NIH grants AG 04085 and AG 04287.)
66. Sucrose stimulation of adenylate cyclase activity in membranes of taste papillae

from pigs and cattle
Michael Nairn, Tal Ronen, Benjamin J.Striem, Martha Levinson and Uri Zehavi
Department of Biochemistry and Human Nutrition, Faculty ofAgriculture, The Hebrew University ofJerusalem,
Rehovot 76100, Israel
The possible involvement of cAMP in ceilular transduction of taste evoked by sucrose in the pig was investigated
in the plasma membrane of isolated pig's circumvallate and fungiform taste papillae. For comparison, some
experiments with membranes from taste papillae of cattle were also conducted. Typical adenylate cyclase
responses to guanine nucleotides and fluoride-activating GTP-binding protein were found in the circumvallate
plasma membranes. In the circumvallate membranes, sucrose stimulated adenylate cyclase activity in a dose-
dependent manner while it inhibited this activity in membranes from either fungiform papillae or from those
of non-sensory epithelium. Similar results were obtained with membranes derived from taste papillae of cattle.
GTP was required in the circumvallate membranes for the stimulatory effect of sucrose. This effect of sucrose
was also dependent on the concentration of membranes used in the adenylate cyclase assay. At high con-
399
ECRO IX: Abstracts
centrations of membranes sucrose had a stimulatory effect, while at low concentrations an inhibition was
observed. The present study provides the first biochemical data on cellular transduction of taste in pig's
taste papillae and supports recent studies suggesting a role for cAMP in sweet taste transduction induced
by sucrose. (Supported by the Mutual Fund of the Hebrew University and by grant 1-100-181 from GIF
(the German-Israeli Foundation.)
67. Sensory astringency and vegetable tanning—the same or different?

M.Naish, M.N.Clifford1 and G.G.Birch
Department of Food Science and Technology, University of Reading, Whiteknights, PO Box 226, Reading
RG6 2AP and 'Department of Biochemistry, University of Surrey, Guildford GU2 5XH, UK

The sensation of astringency is generally recognized to be associated with polyphenolic substances such as
tannins. The word tannin was introduced by Seguin in 1796 to describe extractable matter in certain plant
tissues able to convert hide to leather, and much of the early work on the phenomenon of astringency and
polyphenols developed out of tanning studies. In this context tannins have been defined by Haslam in 1981
as 'water-soluble phenolic compounds having relatively molecular mass between 500 and 3000, and besides
having the usual phenolic reactions they have special properties such as the ability to precipitate alkaloids,
gelatin and other proteins'.
The distinctive structural feature of tannins has been described by Haslam as 'the accumulation within
a molecule of moderate size of a substantial number of phenolic hydroxyl groups, many of which are associated
with an o-dihydroxy or o-trihydroxy orientation within a phenyl ring'.
It has been demonstrated that tannins have considerable affinity for proline-rich proteins (PRP) such as
gelatin, and those that occur in the saliva. By analogy with leather tanning it has been assumed that sensory
astringency is caused by the precipitation of these proteins in the buccal cavity with concomitant loss of
their lubricating effect and production of an unpalatable dry sensation.
We have carried out time/intensity astringency studies on more-or-lcss astringent substances and shown
that no more than one 1,2-dihydroxyphenyl residue is required for a molecule to generate astringency and
that such a sensation is clearly different from bitterness by virtue of its persistence relative to its intensity.
We are seeking a better understanding of the astringent sensation and definitions that are more appropriate
to the sensation rather than to tanning. Evidence will be presented to support our proposals, to provoke discussion
thereof and invite amendments thereto.
68. Receptor cell distribution in the olfactory epithelium of cod

Torild Nilsen
In cod specific behaviour patterns can be evoked by electrical stimulation of olfactory tract bundlets [Deving
and Selset (1980) Science, 207, 559]. This feature permits a description of the anatomical and functional
properties of the substrate that participates in the execution of each behaviour pattern. In the present in-
vestigation the distribution of receptor cells in the sensory epithelium was studied.
The cod's olfactory epithelium is arranged in lamellar foldings perpendicular to a central raphe. There
arc approximately 10-12 lamellae in one rosette. Since the epithelium is covered with kinocilia, the recep-
tor cells are not visible with SEM. The receptor cells show a remarkable uptake of fluorescent dye [Holl
(1981) Stain Technol., 56, 67], and this feature was used to visualize the receptor cells. The olfactory organ
was exposed to a 0.06% solution of Procion Yellow for 5 min. The olfactory rosette was dissected free and
fixed in 2% paraformaldehyde for 2 h. The anterior and posterior surfaces of the lamella were separated.
The one-cell-layer-thick part of the lamella was mounted flat and examined under a fluorescence microscope
and photographed.
In the three anterior lamellae the receptor cells are distributed homogeneously over the lamella surface.
In the rest of the lamellae the receptor cell density was particularly high in special areas. These areas con-
verge against the region between the raphe and the 'thumb'. On a lamella of 24 mm2 there are approximately
40 000 receptor cells.
400
ECRO DC: Abstracts
69. A study on the saliva viscosity in human taste

I.Ohsawa
Department of Oral Diagnosis, Nihon University School of Dentistry, Japan
In human taste, I paid attention to saliva which has the purpose of carrying taste substances to taste buds
or taste cells, and considered the relation between human taste and saliva. Then I considered some ions in
human saliva by semi-quantitative stycks, three anions, SO4, NC^ 2 " and CrO 4 2 ~, were measured. This meant
that the buffer action of saliva pH was affected significantly by protein as I reported in 10th ISOT.
I then paid attention to the sulfate ion in the three anions, because I hypothesized that there was a connec-
tion between gulfate ion and saliva viscosity. Saliva viscosity was said to originate the connective force among
polypeptides of mucin which was reliefed and continuously took place in the molecule reciprocal action of
radical end which was highly ionized. There seemed some relation between sulfate radical end which caused
saliva viscosity, and sulfate ion which I measured. Saliva viscosity was indispendable mucin which affected
total protein volume in saliva and saliva buffer action. Furthermore saliva viscosity seemed to take part in

the diffusion of taste substances in oral cavity and the adsorption to taste cells. I hypothesized that there
seemed some relation between human taste and saliva viscosity.
Subjects were 30 male students, 22 to 25 years old, who attended Nihon University School of Dentistry,
had no systemic diseases, and took no medicines. The method of gastatory test was paper filter disc which
I reported in 10th ISOT. I used paraffin wax in oral cavity for saliva volume measurement for 15 min and
BTB paper (Toyo-roshi Co.) to determine the pH of saliva, and for the measurement of saliva viscosity I
used the cone plate type rotational viscometer. Biorheolizer (Tokyo-keiki Co.).
Results of saliva viscosity were in the region of 6.29 x 10~ 3 — 684 x 10~3 Ns/m2 in the increasing
stress and 9.90 x 10" 3 - 451 x 10~3 NS/m2 in the decreasing stress. The relation among the pH, volume
and viscosity of saliva seemed to be the higher viscosity of saliva the better the result of the gastatory test
ever in the case of greater saliva and lower pH. Even with a lower pH of saliva, in the case of lower viscosity,
the results of the gastatory test were poor and with less saliva, in the case of lower viscosity, the results
were worse. So there seemed to be the strong relation between saliva viscosity and human taste.
70. Dichotic odorous stimulation with eugenol and dJpyridyl: influences upon
hedonic estimates, intensity estimates and chemosensory evoked potentials
E.Pauli, Th.Hummel and G.Kobal
Institut fir Pharmakologie und Toxikologie, UniversiUUsstrafie 22, DS520 Erlangen, FRG
The aim of the study was to investigate the influence of a dichotic odorous stimulation with eugenol and
dipyridyl upon hedonic and intensity estimates as well as chemosomatosensory evoked potentials, i.e. odorants
that not only activate the olfactory but also the trigeminal system.
Forty right-handed subjects participated in the experiments (20 male, 20 female; mean age 25.8 years).
After one training session, they all participated in one experimental session, in which eugenol (43 ng/ml)
and dipyridyl (0.5 ng/ml) were each presented 16 times into the right and the left nostril (200 ms duration,
ISI - 4 0 s). After application of each stimulus the subject indicated the stimulus location and estimated the
hedonic quality and the intensity of the stimulus (visual analogue scale). Subjects performed a single video
game during intervals (tracking performance). EEG was recorded from eight positions of the international
10/20 system (sites: Fp2, F3, Fz, F4, C3, Cz, C4 and Pz versus Al +A2; band width 0 . 2 - 7 0 Hz). After
checking for artifacts the stimulus-linked EEG records were averaged, and the amplitudes and latencies of
the chemosensory evoked potentials (CSEP) were measured. After chronological grouping the records, the
spontaneous EEG was submitted to frequency analysis. Additionally, intensity thresholds were determined,
and the hedonic character of the odorants was once more evaluated (visual analogue scale, item scale).
Eugenol was perceived to be pleasant, whereas subjects pronounced dipyridyl to be neither pleasant nor
unpleasant. Since for both odorants the stimulated nostrils were easily identified, it was evident that they
also excited endings of the trigeminal nerve. The left nostril turned out to be more sensitive than the right
one (intensity estimates, thresholds, localizability of stimuli). Smokers were not as sensitive as were non-
smokers (higher thresholds, lower intensity thresholds, not easily able to locate the stimuli).
Changes in the tracking performance during the experiments (improvement in the beginning, deterioration
in the end) were accompanied by consistent changes in the power densities of the a2- and /S-range.
CSEPs were significantly different in dependence upon the used odorants. Eugenol and dipyridyl evoked
maximum amplitudes at Pz. Latencies revealed a distinct laterality in dependence upon the stimulated side.
401
ECRO EX: Abstracts
There were significant correlations between the CSEP parameters and the intensity estimates and intensity
thresholds.
Changes in the CSEPs in dependence upon the hedonic nature of the odorant and the site of application
were not evident. For both odorants this might be due to the fact that they clearly excited the trigeminal
nerve. Additionally, dipyridyl was esimated as hedonically neutral, so no changes in the olfactory evoked
potential could be expected.
71. Recognition and recall of odours

