Experimental Dermatology 2002: 11: 448455 Blackwell Munksgaard .

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Copyright C Blackwell Munksgaard 2002

EXPERIMENTAL DERMATOLOGY
ISSN 0906-6705

Efcacy of naltrexone on acetylcholineinduced alloknesis in atopic eczema

Heyer G, Groene D, Martus P. Efcacy of naltrexone on acetylcholineinduced alloknesis in atopic eczema Exp Dermatol 2002: 11: 448455. C Blackwell Munksgaard, 2002 Abstract: Atopic eczema (AE) is a chronically pruritic inammatory skin disease. Although the mediators and exact mechanisms eliciting and sustaining pruritus are not completely known, AE patients in clinical trials have been shown to benet under treatment with morphine antagonists. Naltrexone (NAL) is a relatively pure morphine antagonist that blocks the effects of opioids twice as much as naloxone. NAL exhibits minimal pharmacological activity and displaces endorphines at mu- and kappa-receptors without its own intrinsic activity. NALs excellent oral bioavailability and linear increases in the area under plasma concentration-time curve make it ideal for use in experimental studies. We designed our present experiments similar to former experiments evaluating both peripheral cutaneous sensations and central itch procession in order to gain more information about the possible distribution of opioid receptors and their involvement in the pathophysiology of pruritus. Eleven AE patients participated in our double-blind study. Either 25 mg of NAL (NemexinA) or a placebo (PLA) was given to the participants 60 min prior to the acetylcholine (ACH) injection [intracutaneous (i.c.) injection of 0.02 ml of 0.55 M]. A PLA stimulus with buffered saline served as control on the opposite forearm. We used laser Doppler owmetry to measure the vasomotoric changes after ACH injection and recorded the duration and intensity of itch with a visual analogue scale (VAS). Following the evaluation of wheal and are sensation, we obtained the area of itchy skin around the injection site (alloknesis) by gently stroking the surrounding skin with a brush in the centripetal direction towards the injection site. The results were planimetrically evaluated. Oral NAL reduced the perifocal itch signicantly (P 0.009). In four of our observations the area of alloknesis completely disappeared. Itch duration was reduced by 20 s and the intensity of itch was diminished, yet not signicantly. NAL had no signicant effects on cholinergic vasoreactions measured by the laser Doppler (P 0.50) and especially failed to decrease the initial ux response, which is a typical sign of an altered vascular reaction (P 0.25). The decrease of wheal (P 0.008) and are (P 0.01) extension indicates an appropriate dosage of our treatment for this experiment. The most signicant effects of NAL were observed in parameters of itch processing such as alloknesis (P 0.009) and are extension (P 0.01). Therefore we favour the concept that NAL might have a stronger impact on central nervous mechanisms than on peripheral nociceptive structures.

G. Heyer1, D. Groene1 and P. Martus2

1 Department of Dermatology, Friedrich-Alexander University of Erlangen-Nuremberg, Hartmannstrae, Erlangen, Germany; 2 Department of Medical Informatics, Biometry and Epidemiology, Benjamin Franklin Medical Center, The Free University of Berlin, Hindenburgdamm, Berlin, Germany

Key words: atopic eczema pruritus acetylcholine naltrexone (NemexinA) Frau PD. Dr med. habil. G. Heyer, Dermatologische Universittsklinik, Hartmannstrae 14, 91052 Erlangen, Germany Tel.: 49 9131 8533662 Fax: 49 9131 8536175 e-mail: gisela.heyer/derma.med.uni-erlangen.de Accepted for publication 16 October 2001

Abbreviations: NAL: naltrexone; PLA: placebo; ACH: acetylcholine; NaCl: buffered saline; AE: atopic eczema; SDS: self-rating depression scale; LDF: laser Doppler owmetry; VAS: visual analogue scale; VIP: vasoactive intestinale peptide.

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NAL on acetylcholine-induced alloknesis in AE Introduction Pruritus is a major characteristic symptom of atopic eczema (AE) and can often be the most disturbing feature of inammatory skin diseases (1 4). Pruritus results in constant and vigorous scratching complicating the underlying disease by excoriations. Interruption of this scratch cycle may minimize the development of new eczematous lesions. Consequently, relief of pruritus is a treatment objective of paramount importance in clinical management. In AE the physiology of pruritus seems to be altered. Previous neurophysiological studies have disproved histamine as the main mediator (5, 6), and antihistaminic agents, which have been successfully used in other inammatory diseases, lack major antipruritic capacity in AE (710). Other mediators such as endopeptidases, kinins or neuropepides (9, 1113) that have been discussed as possible mediators appear to be largely mediated by a secondary release of histamine by depleted mast cells. Acetylcholine (ACH) causes itch independently (9, 14). Recently, we investigated patients with itchy skin during sweating and administered ACH and a vasoactive intestinale peptide (VIP), both being involved in sweat secretion (15). Both substances elicited pruritus, but in contrast to VIP, ACH-induced itch could not been blocked by the antihistaminic drug cetirizine. Consequently, ACH can be considered as a histamine-independent initiator of pruritus in AE. In the present study, we focused on the action of naltrexone (NAL) on ACH-induced sensations. NAL (NemexinA) is a pure opioid receptor antagonist with a strong blockade of mu-, delta- and kappa-receptors without its own intrinsic activity (16). After oral intake it is rapidly absorbed and results in a large intracellular and extracellular distribution (17, 18). The long-term efcacy of NAL on intractable uraemic and cholestatic pruritus has already been demonstrated in clinical studies (1921). Recently, NAL was examined in AE patients (22, 23). Nevertheless no data is available about the short-term efcacy on cholinergic pruritus. Opiate membrane receptors may be directly involved in either peripheral or central nervous processing of itch or both (24). They are known to inhibit the supraspinal pain perception and to inuence the action potential processing in the dorsal root. Strong itching, especially in the face, has frequently been observed after spinal applications and was prevented by morphine antagonists (25). Apart from central mechanisms, opiate memTable 1. Demographic and experimental data of 11 patients Subject no. 1 2 3 4 5 6 7 8 9 10 11 Sex, age x 33.9 years male 20 years male 28 years male 33 years female 61 years male 30 years female 25 years male 28 years female 40 years female 28 years female 52 years male 28 years SDS score x 39.7 44.5 33.5 50 42 37.5 33.5 32.5 40 40 45 38.5 ACH test: duration of itch: 3.05 min 2.20 2.20 1.07 3.50 2.30 6.02 1.10 3.56 2.02 1.54 2.02

brane receptors might interact with peripheral mechanisms. Intradermal injection of opioid peptides (e.g. FK 33-824 Enkephalin analogue) (26, 27) causes vasocutaneous reactions and potentiates itch in the indomethacin- and histaminedepleted skin. Both effects are inhibited by NAL. Hence mast cells released by histamine or prostaglandins are unlikely to be the cause of this potentiation. Lately Grando et al. discovered cholinergic membrane receptors at the surface of keratinocytes and demonstrated interaction with the release of ACH that is synthesized and stored in human keratinocytes (28). Pathological high release of ACH may stimulate muscarinic receptors on nociceptive tissues and induce electrical impulses directly. This theory is supported by the abnormally high concentration of ACH (29), muscarinic receptors on c-bers (30) and the increased serum concentrations of the endogenous mu-opiate receptor ligand beta-endorphine in patients with inammatory dermatoses (31). Materials and methods
Recruitment for the study was performed among a group of 16 outpatients who had reported a long history of AE. They had been under therapy earlier in the hospital but were in an eczema-free interval during the investigation. They were examined and evaluated by the same physician according to the clinical criteria of Diepgen et al. (32) and Hanin and Rajka (33). Medications (topical or systemic corticosteroids, opiates, UV therapy) were not allowed and if necessary withdrawn at least 8 days before the beginning of the study. In order to determine the presence of depression that may inuence the perception of pruritus (34), we asked our subjects to answer the 20 questions of the self-rating depression scale (SDS) and assigned them with the score according to Zung (35). Afterwards we continued to inject ACH into the skin and evaluated the duration of itch. Four of the 16 patients were excluded from the study because they did not exhibit pruritus. One subject initiated an UV therapy parallel to our experiments and was therefore excluded during the study. The demographical and experimental data of the remaining 11 patients are shown in Table 1.

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Heyer et al. Pretreatment

Pretreatment was with naltrexone (NemexinA, Du Pont Pharma GmbH, Bad Homburg, Germany) in a vehicle dragee [placebo (PLA)]. periments the cutaneous reactions were scanned (Agfa Gevaert N.V., Studioscan II, Kln, Germany) and evaluated planimetrically by Fotoscan Software.