Judith Perkins
Department of Psychology, University of Ulster at Jordanstown, Newtownabbey, Co. Antrim, UK
The classic 'Proustian' view that odour memory is impervious to interference, unique and independent can

be seriously questioned. This study examined the effects of suppression of visual and verbal encoding pro-
cesses on olfactory memory performance. Subjects were allocated to one of four suppression conditions (no
suppression, visual suppression, verbal suppression, and visual-plus-verbal suppression), presented with target
odours, and then tested for either free recall or recognition of these.
Although both types of performance showed a significant overall effect for level of suppression, results
were only partially consistent with predictions drawn from dual coding theory.
72. A multi-element gas and odour detector based on conducting organic polymers
Krishna C.Persaud, Paolo Pelosi1 and Peter A.Payne
Department of Instrumentation and Analytical Science, UMIST, Manchester, UK and 'istituto di Industrie
Agrarie, University of Pisa, Italy
Devices which are capable of objective qualitative and quantitative odour measurements are still not available.
At ECRO VI, Lyons, 1984, we presented for the first time new sensor materials based on conducting polymers
which were capable of odour detection. In the intervening years we have further developed these materials
to produce an array of sensors which contain elements which have broad overlapping specificities of response
similar in some characteristics to biological olfactory receptor arrays. We have developed the electronics
and software necessary to produce a small portable device capable of measuring and discriminating between
many polar volatiles such as aldehydes, alcohols, ketones, amines, and mercaptans. The device is capable
of real time measurements within seconds, is operated at room temperature, requires low power and is not
poisoned by volatiles which cause irreversible changes to tin oxide sensors. A working device will be
demonstrated. Applications include olfactometry, quality control in the food and beverage industries and
environmental monitoring.
73. Partial amino acid sequence of odorant binding proteins from rabbit, pig and
human olfactory tissues
Anna Maria Pisanelli, Jeffrey N.Kcen1 and Massimo Dal Monte
Istituto di Industrie Agrarie, University of Pisa, Italy and 'Department of Biochemistry, University of Leeds,
UK
Soluble odorant binding proteins have been identified in many animal species. Their function is as yet unknown,
but their presence suggests a specialized role in the olfactory system. In this study we have examined the
partial sequence homologies from odorant binding proteins (OBP) purified from rabbit, pig and human olfactory
tissues and have compared them with those of the cow and rat OBPs and aphrodisin from the hamster.
OBPs were purified by affinity chromatography and ion exchange chromatography [methods: Pelosi et
al. (1990), in press]. Peptide fragments were generated by digestion with 1 % V8 protease in 67 mM phosphate
buffer, pH 7.4, 37°C for 2 h. From rabbit and human OBPs two peptides were obtained, while pig OBP
yielded four peptides. The amino acid sequences of these peptides were determined using a Milligen 6600
sequenator.
The results show substantial homologies between the rabbit and cow OBPs with large similarities also
with the pig OBP. The pig protein show large homology with aprhodisin from hamster. The human protein
is currently being sequences!
402
ECRO K : Abstracts
74. Study of some factors affecting intensity/time characteristics of sweetness

response
M.O.Portmann, S.Serghat and M.Mathlouthi
Laborasoire de Chimie Physique Industricllc, Faculty des Sciences, University de Reims Champagne-Ardenne,
B.P. 347, 51062 Reims Ctdcx, France
Intensity/time plots of sweetness produced by sucrose and D-fructose in water al concentrations ranging from
2.3 to 9.2% (w/v) were recorded for solutions at 20 and 35°C. The intensity and persistence power func-
tions were applied to the results obtained with a potentiometer connected to a chart recorder similar to the
SMURF device [Birch el al. (1981) Chem. Senses, 6, 45].
Increasing the temperature of the assessed samples leads to a slight decrease of both the intensity and
persistence of the sweet taste of sucrose and D-fructose. This result is interpreted by reference to solution
properties (apparent specific volume and intrinsic viscosity) of the sugars and their effect on water structure.

The second factor affecting the sweetness response to be studied was viscosity. Intensity/time plots were
recorded for solutions containing 5% sucrose or 4.2% fructose brought to apparent viscosity values of
5, 15, 25 and 35 mPa/s by addition of maltodextrins. It was found that sweetness intensity decreases as
viscosity increases while the persistence remains almost constant.
The results suggest that the sweet taste chemoreception probably proceeds in two steps: the first depends
on the accessibility of the sweet molecule to the receptor site, and the second, which involves opening of
the ion channels, is sensitive to water mobility around the site and the sweetener.
75. A comparison of the pheromone receptor system of four Mamestra moths

(Lepid.: Noctuidae)
Michel Renou and Philippe Lucas
INRA Laboratoire des Midiateurs chimiques Brouessy F78114 Magny les Hameaux, France
A comparison of the organization and sensitivity of the olfactory hairs receptive to pheromone components
has been done in four Namestra species by scanning electron microscopy and single hair recording.
The antennae are about 10 mm long and unbranched. The olfactory hairs are restrained to the ventral sur-
face of the flagellar segments. Two fields of trichoid sensilla can be distinguished. Lateral hairs (LHs) are
longer (up to 110 /«n), hooked, regularly arranged in four or five parallel rows. They are male-specific.
A medial field of shorter (50 /an) trichoid sensilla (MHs) is present. The MHs are more numerous than the LHs.
All the LHs and most of the MHs appeared to house one or two pheromone sensitive receptor neurones.
Stable differences of action potential amplitudes facilitated the measure of individual activity of receptor
neurones in two species. All the LHs housed two pheromone sensitive receptor neurones. A receptor neurone
tuned to Zl l-16:Ac was found in the LHs of the Mamestra species. Zl l-16:Ac is the main component of
the female secretion of three of them. The second receptor neurone in the LHs of brassicae and suasa was
found to be tuned to Z9-14:Ac, a compound not produced by the conspecific females but that inhibits the
sexual behaviour in males. The second LH receptor cell of oleracea and of thalassina was sensitive to a
common minor component of the female pheromone, Zl 1-16:OH. A single type of MHs, with only a recep-
tor cell sensitive to the Zl l-16:Ald was found in thatassina. M.brassicae MHs have not been investigated
yet. MHs of the same sensitivity as LHs were found in oleracea and in suasa together with MHs sensitive
to the minor component Zl 1-16:Aid in suasa, and to two further compounds with unknown function. Z9-14:
Ac and Zl l-14:Ac in oleracea.
Species specificity in the organization of the peripheral olfactory system shows that not only the presence
of receptor neurones specifically tuned to a compound but also their relative number and arrangement is
important to the coding of the pheromone blend.
76. Possible mediatory role for inositol-l,4,5-triphosphate in olfactory transduction

Diego Restrepo, Taufiqul Hugue, D.Lynn Kalinoski, Takashi Kumazawa, Takenori Miyamoto and
John H.Teeter
Monell Chemical Senses Center, Philadelphia PA, USA
An odorant-regulated cyclic AMP cascade has been implicated in olfactory transduction. Recent studies in
the channel catfish (Ictalurus punctalus) indicate that exposure of isolated olfactory cilia to olfactory stimuli
(amino acids) leads to G-protein-mediated increases in intracilliary cAMP. This increase, which occurs at
relatively high stimulus concentrations, could lead to depolarization by opening of cAMP-gated channels
located on the plasma membrane of catfish olfactory cilia [Bruch and Teeter, Chem. Senses, in press). On
403
ECRO IX: Abstracts
the other hand, L-amino acids trigger a rapid G-protein-mediated increase in phosphoinositide (PI) turnover
resulting in the formation of mtracilliary inositol-l,4,5-triphosphate (IP3) which is further metabolized
resulting in rapid formation of other inositol polyphosphates. In addition, L-amino acids elicit a transient
elevation in intracellular calcium in a group of olfactory neurons, and an elevation in [Ca,] can also be
triggered by exposure of olfactory cells to the G-protein agonist AF*~. Both the stimulus and AF 4 " induced
increases in [Ca,] are mediated by an influx of external Ca 2+ which is not inhibited by addition of the L-
type Ca 2 + channel antagonist nimodipine. To characterize this Ca 2 + influx pathway we have measured zero-
current membrane potential in cells under whole cell patch clamp. We find that addition of 8 ^M intracellular
1,4,5-1P3 (but not 1,3,4-IP3) elicits a large transient depolarization which is inhibited by 10 /iM ruthenium
red. In addition, we find that, when isolated olfactory cilia vesicles are fused into phospholipid bilayers at
the tip of a patch pipette, IP3 (1 - 2 0 iM) directly gates a cation channel with a slope conductance in 55 mM
Ba2+ of 79 ± 5 pS. This channel is selective for Ba 2+ (and Ca 2+ ) over K + and N-methyl-D-glucamine
(minimum permeability ratios for Ba 2+ over K + and NMDG + are 7 and 54 respectively) and is inhibited

by ruthenium red at 10 jiM. By studying radioligand binding to isolated cilia membranes we have also identified
a binding site for 3H-IP3 which may correspond with the IP3-gated channel. Cation influx through this channel
could cause receptor cell depolarization, suggesting that G-protein-linked PI turnover may play a mediatory
role in olfactory transduction through opening of an LP3-gated cation channel. (Supported by NIH grant
DC-00566-02, BRSG S07RR05825 and a grant from the Veterans Administration to Dr J.G.Brand.)
77. infrastructure of taste buds in the Australian lungfish, Neoceraiodus forsteri