Alloknesis
Slight mechanical stimulation induces perifocal itch sensation as in the area surrounding an insect bite. Alloknesis has been explained by the excitation of itch mediating central neurons through interneurons. They seem to receive input from low threshold, fast-conducting mechanoreceptor units that are gated by the primary pruritic stimulus-induced input from unmyelinated itch mediated nerve bres (36). In our experiment the area of alloknesis was assessed 10 min after application of ACH by gently stroking the surrounding skin with a soft brush. Starting at a distance of 810 cm from the injection site, the brush was gently passed over the skin in a centripetal direction. The border of the itchy skin was determined from six directions. Each point was marked and later connected to the others. This area was transferred to an acetate sheet and planimetrically evaluated (see cutaneous evaluation).

Solutions of injection
ACH (Acetylcholine ophthalmicum Dispersa, Niederwangen, Germany) was dissolved in saline (NaCl 0.9%, pH 6.7) at a concentration of 0.55 M for intracutaneous (i.c.) injection. Randomly selected, either 0.02 ml of ACH or the control stimulus 0.02 ml of NaCl was injected with a specially devised tuberculine syringe into the left and the right volar forearm of the subject. The concentration was chosen in accordance to the internationally accepted usage of ACH for these experiments.

Study design
This study is designed double blind, PLA and stimulus controlled. For intra-individual comparison of the treatment we held two consultations with each subject. According to the randomization either NAL or PLA was orally given 1 h prior to the session. ACH (stimulus) or buffered saline (control) was injected into the skin of the right and left forearm. Tests were performed between 15.00 and 17.00 hours. Room temperature was kept constant between 20C and 22C with the air humidity between 60% and 65%.

Data processing and statistical analysis

The primary focus of our study concerned the effect of NAL after ACH stimulus. This effect can be quantied by the following contrast: [effect (NAL,ACH) effect (PLA,ACH)] [effect (NAL,NaCl) effect (PLA,NaCl)] This contrast corresponds to the test of an interaction in a two-factorial ANOVA model (factors: stimulus, therapy). The assumption of normally distributed data was not fullled in our study therefore we used Wilcoxons signed rank test for statistical hypothesis testing. The secondary question of our study concerned the usefulness of ACH as stimulus in AE patients. This effect could be quantied by the contrast: [effect (PLA,ACH)] effect (PLA,NaCl)] The level of signicance was 0.05 (two-sided) in all statistical tests. For data management and statistical evaluations, Excel (Microsoft, Redmond, WA, USA) and SPSSWIN 9.0 (SPSS Inc. Chicago, IL, USA) were used. Dadisp (DSP Development Co., Cambridge, MA, USA) was used for editing LDF.

Laser Doppler uxmetry

Using a helium laser (MBF 3D, Moore Instruments Ltd., Axminster, UK), skin blood ux was recorded continuously by laser Doppler owmetry (LDF). Two laser probes were attached proximally and distally to the injection site using double-faced adhesive tape. The surface area of each probe for measurement of the cutaneous blood ow was about 1 mm2. Blood ow was measured with a time constant of 1 s and logged on a personal computer. Prior to the stimulus the baseline ux was determined over 60 s, then NaCl or ACH was injected into the skin and the increased blood ow was monitored during an observation period of 6 min. For evaluation, the LDF data were normalized to the average baseline values recorded during the rst minute prior to the injection.

Sensory rating
After the injection of ACH or buffered saline, the volunteers were asked to quantify their itch sensation on an electronic visual analogue scale (VAS). The left end of the 10-cm scale was dened as no sensation and the right end as maximal sensation. In 10-s intervals and using a tonal reminder over headphones, the subjects rated their itch intensity. The sensory rating stopped at the 6th min, as well as LDF measurements. The sensation rating data were evaluated by calculating the mean values of the reported ratings (area under the curve). Other sensations were reported verbally and further documented on questionnaires. Stinging and burning were regarded as painful sensation. Tickle was dened as minimal sensation and in contrast to prickle listed with the non-/minimal sensation group.