(Dipnoi)
K.Reutter
Anatomical Institute, University of Tubingen, FRG
For the first time, the taste buds (TBs) of the Australian lungfish are described by using TEM techniques.
The study is fecussed to the TBs which occur in the fish's mouth cavity. In dependence to their actual loca-
tion, shape and size of TBs varies, but at all, their structural organization is uniform. TBs are situated within
the statified squamous epithelium and are ovoid in shape if cut longitudinally. They are 8 0 - 100 yxn long
and 70—80 pm wide. A TB rests on the top of a dermal papilla and depresses it in a U-like manner. The
apex of the TB is formed by a pit-like structure, the TB-porus. A TB consists of about 6 0 - 8 0 cells which
belong to four distinct cell types: elongated light and dark sensory cells form the sensory epithelium in the
stricter sense and reach with their distal endings the TB porus; basal cells form the actual basis of the TB;
granular cells are located also near the basis of a TB. A real nerve fibre plexus does not exist but nerve
fibres are intermingled with the basal processes of light, dark and granulated cells and also with basal cells
and synapse with them. Within the TB-porus, the apical structures of light and dark sensory cells converge
and interweave themselves: light sensory cells possess only one large villus, dark cells either several small
and often divided ones or bundles or stereocilia-like microvilli. The cytoplasm of light sensory cells is rich
in longitudinally arranged filaments, in microtubuli and has a well developed smooth endoplasmic reticulum.
Contrarily, the cytoplasm of dark sensory cells is nch in Golgi fields and contains lots of vesicular struc-
tures. These are regularly filled with electron-dense material. Strands of filaments are numerous. The granulated
cells possess a relatively clear cytoplasm rich in organelles. Their granular aspect is caused by dense-cored
— 160 nm vesicles. These cells do not reach the TB-porus. Some of their lobar processes are attached to
the basal lamina. The disc-shaped basal cells are arranged transversally to the longitudinal axis of the TB.
Their cytoplasm is rich in organelles, too. They often do not lie consequently on the underlying basal lamina:
occasionally, some nerve fibres lie between a basal cell and the basal lamina. In my opinion, in the TBs
of the Australian lungfish the light sensory cells are the sensory cells in a stricter sense; the dark sensory
cells may possess a secretory function, too; the granulated cells are possibly paracrine cells and the basal
cells may function as interneurons or are even mechanoreceptive, comparable to Merkel cells [see Reutter
(1987) Ann. NY. Acad. Sd., 510, 570]. These data are of special interest in view of the vertebrate TB phytogeny.
(Supported by DFG 225/10-1.)
78. Ciliary spike responses to neurotransmitters recorded from olfactory mucosa

slices
J.F.Rosin, D.Trotier and P.MacLeod
Neurobiologie Sensorielle, EPHE, I Av. des Olympiades, F-91305 Massy, France
In order to study the sensitivity of the frog olfactory neuroreceptors to several neurotransmirters, transmucosal
slices (thickness 200—300 /an) were prepared. They were secured into a 70 /d perfusion chamber (D.Trotier,
ISOT X, 1990).
404
ECRO IX: Abstracts
Under microscopic control, a patch-clamp pipette (tip diameter 0.8-1.2 /un), filled with Ringer solution
(7 — 16 MQ), was gently moved towards the epithelium surface. By applying a weak negative pressure (20-60
cm of water), a few (1 -15) long immotile cilia converged and gradually entered the pipette. As half of
the cilia length was inserted, a spike activity appeared, which could be recorded up to 1 h without appreciable
modifications.
These 10 ms spikes corresponded to part of an outward ciliary current returning to the action potential
generating membrane. Their amplitude was usually 2 0 - 3 0 pA with a signal-to-noise ratio up to 15 when
filtered with a 100—300 Hz band-pass. Several amplitude classes were often observed; these were stable
enough to allow discrimination of the corresponding unicellular activities.
The usual basal firing pattern was an irregular succession of 10-20 s of low activity and of 1 - 3 s of
higher activity periods, similar to in vivo conditions. Some cells had a basal quasi-periodic bursting activity.
Chemicals were applied close to the recording site by pressure ejection from a pipette.
Nine agonists corresponding to the main classes of neurotransmitter receptor sites have been tested at

10—500 pM (about 40 units). They induced responses similar to odorant and isobutylmethylxanthirte
applications (10-500 /iM).
Most of the cells responded to 2 —4 s stimulation by an increase of their firing rate (maximum 30 spike/s).
For high concentrations, spike amplitude decreased, then the firing activity paused for 10-15 s. Sometimes,
multiple damped bursts occurred. By contrast, related competitive antagonists seemed to be ineffective when
used as stimuli. When present in the perfusion medium, these antagonists were apparently unable to modify
the sensitivity of the cells to the puffer application of the corresponding agonists.
79. Morphometric analysis of male antenna! lobes in the moth Manduca sexta
J.P.Rospars and J.G.Hildebrand1
Laboratoire de Biomitrie, INRA, F78026 Versailles, France and 'ARL, Division of Neurobiology, Univer-
sity of Tucson, AZ, USA
As part of our continuing comparative studies of the functional organization of antennal lobes (ALs), especially
of their interspecific variations and sexually dimorphic components [e.g. Rospars and Chambille (1989).
In R.N.Singh and N.J.Strausfeld (eds) Neurobiology of Sensory Systems. Plenum Press, New York], we
have begun an investigation of the glomerular neuropil of the sphinx moth Manduca sexta. ALs of male
individuals have been analyzed morphometrically from serial sections stained by Bodian's silver proteinate
method. Profiles of glomeruli, cell groups, and fiber tracts on each section have been traced, aligned and
digitized. We have further developed microcomputer programs to study these quantitative data. The position
and volume of AL components have been computed, visualized in 3D space, and analyzed statistically. We
have observed the following (i) The number of glomeruli per AL is almost constant (64 ± 1, macroglomerular
complex not included), (ii) Glomeruli are present in different ALs as individually identifiable and homologous
units. Identification criteria permit their classification in these categories. Glomeruli in the first class have
been recognized as homologous only on 3D computer reconstructions, mainly from their relative positions.
Glomeruli in the second and third classes can be recognized on sections from their characteristic morphological
features or proximity to landmarks such as fiber tracts and cell groups, or from their position relative to
one another, (iii) Only three exceptions to this identification rule have been found. Each anomalous glomerulus
is present in only one AL. (iv) Some glomeruli can be distinguished by their exceptional sizes. They include
the very large macroglomerular complex (more than 200 /an in diameter), which is made of at least two
bulky subunits, glomerulus No 20 (about 100 /jm in diameter), and glomerulus No 64, which receives afferent
neurons from the labial palp pit organ. The other 'ordinary' glomeruli are in the range 50—80 ym in diameter,
(v) The volumes and positions of homologous glomeruli are conservative as shown by statistical analyses
and various methods of automated pairing. The variability measured in Manduca is similar to that observed
previously in the cockroach Blaberus and the moth Mamestra.
80. Confidence interval for olfactory thresholds

Jacques Rouault and Patrice Girard
Laboratoire de Physiologie de la Chimioriception (URA 1190), CNRS 91198 Gif-sur-Yvette Cedex, France
The aim of the French AF NOR X43-101, the American ASTM E679, and the German VDI 3881 standards
is to estimate olfactory thresholds and confidence intervals for odorants [standardized values are presented
iaDevosetal. (1990) Standardized Human Olfactory Thresholds, IRL Press, Oxford]. These standards call
for a probit analysis based upon an Arcsine transformation of observed frequencies, assuming that the result
follows a normal probability law. There are several other resolutions for the confidence interval for a pro-
405
ECRO IX: Abstracts
portion, some ones being much better, and/or well suited to small samples [Rouault (1988) Mimoire de
these. Analyse ecologique et exp£rimentale de la competition entre Drosophila metanogaster et D.simulant.
Interaction avec le parasite Leptopilina boulardi. DeVeloppement et application de techniques statistiques
propres au traitement de trfcs petits echantillons, 397 pp. ISBN 2-907103-00-8].
The proposed method, which is compatible with the different experimental designs, consists in four steps:
(i) for a given value of the first kind risk, the observed proportions are converted into confidence intervals
by an appropriate method; (ii) we choose a 2-parameter mathematical function (straight line, logistic, nor-
mal distribution function, . . .); (iii) then we arbitrarily vary the two parameters and we retain the values
such as the function intercepts all the observed confidence intervals; (iv) finally, interception at the level
0.5 of the functions corresponding to retained values of parameters provides the confidence interval for threshold
dilution. A Pascal listing is available.

81. Does the first odor reaching the mucosa suppress or mask the second one?
C.Rouby, M.Vigouroux and A.Holley
Physiologie Neurosensorielle, Universiti Claude Bernard, CNRS URA 180 69622 Villeurbannc Cedex, France
This study was designed to test the hypothesis that (i) the minimal stimulus duration required for the percep-
tion of odor quality is different across odorants, and (ii) in a binary mixture, the odor which is perceived
with the shorter minimal duration will suppress the quality and/or intensity of the other. For this purpose
we built an olfactory stimulator allowing a very precise temporal control of the odorant stimulus. A previous
study in 1986 attested that a very short stimulus duration (50 ms) could be detected and recognized. However,
temporal integration processes are rather long in olfaction and allow odor interaction with receptors after
stimulus cessation. Therefore, we decided to investigate minimal stimulus duration by a masking paradigm:
the target odor is presented briefly (200 ms) and followed by a second, different odor, the mask (600 ms).
The delay between target and mask is varied in six different steps of 100 ms: the mask starts in advance,
or synchronous with the target, or 100, 200, 300 and 400 ms after.
The intensities of target and mask were evaluated, and the quality,of the mixture recorded: target alone,
mask alone, or a mixture. In the first part of the study, cineole was the target, hexenol the mask; the paradigm
was reversed in the second part.
Target odor and masking odor were sent either in the same nare (physical mixture) or in left and right
nare (dichorhinic mixture).
The results show that the first odor reaching the mucosa seldom dominates the second one (frequency:
10%) even when the delay between target and mask is 400 ms. On the contrary, backward masking is very
frequent (35%). Thus the first odor reaching the mucosa does not prevent the perception of the second one
and this does not allow us to measure a precise critical duration.
However, perception of such stimuli cannot be predicted by mixture interaction models: perception of
both odors occurs in only 55% of the stimulations, attesting a large amount of complete masking, which
also occurs when intensities of components are equal.
Dichorhinic mixtures give significantly more complete masking than physical mixtures. Hexenol is a better
mask than cineole, which acts more as an intensity suppressor.
The data suggest that interactions between physical and dichorhinic mixtures rely on different mechanisms,
and that quality masking and intensity suppression mechanisms could also differ. (This research was sup-
ported by a grant from Roure (No 820339.)
82. The formation of ternary inclusion complexes of gelatinized potato starch with
(—)limonene and menthone—a binding model with synergism
M.A.Rutschmann and J.Solms
Department of Food Science, ETH, 8092 Zurich, Switzerland
Starch inclusion complexes with hydrophobic ligands having a polar group in addition to a dominant non-
polar portion of the molecular (e.g. ketones, alcohols) have been described in several publications. In these
complexes starch displays individual populations of binding sites, which can be characterized by the affinity
spectrum method. Much less is known about the complex formation of starch with totally non-polar hydrophobic
compounds, like (-)limonene. With hydrophobic bonding being the only stabilizing force, non-polar
hydrophobic compounds cannot co-stabilize their complexes by dipolar interactions and display only limited
affinities. In the present study the formation of starch inclusion complexes of (-)limonene in the presence
406
ECRO IX: Abstracts
of menthone was investigated. Menthone has a molecular structure with polar as well as non-polar portions
and is able to co-stabilize (—)limonene inclusion by forming mixed complexes. The corresponding binding
parameters will be presented and discussed. The ternary system of starch, menthone and (-)limonene is
a possible model for specific binding of flavor compounds with synergistic effects.
83. Measurement of pheromone concentrations in the field with EAGs

A.E.Sauer, G.Karg, U.T.Koch and J.J.de Kramer
FB Biologie, Universiiat Kaiserslautcrn, FRG
It has been concluded from analyses by Murlis and Jones in 1981 that pheromones are propagated in the
field in the form of discrete 'odour puffs'. Receptive insects would thus encounter highly fluctuating pheromone

concentrations. However, these concepts have been derived from the dispersal behaviour of ions in an abiotic
environment and it is possible that pheromones may behave differently under field conditions due to the
complexity of their interactions with real biotopes. We have developed a robust and portable apparatus for
the measurement of pheromone concentrations under field conditions. It uses the antenna itself as a sensitive
and specific pheromone detector. Shock-proof contact with the electrodes is maintained by fixing the antenna
in a specially-shaped plexiglass holder mounted within a glass tube. Continuous airflow through the tube
is generated by a suction pump, and the incoming air can be purified by passage through a charcoal filter.
This allows the instrument to be zeroed and calibrated by the application of pheromone pulses of known
concentration. Removal of the filter allows the flow of non-purified air over the antenna which responds
by generating an EAG as a measure of the ambient pheromone concentration.
84. Ca2+-gated and cAMP-modulated cation conductance on Xenopus olfactory

cilia
D.Schild and J.Bishofberger
Physiologisches Institut der UniversitOt D-34 Gottingen, FRG
Over the last years, the voltage-gated currents in amphibia olfactory receptor neurones have been analyzed
using the patch clamp technique. However, almost no electrophysiological evidence concerning a satisfying
biophysical characterization of second messenger gated currents has been reported. On the basis of Ussing-
chamber measurement of currents across the whole mucosa we concluded that not only cAMP but also kinase
C, IP 3 and Ca 2 + were involved in the transduction process. Here we show the involvement of Ca 2+ in the
whole-cell mode of the patch clamp technique. The preparation and technical details are given in a previous
publication. In the voltage-clamp with all voltage-gated currents except the Ca 2 + current blocked (Cs + ,
TEA"1", TTX), the experimental outcome depended on whether the cells had retained their cilia or not. In
cells with cilia a conductance g^ activated in parallel (i.e. at the same potentials) with the Ca 2+ -inward cur-
rent. An increase in intracellular Ca 2+ activated the ciliar conductance gc. The corresponding current was
carried by various cations as shown by substitution experiments, g,. was modulated by the intracellular cAMP
concentration and blocked by amiloride. The existence of a Ca2+-activated cation conductance on olfactory
cilia leaves little space for speculation concerning its biological role. The most probable interpretation seems
to be a receptor-mediated IPyCa2"1" pathway.
85. Expression of the von Ebner's gland protein during ontogenetic development
of the rat
Hartwig Schmale, Kai Kock and Michael BlSker
InstituxfUr Zellbiochemie und klinische Neurobiologie, UnivtrsiUU Hamburg, D-2000 Hamburg 20, FRG
The secretory von Ebner's gland (VEG1) protein is highly expressed in rat von Ebner's glands which pro-
duce saliva that fills the furrows of vallate and foliate papillae. Molecular cloning and sequence comparison
have shown that the VE01 protein belongs to a protein superfamily of hydrophobic molecule transporters
and enzymes binding lipophilic substrates such as the recently characterized prostaglandin D synthetase.
It has been postulated that the VEG1 protein could be necessary in pre-receptor events, particularly in binding
of lipophilic sapid molecules [Schmale et al. (1990) Nature, 343 366]. We have now identified cDNA clones
for a VEG2 protein that is about 95% identical in cDNA and protein sequence with VEG1. Northern blot
407
ECRO DC: Abstract*
and in situ hybridization analysis using specific oligonucleotide probes indicates that the level of VEG2 mRNA
is at least ten-fold lower compared to that of VEG1 mRNA.
In order to determine the onset of VEG protein expression during ontogenesis, we performed im-
munocytochemistry with specific antibodies directed against the VEG protein as well as in situ hybridization
on von Ebner's gland tissue sections of fetal and postnatal rats. VEG protein expression begins at postnatal
day 2 in small alveoli branching off from von Ebner's gland ducts and increases with further growth and
terminal differentiation of the gland. Thus, VEG protein expression starts shortly before the first mature
taste buds appear in the gustatory epithelium. This correlation emphasizes the functional relationship of von
Ebner's glands and taste buds which are both derived from an imagination of tongue epithelium during
embryogenesis.
86. FMRFamide-Hke immunoreactivity in the olfactory brain of the spiny lobster,