Results Evaluation of the SDS SDS levels above 51 outline a depressive symptomatic disorder, values above 61 accounts for a major depressive disorder. The mean SDS score of our subject group measured 37.9 with a range of 32.5/50. Reporting and rating of sensation As outlined in Fig. 1, NAL almost eliminated ACH-induced alloknesis in AE (P 0.009). In four of our observations the area of alloknesis was not detectable because the patients felt less than three itchy points after the mechanical stimulation. The others resulted in an alloknesis area of 1.02 cm2 with a maximal area of 2.39 cm2. All of the subjects

Cutaneous reactions
Wheal and are areas were outlined on transparent paper 10 min after ACH or buffered saline injection. Following the ex-

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NAL on acetylcholine-induced alloknesis in AE

Figure 1. Alloknesis area in mm2 elicited 10 min after ACH stimulus in subjects with NAL or PLA. The distribution of alloknesis is given as box plots. Condition (cond.) 2/4 indicates sessions with ACH injection, condition 1/3 outlines the saline control. The median, the 25th and the 75th percentile are presented. NAL reduced the perifocal itch signicantly. We found signicance between stimulus and the control.

Figure 2. Boxplots in cm2 showing the are development at the end of the session. Condition (cond.) 2/4 indicates sessions with ACH, condition 1/3 outlines the saline control. Median, 25th and 75th percentiles are presented.

with PLA felt more than three single itchy spots after mechanical stimulation, two of them with alloknesis areas bigger than 3 cm2. In the arm used as a stimulus control, the alloknesis measured 40 mm2 after NAL and 70 mm2 after PLA. In fact, NaCl could not effectively provoke alloknesis as compared with ACH (P 0.005). Following the injection of ACH, NAL decreased itch intensity from 5.96 to 4.95 arbitrary units, yet not signicantly (P 0.53). The maximal itch values were slightly increased but without a signicant effect (compare Table 2). Similar to the results of our pre-examinations, all subjects initially felt pain that slowly turned into itch. They started rating the itch after the 36th s and then continuously until the 3rd min. NAL succeeded to stop the sensation of itch 20 s earlier compared with

PLA (P 0.29). None of our subjects with NAL reported an itchy skin after the end of the session compared with three subjects with PLA. The stimulus control caused predominantly burning in our patients. However, NaCl elicited itch in three subjects but only to a very small extent, so that comparisons to ACH remained signicant as can be seen in Table 2.

Assessment of cutaneous vasoreactions NAL reduced are size signicantly from 9.2 cm2 to 5.87 cm2 as compared with PLA (P 0.01). Wheal size was diminished from 261 mm2 to 182 mm2, which is a signicant change (P 0.008). The effect of ACH was signicant for the area of the

Table 2. Mean, SD, range of pruritus given as the area under the curve (AUC) in the parameter of itch intensity. In addition, evaluation of maximal VAS rating is shown Sensory parameters Intensity of itch 0.530.003 Max. itch rat. 0.48 Area of sis (cm2) Effect of NAL P-values 1.06 (0.36.2) 0.01 0.009 Effect of ACH P-values 4.95 (0.311.5) 0.15 (0.011.1) 0.005 NAL/NaCl (cond. 1) 1.18 (0.33.7) 0.70 (0.11.4) 0.04 (00.45) NAL/ACH (cond. 2) 5.96 (2.615) 0.25 (00.6) 1.02 (02.39) 0.68 (01.8) 0.07 (00.78) PLA/NaCl (cond. 3) PLA/ACH (cond. 4)

1.87 (03.11)

All sensations were recorded in 10-s intervals during the observation period of 6 min on a VAS with the endpoints of no itch and unbearable itch. No signicant effects of NAL were found for intensity of itch and max itch rating. Signicant effects of ACH were found in all sensory parameters. The area of alloknesis is detected by stroking the skin with a brush. Less than three itchy points cannot be detected as area. Alloknesis was eliminated in four observations. The effects of NAL and ACH were signicant.

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Heyer et al.
Table 3. Mean, SD, range of cutaneous parameters are shown Cutaneous sensations Area of are 0.010.003 Area of wheal 0.0080.003 Effect of NAL P-values 241 (01538) 113 (41281) Effect of ACH P-values 5865 (180912890) 182 (64314) NAL/NaCl (cond. 1) 1036 (06690) 159 (28307) NAL/ACH (cond. 2) 9265 (396114909) 261 (95409) PLA/NaCl (cond. 3) PLA/ACH (cond. 4)

Wheal and are areas were detected with a scanner system following the evaluation of sensory parameters. Signicant effects of NAL were found for the area of wheal and are. Signicant effects of ACH were found for all cutaneous parameters.

Table 4. Mean and SD of the most important vascular reactions measured with the laser Doppler owmetry Vascular reactions T63% max laser 1 T63% max laser 2 Max ow laser 1 Max ow laser 2 Intensity laser 1 Intensity laser 2 End ux laser 1 End ux laser 2 NAL/NaCl (cond. 1) 106/34 173/72 0.09/0.11 0.11/0.14 0.73/0.94 2.31/2.87 0.02/0.04 0.09/0.13 NAL/ACH (cond. 2) 103/76 106/42 0.13/0.18 0.45/0.64 0.92/0.88 4.55/4.54 0.02/0.05 0.17/0.19 PLA/NaCl (cond. 3) 139/77 152/77 0.11/0.10 0.39/0.63 0.84/0.94 3.66/3.97 0.02/0.09 0.10/0.03 PLA/ACH (cond. 4) 145/79 140/62 0.13/0.18 0.37/0.21 0.94/1.06 6.05/3.53 0.03/0.04 0.28/0.18

In order to distinguish between initial vasodilatation and the axon reex are, we installed one laser probe closer to the injection point and the other in a distance of 1 cm from the point. All conditions are measured in mV. The intensity is calculated as AUC.

wheal (P 0.003) and the area of the are (P 0.003) (Fig. 2 and Table 3). Assessment of vascular reactions Soon after injection of ACH or saline, blood ow increased (compare Table 4). This was observed in every volunteer. Sixty-three per cent of the peak ow was recorded between the 103rd s (condition 2) and the 145th s (condition 4) in laser 1, the proximal laser and between 106th (condition 2) and 173th s (condition 1) in laser 2. The maximal LDF of the distal laser reached 0.13 arbitrary units (a.U.) with NAL application compared with 0.45 a.U. with PLA. After the 3rd min, LDF slowly decreased and reached end-ux levels close to the initial base values. No signicant changes were seen in the AUC; the parameter of intensity that comprises ux changes over the course of time. In comparison to the LDF values of ACH, the mean NaCl values rated lower indicating a less pronounced effect on the blood system. The distally placed laser probe laser 2 revealed continuously higher values than the laser probe close to the injection site. This is in agreement with published results and demonstrates the spread of erythema with a more pronounced vasodilation in the periphery. Discussion In some forms of dermatitis, notably AE, scratching behaviour secondary to pruritus is postulated 452

to play a major role in initiating and prolonging a cutaneous inammatory response. Therefore, an antipruritic agent has to sufciently suppress the itch induction in order to minimize the development and progression of the eczematous status and thereby reduce the necessity for extended periods of high potency therapies, e.g. corticosteroids. Efcacy and safety of morphine antagonists have been frequently observed (36, 38). Other studies have impressively demonstrated the favourable risk-to-benet ratio of NAL in diseases with intractable pruritus (1719, 36) and the inhibition of scratching behaviour in ICR mice (37). Recently, Metze and his group examined the antipruritic threshold of NAL (22,23). In a pilot study with AE, psoriasis, cutaneous lymphoma and other patient groups, they achieved a signicant therapeutic response in 35 of 50 patients within a week. As demonstrated in former studies, chemical mediators other than histamine play an important role in the chemical activation of sensory neurons in AE (6, 7, 12, 13). One of these may be ACH a neurotransmitter that is also increased in AE skin (29) and may be directly involved in the pathomechanism of pruritus (6, 1315). Recently, our group discovered that intracutaneous injection of ACH evoked itch in inammatory diseased skin of psoriasis and AE (1, 14). Guided by the idea that actions of ACH on the nociception might be modulated by morphine derivates and depend on the opioid receptor system, we established this study to gain some information about:

NAL on acetylcholine-induced alloknesis in AE 1. The efcacy of NAL on pruritus perception, conductance and central itch progression caused by ACH injection. 2. Bioavailability of NAL seen in the cutaneous parameters of are and wheal. 3. Risk-to-benet ratio of the experimental dosage of 25 mg of NAL. Pruritus is a very subjective sensation and has to be regarded with respect to psychology, anxiety, depression and stress perception (39). As depression has often been shown to increase the central itch perception and encourage the subject to react (scratch), we only recruited individuals free of psychiatric diseases and performed tests as the SDS to determine the relationship of personality and depression. We invited volunteers from our research list that have participated in former experiments of our group. They had no psychogenic excoriation and responded to ACH with itch. We tested them with the SDS and accepted scorings below 50. None of them had to be refused from participation. A week later we started our experiments. The injection of ACH affects the cutaneous blood vessels causing vasodilatation measured by the LDF and observed as an initial erythema next to the injection site. In addition, a wheal appears as a direct vasotropic effect of ACH and plasmaextravasation (6, 40, 41). Parallel to the itch perception, the electric impulses antidromically reach the surface of the skin causing a are (efferent part of an axon reex). In contrast to former experiments with NAL (7), our study reveals signicant changes in cutaneous parameters: wheal and are. Similar strong diminution after stimulus control goes along with a less specic interaction of NAL on cutaneous sensations. NAL is known for its rapid absorption after oral intake and a sufcient intracellular and extracellular distribution. In the extensive rst pass of the liver, NAL undergoes extensive glucoronide conjugation. After metabolism blood concentrations are 3.4% for NAL and 73.5% for beta-6-naltrexol. So, it seems likely that NAL itself has only minor effects. Therefore it remains unclear if NAL or other substances cause the unspecic interaction with wheal and are. As mentioned before, ACH injection leads to a triple response (neurogenic inammation) with initial vasodilatation, wheal eruption and a are reaction. It is concluded that mechano-insensitive Cnociceptors and not polymodal C-units mediate the axon reex are in human skin. We used two laser probes at a distance of 1 cm to detect the vasodilatation that goes along with the initial erythema on the skin surface before the are expansion (as sign of the spinally regulated axon reex) starts. Our LDF results revealed no signicant ux differences between the test substances. Neither NAL nor PLA caused an additional increase of ux response, which might support the idea of a peripheral interaction of NAL with the histamine release from mast cells or other vasodilatory agents. In fact, ACH injection leads to a pronounced vasodilatation in each subject. Similar to former studies with AE patients, 63% of the peak value was already noticed within 2 min, with elevated peak values in the proximal laser. This is not an indication for a medication effect but explainable by the overlapping of initial erythema and enhancing are response possibly due to the release of a calcitonin gene-related peptide of the nerval branches stimulated by the axon reex (42). Maximal ux values (P 0.50) and AUC results (P 0.60) were also similar indicating no particular vasotropic interaction of the therapy. Itch intensity decreased from approximately 6 to 5 arbitrary units, yet not signicantly (P 0.50). The maximal itch rating values were similar in all patients indicating that NAL was ineffective in prompt itch relief. This is in accordance with the studies of Metze et al. (22, 23) concerning the clinical relief of itch in AE patients and should be considered for clinical application of NAL in AE. Pruritus is mediated through unmyelinated nociceptive nerve bres to the central nervous system (43, 44). C-bres enter the dorsal horn of the grey matter of the spinal cord, synapse there with secondary neurons that cross over the contralateral spinothalamic tract and ascend to the thalamus (45). From there, tertiary neurons relay itch to the level of conscious perception in the cerebral cortex. According to the gate theory, itch is due to a combination of peripheral excitation and central inhibition. When AE patients feel itch after slight mechanical stimulation of their skin (alloknesis), this is explained by the excitation of central itch transmission neurons due to reduced gating. In four of our 11 participants, the area of alloknesis was completely eliminated. The others showed more than three single alloknesis spots and could be evaluated as an alloknesis area. However, after the 6th min the mean value for them was 53% lower than the PLA value. Obviously NAL interferes strongly with central itch processing. Conclusions 1. NAL diminished the nociception of cholinergic pruritus in every subject, yet not to a point of complete relief. On the other hand the decrease of are size and alloknesis suggests a central inhibition of the axon reex rather than a blockade of nociception in the periphery. 2. The decrease of cutaneous parameters, e.g. 453