Panulirus argus
M.Schmidt and B.W.Ache
Whitney Laboratory and Departments of Zoology and Neuroscience, University of Florida, St Augustine,
FL 32086, USA
In our search for potential neuromodulators of the olfactory brain we studied the distribution of FMRFamide-like
immunoreactivity in the deutocerebrum of the Caribbean spiny lobster, Panulirus argus, a crustacean whose
peripheral olfactory system is well characterized. A polyclonal antiserum raised against FMRFamide (Dr
E.Weber, Oregon Health Science University) and visualized by the PAP method was used to localize im-
munoreactivity in 100 /un thick vibratome sections of desheathed brains which had been fixed in 4% parafor-
maldehyde. FMRFamide-like immunoreactivity was found in all five neuropils of the deutocerebrum: the
olfactory (OL), the parolfactory (PL), and the accessory lobes (AL), and the lateral (LAN) and medial antennular
neuropiles (MAN). In the LAN and MAN many fine and densely packed afferent fibres originating from
the antennular nerve and some terminals of interneurons are immunoreactive. Immunoreactive terminals of
intemeurons are present in all glomeruli of the OL and the AL, and in the periphery of the PL. Of the three
layers of the columnar OL glomeruli, the subcap stains most intensely, whereas the base and the cap are
stained more lightly. In the AL the small, spherical glomeruli of the central layer are stained more intensely
than are the larger glomeruli of the lateral and medial layer. The immunoreactive terminals in the OL, AL
and PL belong to interneurons with immunoreactive somata in the ventral paired mediolateral cluster (VPMLC)
and the ventral paired posterolateral cluster (VPPLC). In the VPMLC most of the several 1000 small globuli
cell somata and about 10 larger somata are immunoreactive. In the VPPLC only a subpopulation of the globuli
cells shows immunoreactivity. Our results show that FMRFamide-like peptides are widespread in the olfac-
tory brain of the spiny lobster. Biochemical and electrophysiological experiments are under way to elucidate
the potential function of these peptides in the central processing of olfactory information. (Supported by
DFG award Schm 738/1-1 and NSF award # 88-10261.)
87. A computerized system to analyse olfactory acuity and searching strategies in

the ferret
Manuel Schneidereit and Raimund Apfelbach
Department of Zoology, University of Tubingen, Aufder Morgenstelle 28, D-7400 Tubingen, FRG
It is well established that in the carnivorous ferret olfaction plays a dominant role in the recognition and
detection of prey objects [Apfelbach (1986) Behav. Proc., 12, 363]. Yet, there is little known about the
olfactory capacities in this mammal. This lack of knowledge is partly due to inadequate testing procedures
for the ferret. We therefore present a computerized Y-maze which allows us to ask for odor discrimination,
searching strategies and even threshold evaluation.
Odor vapors are generated and diluted with air to the desired concentration in an olfactometer and are
subsequently delivered to the animal located in the Y-maze. The Y-maze is made of plexiglass; at seven
points infrared sensors are installed to monitor the animals movements and behavior. The hardware consists
of a microprocessor INTEL 8052 AH with a 16 kB RAM and an EPROM 27128, one seriell interface (RS
232), 24 output ports and 16 input ports including one printer port. Accuracy of the system is 5 ms. The
software includes three programs introducing the stimuli and the positive reinforcement to the animal step
by step, and one testing program. It also controls the valves delivering the odor/air stimuli and the infrared
sensors. All programs are resident stored in an EPROM. Besides the computerized monitoring additional
408
ECRO IX: Abstracts
observations may be added by the experimentor via a keyboard. In each experiment up to 1600 data are
recorded and stored. Data are printed at the end of the experiments. Data can be transfered via an RS 232
(ASCII) in a data base system (Open Access) which allows further analysis.
88. Odour quality? An intriguing problem!

P.Schudel
Givaudan Forschungsgeseltschaft AG, DQbendorf, Switzerland
Odour and quality are perceptual terms. Natural science's methodology is based on certain criteria, agreed
upon since Galileo Galilei. It does not give us access to such qualities we perceive as odours. As a conse-
quence, we as scientists can look at odour as information bound to molecules or electromagnetic fields within
our brains as information carriers. Therefore, the related problematic nature of describing odours forms

an ongoing topic of discussion in view of improvements within fragrance research. Based on the plausible
assumption that the highly dynamic, interrelated, complex patterns of activated and non-activated brain neurons,
emerging while smelling, do not represent what we perceive as odour, we can conclude that everything we
smell (as well as touch, hear, taste and see) is no more than a human concept of what there is. It will be
shown that, therefore, odour quality is a question: (i) components, (ii) blend, (iii) receptor and neurological
status of the individual smelling as well as (iv) the individual's interpretation (human concept of the brain
signals).
89. Inhibition of rat olfactory cytochromes P-448 by odorants

Michael S.Skinner, Stan Lovett and George H.Dodd
Ofaction Research Group, University of Warwick, Coventry CV4 7AL, UK
We have previously reported that the odorant and specific olfactory toxin 3-trifluoromethyl pyridine (TF)
is a potent inhibitor of rat olfactory P-448 (P450IA1 and P450IA2) -dependent activity.
We now report that the odorant 3-methylindole (3MI), another specific olfactory toxin, is also a potent
inhibitor of rat olfactory P-448-dependent activity.
We have examined the effects of a large number of odorants, including some methylene dioxyphenyls,
on rat olfactory P-448-dependent activity. We report that some are potent and selective inhibitors of this activity.
These results are discussed in relation to current ideas about peri-receptor events in olfaction.
90. Receptor-site mapping in olfaction and the molecular basis for olfaction
Everett W. South wick
Philip Morris Research Center, Richmond VA 23261, USA
The correlation of chemical structure with odor properties is only in its infancy. The odor profile methodology
developed by ASTM has been automated in our laboratories. Odor data for nearly 1400 compounds are stored
in a searchable data base in the form of 'odor profiles'.
Factor analysis of the odor profile database has been applied to identify compounds, i.e. acylpyridines,
with a high factor score for popcorn-nutty. A systematic study of the structural requirements for popcorn-
nutty character in this series of compounds has led to the delineation of a receptor-site map. The latter has
been interpreted to provide an hypothesis for the molecular basis of olfaction which parallels that in vision.
91. The bitter taste of sucrose octaacetate is IP3 mediated

A.I.Spielman, T.Huque2, J.G.Brand1 and G.Whitney2
New York University College of Dentistry, 'Monell Chemical Senses Center, Philadelphia and 2State Univer-
sity of Florida, Tallahassee, USA
The peripheral mechanism of mammalian bitter taste has been the focus of recent investigations [Spielman
et al. (1989) Brain Res., 503, 326]. The purpose of this study was to understand the nature of the second
messengers involved in the signal transduction of bitter taste of sucrose octaacetate (SOA).
Mice belonging to the B6.SW and C57B1/6J strains are congenic and differ only in their tasting abiiry
of SOA [Whitney and Harder (1989) Behav. Genet., 16, 559]. Taste tissue from vallate and foliate papillae
409
ECRO DC: Abstracts
from both strains, taster and non-taster of SOA, were used to obtain membrane preparations [Spielman et
al. (1989) Oiem. Senses, 14, 841]. Accumulation of IP3 was monitored in the presence of SOA (10~ 4 M)
alone, or in association with hydrolysis resistant GTP analogoues (10~ 3 M) or A1F~4 (10~ 2 M).
Membrane preparations derived from taste tissue of the SOA taster strains demonstrated a 180% increase
in IP3 accumulation over non-taster (control) mice-derived preparations, but only in the preparation of SOA,
GTP7S and 278 nM Ca 2 + . IP3 seems to be the second messenger of the bitter taste of sucrose octaacetate.
SOA tasting thus appears to be a Ca 2+ dependent process that requires the presence of a GTP-binding pro-
tein. (This study was supported by BRSG funds from Monell Chemical Senses Center, Philadelphia and
by BRSG grants RR 07062 and RR 05332 from New York University.)
92. Quantifying the gustofacial and nasofacial reflexes

J.E.Steiner

77K Hebrew University Hadassah, Faculty of Dentistry and Medicine, Jerusalem, Israel
The gustofacial and nasofacial reflexes, described in our earlier studies, are reliable and valid indicators
of the organism's motivation, towards the quality and hedonics of triggering stimuli. Based on these expressive
behavioral reactions it is easy to classify the stimuli as (i) liked, palatable ones, in contrast to (ii) disliked,
aversive ones; or as (iii) indifferent ones. Careful systematic analysis of video-documented sequences of
the elicitation of the behavioral stereotypes present adequate methods to quantify and to compare different
degrees of liking towards a chemostimulus; this for theoretical studies or for assessment of consumer liking.
In a previous study we introduced the analysis of expressive motion sequences according to a notational
system. The relative occurrence of the individual motion components across all tested individuals under each
stimulus-conditions, expressed in percentage, was found as an easy-to-handle highly indicative value [Ganchrow
et al. (1983) Inf. Behav. Dev., 6, 189]. Later, neonate infants' liking of glutamate salts and other tastes
was quantified by similar procedures [Steiner (1987) In Umami: a Basic Taste, Dekker, New York]. Besides
quantitative analysis of motion features, numerical hedonic estimates of the entire response sequence can
serve as a quantitative measure. Such ratings can be correlated with subjects' self-estimates. Finally, latency
and duration of expressive behaviors in relation to stimulus presentation can be used as indicative parameters.
In recent studies we used these quantitative methods. Among other findings, the studies evinced that (i)
quantification of behaviors, indicative for liking of chemostimuli in animals is easily achieved; (ii) interstimulus
rinses have only little or no effect on the temporal and spatial organization of the neonate's gustofacial response
to different, subsequently presented tastes; (iii) autistic adolescents' taste and odor liking is similar to that
of their normal agemates; (iv) patients affected with Alzheimer's disease differentiate between pleasant and
aversive tastes and odors as do their normal age-mates; (v) in psychic depression, liking of sweet is dif-
ferent, but dislike of bitter is similar to that of control subjects. (Sponsored by a grant from Mrs and Dr
Sigg, Switzerland.)
93. Second messengers modulate ion channels in cultured olfactory receptor neurons
from male Manduca sexta pupae
M.Stengl, R.B.Levine and J.G.Hildebrand
ARL Division of Neurobiology, University of Arizona, Tucson, AZ 85721, USA
Males of the moth Manduca sexta locate their mating partners by detection of species-specific female sex
pheromone via male-specific olfactory receptor neurons (ORNs). To investigate the primary sensory transduction
mechanisms of pheromone detection, antennae from stage 3 male M. sexta pupae were dispersed into single
cells and kept in long-term primary cultures. Among the diverse antennal cells in our cultures, the ORNs
could be identified with two monoclonal antibodies, one of which specifically recognizes mature pheromone-
sensitive ORNs in situ. Patch-clamp studies demonstrated that the cultured ORNs express at least three
different kinds of K + channels and at least one type of a TTX-blockable Na + channel after 3 weeks in vitro.
In 3-week-old cultures about 30% of the ORNs responded to puffer-pipette-applied DMSO extracts of female
pheromone glands. In most cases an inwardly rectifying unspeciftc cation channel (L^,) opened in cell-attached
recordings. Current through this unspecific cation channel was the only inward current observed in ORNs
at potentials more negative than the resting potential (-60 to - 8 0 mV). Since G-proteins and second messengers
appear to play a role in insect olfaction, we investigated their possible involvement in the gating of the 1^,.
With GTP7S included in the patch-pipette during whole-cell recordings from 2-4-week-old cultured ORNs
(n = 11), in at least 27% of the experiments an inward current appeared at hyperpolarizing potentials. With
410
ECRO K : Abstracts
IP3 included in the patch-pipette (n = 20), an even stronger and quicker inward current developed during
hyperpolarization in the whole-cell mode in at least 45 % of the cultured ORNs. Application of phorbol ester
(n = 34) with a puffer-pipette in the cell-attached mode or included in the patch-pipette opened the 1 ^ or
was followed by depolarizing inward currents at hyperpolarizing potentials in 49% of the ORNs in vitro.
Future experiments will examine whether the observed second messenger or G-protein modulation of the
ion channels in cultured ORNs interacts with their ability to respond to their species-specific pheromone.
(Supported in part by NIH grant AI-23253.)
94. Intracellular generation of cyclic AMP in the circumvallate taste buds cells
of the rat in response to extracellular signaling by sucrose
Benjamin J.Striem, Michael Nairn and Bernd Lindemann1
Department of Biochemistry and Human Nutrition, Faculty ofAgriculture, The Hebrew University ofJerusalem,

Rehovot 76100, Israel and 'Department of Physiology, UniversitOt des Saariandes 6650 Homburg, Soar, FRG
Epithelial sheets rich in taste buds and free of muscle tissue were isolated as the U-shaped cleft which surrounds
the circumvallate papilla of the rat tongue. The circumvallate tissue from one tongue (8—14 /ig protein)
was cut in two approximately equal parts which were incubated with phosphodiesterase inhibitor and zero
or 150-600 mM sucrose. After 6 min of incubation, tissue was washed, the cells permeabilized and their
cAMP content determined by radioimmunoassay. Paired estimates from the same animal showed a significant
sucrose-induced cAMP production. This increase in intracellular cAMP was dependent on the sucrose
concentration and was suppressed by about 65% when 50 mM of methyl 4,6-dichloro-4,6-dideoxy-a-r>
galactopyranoside (kindly provided by Dr William Jakinovich Jr), a known competitive inhibitor of the sucrose
sweet taste, was added. Epithelial sheets free of taste buds did not respond to stimulation either by sucrose
or by the sweet taste inhibitor. Results are in line with previous suggestions that cAMP is involved in sucrose
sweet-taste transduction in the rat. (Supported by grant 1-100-181 from GIF (the German - Israeli Foundation)
and by the Deutsche Forschungsgemeinschaft (SFB 246, project Cl).)
95. Characterization of olfactory receptor cells in insects: electrophysiological and

immunological approaches
J6rg Strotmann, Erwin Tareilus, Jorg Wegener and Heinz Brcer
The initial electrical event in olfactory receptor cells stimulated by odorants is a graded membrane depolarization,
i.e. a receptor potential, supposed to be generated by activation of specific ion channels. In order to explore
the ion channel endowment of antennal receptor cells, patch-clamp recording techniques have been employed
studying dissociated antennal cells; the receptor cells were readily identified by their bipolar morphology.
In cell-attached and -excised patch—clamp analysis Ca2+-dcpendent K+-channels displaying different con-
ductances have been recorded and further characterized. In inside-out patches applying different
Ca2+-concentrations it was demonstrated that elevated calcium concentrations greatly increased the open-
ing probability of the channels. Furthermore a low-conductance, voltage dependent K+-channel type was
recorded; in embryonic antennal cells a slow-inactivating Na+-channel has been detected.
Towards a chemical identification of olfactory receptor cells, immunological approaches, including the
hybridoma technique to generate monoclonal antibodies, have been employed. In differential screening assays,
monoclonal antibodies were detected which selectively react with antennal membrane preparations. One of
the antibodies recognized a protein that was exclusively expressed in receptor cells and heavily labelled the
antennal nerve. Another monoclonal antibody selectively labelled a population of supporting cells and showed
no reaction with the antennal nerve. The epitope was only accessible just before hatching and was not detectable
if protein synthesis was blocked.
96. A truffle-hunting apparatus based on conducting polymer sensors

Thierry Talou and Krishna C.Persaud1
Laboratoire de Chimie des Agroressources, ENSCT, 31077 Toulouse Ctdtx, France and 'Department
Instrumentation and Analytical Sciences, UMIST, Manchester M60 1QD, UK
Black truffles are the fruiting bodies of the hypogeous fungus Tuber melanosporum vitt which grow in sym-
biosis with certain trees, especially oaks. During its limited harvesting season, mainly winter, one finds them
411
ECRO K : Abstracts
in several regions of southern Europe, especially Spain, Italy and France. Their harvesting is a delicate and
always uncertain operation. Indeed, because truffles remain underground even after maturation, the acute
sense of smell of certain animals is required to find them. Traditionally, pigs, dogs, and the presence of
a particular fly (genus Suillia) which is attracted by truffle odour, are used to localize underground truffles.
Although efficient, animals have a short concentration span and tire rather quickly. So, electronic odour
detectors capable of localizing buried truffles may provide a more efficient way for truffle harvesting. After
testing portable electronic gas detectors (Talou et al., ECRO VIII, poster 131), we have conducted suc-
cessfully field tests with a self-constructed hand-held 'sniffer' using conducting polymer sensors. The system
consisted of a small pump drawing air (2.5 L/min) over the sensor surface which has multi-element odour
sensors based on electrically conducting polymers (electrochemically polymerized heterocyclic molecules).
The array of sensors operates with rapidly reversible adsorption/desorption kinetics of change in electrical
conductance when exposed to polar volatiles. The sensor readings were integrated and displayed on a small
digital meter which gave a measure of odour concentration. Sensors were selected to have higher specificity
to truffle odour than to the surrounding humus odour. The efficiency of this prototype appears to be quite

good, and rigorous field trials will be planned for the next truffle production season.
97. The behaviour of the scent-tracking dog