Heyer et al. erythema or wheal size outlines the bioavailability of NAL and its metabolites. 3. The dosage of 25 mg is sufcient for experiments. None of our subjects suffered from withdrawal symptoms like nausea, vomiting or other side-effects which we had observed with higher dosages. The often-advised 50 mg should be reserved for long-term clinical management of patients, which are refractory to other therapies.
14. Heyer G, Groene D. Acetylcholine as pruritugenic mediator. 7th Congress of the European Academy of Dermatology Venereology, JEADV 1998: 11: 59. 15. Rukwied R, Heyer G. Administration of acetylcholine and vasoactive intestinal polypeptide to atopic eczema patients. Exp Dermatol 1999: 8: 3945. 16. Nemexin Fachinformation. Du Pont 1999. Alendorf: Bundesverband der Pharmazeutischen Industrie e. V., 1999. 17. Porter S J, Somogyi A A, White J M. Kinetics and inhibition of the formation of 6 beta-naltrexol from naltrexone in human. Br J Clin Pharmacol 2000: 50: 465471. 18. Pauli-Magnus C, Mikus G, Alscher D M et al. Naltrexone does not relieve uraemic pruritus: results of a randomized, double blind, placebo-controlled crossover study. J Am Soc Nephrol 2000: 11: 514519. 19. Wolfhagen F H, Sternieri E, Hop W C, Vitale G, Bertolotti M, Van Buuren H R. Oral naltexone treatment for cholestatic pruritus: a double-blind, placebo-controlled study. Gastroenterology 1997: 113: 12641269. 20. Bergasa N V, Talbot T L, Alling D W. Controlled antipruritic trial of naloxone infusions for the pruritus of chronic cholestasis. Gastroenterology 1992: 102: 544549. 21. Peer G, Kivity S, Agami O et al. Randomised crossover trial of naltrexone in uraemic pruritus. Lancet 1996: 348: 15521554. 22. Metze D, Reimann S, Luger T A. Effective treatment of pruritus with naltrexone, an orally active opiate antagonist. Ann N Y Acad Sci 1999: 20: 430432. 23. Metze D, Reimann S, Beissert S, Luger T. Efcicacy and safety of naltrexone, an oral opiate receptor antagonist, in the treatment of pruritus in internal and dermatological diseases. J Am Acad Dermatol 1999: 41: 533539. 24. Feuerstein T J, Albrecht C, Wessler I, Zentner J, Jackisch R. Delta 1 opioid receptor-mediated control of acetylcholine release in human neocortex slices. Int J Dev Neurosci 1998: 16: 795802. 25. Summereld J A. Naloxone modulates the perception of itch in man. Br J Clin Pharmacol 1980: 10: 180183. 26. Fjellner B, Hgermark . Potentiation of histamine-induced itch and are responses in human skin by the enkephalin analogue FK 33-824, beta-endorphin and morphine. Arch Dermatol Res 1982: 274: 2937. 27. Nissen J B, Kragballe K. Enkephalins modulate differentiation of normal human keratinocytes in vitro. Exp Dermatol 1997: 6: 222229. 28. Grando S A, Kist D A, Qi M, Dahl M V. Human keratinocytes synthesize, secrete and degrade acetylcholine. J Invest Dermatol 1993: 101: 3236. 29. Scott A. Acetylcholine in normal and diseased skin. Br J Dermatol 1962: 74: 317322. 30. Schmelz M, Schmidt R, Bickel A, Handwerker H O, Torebjrk H E. Specic C-receptors for itch in human skin. J Neurosci 1997: 17: 80038008. 31. Bigliardi-Qi M, Mei Bigliardi P L, Eberle A, Rui T. Beta endorphin stimulates cytokeratin 16 expression and downregulates mu-opiate receptor expression in human epidermis. J Invest Dermatol 2000: 114: 527532. 32. Diepgen T L, Fartasch M, Hornstein O P. Evaluation and relevance of atopic basic and minor features in patients with atopic dermatitis and in general population. Acta Derm Venereol 1989: 144: 5054. 33. Hanin J M, Rajka G. Diagnostic features of atopic dermatitis. Acta Derm Venereol (Suppl )(Stockh) 1980: 92: 4447. 34. Zung W W K. A self-rating depression scale. Arch General Psychiat 1965: 12: 6370. 35. Gupta M A, Gupta A K. Depression modulates pruritus

Acknowledgements
This work was supported by the Deutsche Forschungsgesellschaft, SFB 353. We wish to thank Professor Dr med. H. O. Handwerker, PD. Dr med. habil. M. Schmelz and Dr rer. biol. R. Rukwied for their support. Thanks also to R. Mick for his revision of the manuscript.

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