Aud Thesen
The ability of a dog to follow a scent U remarkable by any measure. It's a question about life and death
for wild dogs to follow a prey in the right direction. Once he has found the track, the dog makes his decision
as to direction in only a few seconds. Whether the subject walks forwards or backwards, the dog is still
able to track direction. The dog easily detects continuous trails, but is unable to detect in which direction
the subject has moved [Steen and Wilsson (1990) Ada Physiol. Scand.]. These findings indicate that the
directional cues to the dog's nose are contained in discontinuous and not continuous trails. It is probable
that the dog is capable of perceiving the minute differences in odor concentrations between consecutive footsteps
of a track [Derving (1990) Cell Biol, 39s, 271].
To achieve a better understanding of how the dog is able to determine the track direction, I have performed
simultaneous video recordings of the dog and his sniffing activity. The sniffing activity was monitored with
a wireless microphone fastened to the forehead of the dog. The data will be analysed to observe: (i) the
dog's behaviour in relation to the footprints in marked areas, (ii) the variation of sniffing activity at the point
of track detection (iii) and the time interval between detection and determination of track direction. Further,
the recordings will be used to ascertain the way the olfactory sense is used when the dog is tracking. A
videorecording of the dog's behaviour will be demonstrated at the poster presentation.
98. Selective discrimination of bark beetle olfactory receptor cells among host
volatiles
Bjem Age Temmeras
Department of Zoology—AVH, University ofTrondheim, N-7055 Dragvoll, Norway
How odours of the complicated mixtures from trees are received by the receptor cells of bark beetles is
of major importance in any manipulating plan using host or non-host volatiles. This regards both which com-
pounds of the blend can be received by the antenna and how these compounds activate the olfactory system
of the beetles. In bark beetles olfaction is considered to be the most important sense for host recognition
and acceptance. In the common spruce beetle, Ips tyopgraphus it is furthermore demonstrated that the beetles
discriminate between different ages and quality of the trees, and prefer a host of certain condition. These
beetles should therefore be a proper species to find out which components of the host odorous blend really
activate olfactory receptor cells and how the discrimination system is on the receptor level.
In previous electrophysiological studies it has been difficult to find out how host volatiles arc received
by the receptor cells, mainly because adequate synthetic compounds have not been available. In later studies
[TBmmeras and Mustaparta (1987) /. Comp. Physiol., A 161, 705] Unking gas-chromatographical (GC) separa-
tion of the host odour bouquet to single-cell recordings, the questions about which constituents of the host
volatiles are important for the beetles and how individual cells receive each constituent, in principle can
be solved. While the results clearly showed which of the odour components, e.g. the major ones, were of
no significance for the beetles, it was much more difficult to point out among the minor components which
412
ECRO DC: Abstracts
were important cues. These complications are related to problems previously encountered during analysis
of plant odours, and may concern stabilities of the volatiles in air, low amounts of the particular compounds,
adsorption in the extraction medium, changes at high temperature in the GC-oven and interaction with the
medium in the GC-column.
To minimize the problems of loosing compounds, now new extraction procedures are introduced. The
use of logs inside 200 1 aluminium drums sucking cleaned air through and using Porapak as traps, gave extracts
containing a larger variety of odorous compounds with molecules of unchanged structure—as far as known.
Also, in the GC-procedures improvements have been done, e.g. different capillary columns and new split
system, resulting in more compounds unchanged through the column and the transfer tube to the preparation.
The cells recorded using the mentioned improvements often showed another picture than previously. Almost
all cells first responding to a piece of bark stimulated via syringe, now also responded to fractions from
an extract used as stimulation via the GC. Most cells responded to more than one fraction, two or three,
and the cells can be grouped according to responses to specific fractions with no overlap. It therefore seems

that the methods used really can identify compounds olfactory active on the beetles antenna and open for
tests of their behavioural meaning in the choice of suitable host.
99. Nervous activity of the lateral olfactory tract of the cod upon chemical stimula-
tion of the olfactory organ
Arne Torvik
In cod specific behaviour patterns can be evoked by electrical stimulation of olfactory tract bundlets [Deving
and Sclset (1980) Science, 207, 559]. This feature permits a description of the anatomical and functional
properties of the substrate that participates in the execution of each behaviour pattern. The cod react to
taurocholatc with snapping movements [Hellstrem and Eteving (1986) Behav. Brain Res., 21, 155] and glycine
with bottom food search [Ellingscn and Daving (1986) / . Chem. Ecol., 12, 155]. These behaviour patterns
are also seen when the medial and the lateral parts of the lateral olfactory tract (LOT) are stimulated electrically.
In the present investigation the nervous activity of bundlets of the LOT was studied while stimulating the
olfactory organ with taurocholate and glycine at concentrations between 10"' 3 and 10~6 M. Multi-unit
activity was transferred to standardized pulses and counted in 1 s time bins. Nervous discharges increased
over the background spontaneous activity with concentrations of glycine and taurocholate at about 10~ 10 M.
In some of the preparations the increase in nervous activity in the medial and lateral part of the LOT was
dependent upon the substance tested. The results of the present investigation will be discussed in relation
to the organization and spectrum of fiber diameters of the axons in the LOT.
100. Isolation of sex pheromone of female yellowfin baikal sculpin, Cottocomephorus

grewingki (Dyb.)
R.B.Valeyev and T.M.Dmitrieva
Institute of Biology, University of Irkutsk, USSR
Elucidation of chemical structure of sex pheromones is the main and difficult question of fish chemocom-
municalion. We are undertaking an attempt to isolate and to identify these physiologically active compounds.
The subject of research is the endemic of lake Baikal, namely, yellowfin Baikal sculpin, having three popula-
tions. Its spawning period takes place in March, May and August. It allows us to intensify the work. Both
male and female of this species widely use the pheromonal signals for various interrelations between themselves
during spawning period. We have found that a female sex pheromone is released with ovarian fluid after
ovulation.
The aim of this research was to isolate the female sex pheromone of the yellowfin Baikal sculpin. Five
fractions have been obtained from 'eggs wash off* by successive extraction with organic solvents of increas-
ing polarity. The biotesting, consisting in the registration of typical patterns of male sexual behaviour, have
shown that the hexane fraction is active. Three active components of female pheromonal signals have been
isolated from the hexane fraction by TLC. One of these components is a primer pheromone and induces
spermination. Another isolated component stimulates male spawning behaviour.
Thus, it was established that the female sex pheromone in yellowfin Baikal sculpin consists of at least
three components, having various function.
413
ECRO K : Abstracts
101. How do we test what the nose tells the brain about odours?
Steve Van Toller, John Behan1, Martin Kendal-Reed and Pauline Howells1
Department of Psychology, University of Warwick, Coventry, CV4 7AL and 'Quest International, Ashford,
Kens, UK
Over the last ten years the Warwick Olfaction Research Group has used psychophysiological techniques to
investigate human responses to odours [Van Toller (1988) Internal. J. Cosmet. Sci., 10, 171]. Recently,
these studies have involved the use of real time electrical activity of the cortex using a technique involving
topographic mapping of the brain [Duffy (1986) Topographic Mapping of Brain Electrical Activity. London,
Butterworth]. The technique used at Warwick (Neuroscience Series III Brain imager) collects EEG activity
whilst the subject is being presented with an odour. This is achieved by fitting the subject with a cortical
array of 28 electrodes, inset into a fabric headcap. At the Eighth ECRO Congress, preliminary qualitative
results were presented showing typical topographies obtained whilst subjects were smelling various odours.
Since 1988 the research has been extended and further qualitative and quantitative findings obtained.

The basic experimental paradigm used in the Warwick laboratory will be presented and its potential examined.
In addition, some of the latest results will be shown. These will include cortical activity obtained from subjects
smelling the compound 5-alpha-androstan-3-one. The olfactory psychometric response to this interesting
biological compound falls into three categories: (i) specific anosmia to the smell; (ii) pleasant rating of the
smell; (iii) unpleasant rating of the smell. Cortical activity from each of these three odour evaluation categories
show distinct patterns. Cortical patterns will also be presented from subjects who lack a sense of smell.
102. Subfamilies of odorant binding proteins associate with specific olfactory

receptors
R.G.Vogt and M.R.Lemer
Yale University School of Medicine, New Haven, CT 06510, USA
In contrast to aquatic animals which smell water-soluble odorants, terrestrial animals smell lipophillic odorants
which do not readily dissolve in the mucus or lymph surrounding olfactory neurons. Odorant binding pro-
teins (OBPs) have been described from terrestrial insects [Vogt and Riddiford (1981) Nature, 293, 161; Gyorgyi
et al. (1988) Proc. Natl Acad. Sd. USA, 83, 1418; Vogt et al. (1989) J. Neurosci., 9, 3332; Raming et
al. (1989) FEBS Let., 256, 215] and vertebrates [Pelosi et al. (1982) Biochem. J., 201, 245; Pevsner et
al. (1988) Science, 241, 336; Pelosi et al. (1990) In Chemosensory Information Processing. NATA AS1
Series H, Cell Biology, vol 39, pp. 125- 130. (Springer-Verlag, Berlin]. These OBPs reside in the fluid
surrounding olfactory neurons, and are thought to help solubilize lipophillic odorants into this fluid and thus
allow efficient movement of odorants through the fluid to the odorant receptors. Insect and vertebrate OBPs
show no cross phyla homology, based on full length amino acid sequence analysis. This is consistent with
these animal groups becoming terrestrial at independent times, adapting their olfactory systems to terrestrial
life by selecting available but different gene products for OBP source material.
The insect OBPs so far described have been male-specific pheromone-binding proteins (PBPs), associating
with pheromone-specific neurons. However, if OBPs are required for terrestrial olfaction, they should also
associate with non-pheromonal or general olfactory neurons. Such neurons are present on both sexes, while
the pheromone specific neurons are found only in males. By comparing N-terminal amino acid sequences
of antenna] specific proteins we have identified additional subfamilies of insect OBPs which associate with
general-odorant specific neurons. We have called these general-odorant binding proteins, or GOBPs. A striking
and contrasting feature of the PBPs and GOBPs is that the PBPs from different species are quite variable
in sequence, while the GOBPs from these species are nearly identical. This difference follows a pattern of
molecular tuning of OBPs toward specific odorant ligands. As such, not only do different insect OBPs mark
specific odorant pathways, but they may also act as peripheral filters determining what odorants will be
solubilized and thereby gain access to receptors.
103. Isolation and molecular cloning of a tissue specific membrane protein which
is a major component of olfactory dendrites and which possesses odorant binding
properties
R.G.Vogt and M.R.Lemer
Yale University School of Medicine, New Haven, CT 06510, USA
Odorant detection is thought to be mediated through receptor proteins associating with the membrane of
olfactory cilia/dendrites, though the isolation of such proteins has been elusive. A 69 kd protein of the silk
414
ECRO IX: Abstracts
moth Antheraea polyphemus was previously implicated as an olfactory receptor protein: it was a component
of the olfactory dendrite membrane; it could be labeled with a photoaffinity analog of a sex-pheromone odorant,
and labeling could be blocked by scx-pheromone; it was specific to olfactory tissue [Vogt et al. (1988) J.
Biol. Chem., 263, 3952]. In recent studies, in vivo labeling with 33S-methionine has shown that a 69 kd
protein is a major component of olfactory dendrite membranes in two moth species, that it is absent from
other neuronal tissue, and that it is developmentally expressed at a time coincident with other olfactory pro-
teins. We have isolated olfactory dendrite membranes from 800 male A.potyphemus antennae, and elec-
trophoretically purified the 69 kd protein in sufficient quantity to obtain a partial N-terminal amino acid
sequence. We have designed PCR primers to the N-terminus, and have synthesized, purified, cloned and
sequenced an 89-base PCR product corresponding to the N-terminal region. Based on this information, we
constructed a 65-base oligonucleotide which was used as a probe for Northern blot analysis of mRNA. Both
PCR and Northern blot analyses confirm that the mRNA encoding the 65 kd protein was specifically expressed
in olfactory tissues. We have successfully used the 65-base probe to screen oligo-dT and random primed

cDNA libraries generated against antennal mRNA. The properties of the 69 kd protein will be presented,
with respect to tissue distribution and deduced amino acid sequence.
104. Linked GC-single cell recordings in identification of host odours for the pine
weevil (Hylobius abietis)
A.Wibe and H.Mustaparta
Department of Zoology, University of Trondheim—A VH, Norway
A central question in insect—plant relationships is which components of the volatilises from plants are used
by herbivorous insects in recognition of host plants. The pine weevil (Hylobius abietis), feeding on pine
and spruce, is strongly attracted by odours from their hosts. The aim of the present study was to identify
host odour components of the pine weevil, by linking gas chromatography (GC) to electrophysiological
recordings from single receptor neurons (using tungsten microelectrodes).
Odours from host plant seedlings were trapped by a polymer (Porapak Q) using 'headspace' procedure,
eluted by hexane and used as test extract. When a receptor neuron responded strongly to vapour of host
material and an extract, a sample of the latter was injected into the GC column. A fused silica capillary
column with a split-system was used, leading half of the effluent to the GC-detector and the other half to
the insect antenna. After separation, the fractions were thus tested one by one on the receptor neuron and
recorded simultaneously with the GC-detector recording.
Successful recordings have been carried out for 15 receptor neurons. When a fraction triggered a response
in a host odour neuron, the spike frequency rose from 0 - 5 imp/s to 2 0 - 5 0 imp/s. Some neurons responded
to only one fraction in the extract, whereas others responded to two or three. Nine different fractions trig-
gered responses in the host odour receptor neurons. Six of them activated more than one unit. Three frac-
tions, belonging to the major constituents in the extracts, were analysed using connected GC/TR (infrared
spectroscopy) and GC/MS (mass spectroscopy), and identified as a-pinene, /3-pinene (which confirms earlier
results, testing synthetic compounds on the receptor cells) and limonene.
In conclusion, each host odour receptor neuron in the pine weevil seem to possess a high specificity for
one or a few components in the odour extract. Furthermore, the results demonstrate that the linkage of GC
with electrophysiology is a suitable technique in the process of identifying host odours for the pine weevil.
105. Adult low vertebrates functionally and morphologically regenerate peripheral

and central olfactory pathways
H.P.Zippel
Physiologisches Institut der UniversitOt, Humboldtailee 23, D-3400 Gdttingen, FRG
The present behavioural and neuroanatomical investigations in adult goldfish have been designed (i) to examine
the relationship between morphological and changes in the perception of spontaneously preferred olfactory
(food) stimuli before and after inflicting lesions in the olfactory system, (ii) to test the animals ability to
respond to a stimulus learned preoperatively following regeneration, (iii) to investigate the animals ability
to discriminate between two adours, one of which had been rewarded preoperatively ('specific' regenera-
tion), and (iv) to test for functional and morphological regeneration pathways, and after partial ablation of
olfactory bulbs. After receptor axotomy and following rostral bulbectomy surprisingly short functional regenera-
tion ( 7 - 1 0 days) was observed. At this time degenerative processes are at their maximum in other studied
415
ECRO K : Abstracts
vertebrates. From previous observations [Zippel et al. (1970) Z. vergl. Physiol., 69,79] and from the HRP-
cases reported here, it is evident that innervation of the goldfish olfactory bulb is comparable to that of intact
animals. Behaviour acquired in a difficult, long-lasting discrimination training is present after regeneration
(i.e. highly specific). Virtually nothing is known in vertebrates about the central projections after surgical
interruption of connection with the telencephalon. After complete bilateral bulbectomy neither a functional
regeneration [Zippel et al. (1981) In Lesion-inducedNeuronal Plasticity in Sensorimotor Systems, pp. 377-394,
Springer], nor a morphological regeneration (Grazaidei, pers. comm.) was found. From the present data
it is however evident that after surgical interruption of the medical and the lateral substracts and even after
bilateral caudal bulbectomy, a functional and a morphological (HRP-injections into the telencephalon, and
into the olfactory bulb) regeneration occurs. An olfactory bulb of half the normal size and extremely thin
connections to the telencephalon are evidently sufficient for regaining pretraining behaviour. Whether or
not all the afferent and efferent pathways previously described for the goldfish [Bartheld et al. (1984) Cell
Tiss. Res., 238, 475] are regenerated still remains open.

106. Permanent olfactory dysfunction and intermittent exposure to H2S
A.P.J.Hendriks and P.Boelens
Psychological Laboratory, Utrecht University, Sorbonnelaan 16, 3584 CA Utrecht, The Netherlands
The initiative for this study was taken by one us (P.B.) after knowledge of olfactory complaints of some
workers who had been regularly exposed to hydrogen sulfide.
Three groups are studied. Group I consists of 1 female and 35 males (mean age 39.58; SD 4.99) who
have been intermittently exposed to H2S during their professional activities for at least 2 years. Concentra-
tions varied, but were most of the time below the TLV of 10 p.p.m. In addition, these employees have been
exposed to benzene. On chronic exposure, this compound is considered responsible for permanent hyposmia
[Ferrari and Jemmi (1966) Ada Biomedica Aleneo Parmense, 37, 207]. Group II consists of 35 male employees
(mean age 38.63; SD 5.73) with exposure to benzene under the same working conditions as group I. Group
m consists of 1 female and 28 male employees (mean age 39.24; SD 6.67) with no known exposure to either
H 2 S or benzene. All 100 employees work for the same company.
The employees have been individually tested with the odour identification test GITU. This test consists
of 36 well-known natural odourants. Subjects must find each odourant's right name by selecting one of four
odour descriptors. The test has been described elsewhere [Hendriks (1988) Rhinol., 229]. From the subjects
used to calibrate the test an extra control group of 36 persons, group IV, has been formed that matches group
I with respect to number, sex, age and smoking.
Mean GITU-scores and standard deviations are; for group I 31.28 and 7.63; for group II 34.86 and 1.19;
for group III 34.55 and 1.64; for group IV 34.65 and 1.66. The scores of group I are significantly lower
than the scores of each of the three other groups, while the scores of these other groups do not differ from
one another (Wilcoxon-Mann-Whitney rank sums test; P = <0.05, one-tailed).
From all 100 employees, only five have a Grnj-score less than 30. These five belong to group I. Their
scores indicate anosmia (scores of 8, 13 and 13) or severe hyposmia (scores of 14 and 18). The incidence
of permanent anosmia for persons up to 50 years of age has been reported to be about 1.4% [Wysocki;
and Gilbert (1989) Ann. N. Y. Acad. Sci., 561, 12]. If this figure is a reliable estimate, then the probability
of finding at least 3 anosmics in a group of 36 persons under 50 can be calculated, using the binomial distribution
to be 0.0139.
It is concluded that H2S may cause serious olfactory dysfunction under the professional conditions as
described for group I.
416

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Chemical Senses Vol.16 no.4 pp.

Ninth Congress of the European Chemoreception Research

Downloaded from http://chemse.oxfordjournals.org/ by Sergey Novikov on February 16, 2014

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4. Odor response distributions and psychophysical functions generated

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10. Odor identification: filling in some blanks

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15. Preimaglnal conditioning of a solitary bee to nest odors

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25. Inhibition of the mitochondrial respiratory chain by perfumes and perfume

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28. Ion channels from insect olfactory receptor neurons

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34. Olfactory evoked potentials in clinical diagnosis

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36. Very short-term recognition memory for odors

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39. Pheromonal steroids of male yeUowfin Baikal sculpin Cottocomephorus grewingki

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46. Olfactory P300: lateralization of hedonics and discrimination performances

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51. Threshold perception of banana odour by a frugivorous bat: a case of

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54. Taste thresholds, detection probabilities and stimulated area