by mapping the spinal cord in the rostro-caudal axis while recording cord dorsum

Transcription

1 J. Physiol. (1984), 357, pp With 8 text-figures Printed in Great Britain DORSAL ROOT POTENTIALS ARE UNCHANGED IN ADULT RATS TREATED AT BIRTH WITH CAPSAICIN BY F. CERVERO AND M. B. PLENDERLEITH From the Department of Physiology, The Medical School, University of Bristol, University Walk, Bristol BS8 1TD (Received 6 June 1984) SUMMARY 1. The possible contribution of non-myelinated afferent fibres (C fibres) to the mechanisms of primary afferent depolarization (p.a.d.) in the spinal cord of the rat has been investigated. 2. Dorsal root potentials (d.r.p.s) were recorded in the lumbar cord of normal adult rats, of adult rats which had been injected at birth with a solution of capsaicin (50 mgkg-' s.c.) and of adult rats which had been injected at birth with the drug vehicle only. 3. D.r.p.s were recorded from the dorsal rootlet that entered the spinal cord in the main area of termination of the tibial nerve. The location of this area was assessed by mapping the spinal cord in the rostro-caudal axis while recording cord dorsum potentials evoked by A-fibre volleys from the tibial nerve. 4. No differences in peak amplitude, area or time to peak amplitude were observed between the d.r.p.s evoked in control and capsaicin-treated rats by stimulation of the tibial, sural or common peroneal nerves. 5. The relation between the size of incoming A volleys to the spinal cord and the size of the d.r.p.s evoked by them was unaffected by the neonatal capsaicin treatment. 6. Rats treated at birth with capsaicin showed a virtual absence of afferent C fibres as assessed from the lack of C waves in the compound action potentials evoked in each of the three nerves studied after antidromic stimulation of the dorsal roots. 7. The presence of p.a.d. in control and in capsaicin-treated animals was also established using the technique of excitability testing of primary afferent fibres (Wall, 1958). No differences were observed between the p.a.d. recorded in control and in capsaicin-treated animals using this technique. 8. D.r.p.s and p.a.d. (assessed by excitability testing of primary afferent fibres) were of similar magnitude in control and in capsaicin-treated rats anaesthetized with either sodium pentobarbitone or urethane. 9. It is concluded that p.a.d. of myelinated afferent fibres produced by incoming volleys in myelinated afferent fibres is not affected by a life-long loss ofnon-myelinated afferent fibres.

2 358 F. CER VERO AND M. B. PLENDERLEITH INTRODUCTION Treatment of neonatal rats with capsaicin (50 mgkg-1) (8-methyl-N-vanillyl- 6-nonenamide) results in the permanent loss of up to 95 % of the non-myelinated (C) afferent fibres of these animals (Jancso, Kiraly & Jancso-Gabor, 1977; Nagy, Vincent, Staines, Fibiger, Reisine & Yamamura, 1980; Nagy, Hunt, Iversen & Emson, 1981; Nagy, Iversen, Goedert, Chapman & Hunt, 1983). In addition, a small but significant reduction in the number of fine myelinated (Ad) afferent fibres has also been reported at capsaicin doses of 50 mgkg-' and higher (Nagy et al. 1983). Morphometric analysis of the dorsal root ganglia (d.r.g.) of rats treated at birth with capsaicin has shown a 70% loss of small dark d.r.g. cells, a cell type usually associated with small afferent fibres (Lawson, 1981). Furthermore, neurochemical studies of the dorsal horn of the spinal cord in such animals have demonstrated a severe reduction in the levels of substance P, somatostatin and fluoride-resistant acid phosphatase (FRAP), all of which are known to be present in the terminals of small-diameter afferent fibres (Jessell, Iversen & Cuello, 1978; Nagy et al. 1980, 1981). The type I synaptic glomeruli in the substantia gelatinosa of the spinal cord, which probably contain the central terminals of afferent C fibres are also reduced by approximately 90 % following neonatal capsaicin treatment (Ribeiro da Silva & Coimbra, 1984). However, nonmyelinated efferent fibres of the autonomic nervous system do not appear to be affected by neonatal capsaicin (Cervero & McRitchie, 1982). Therefore, this treatment results in the permanent loss of a population of afferent fibres including many of those that transmit noxious sensory information from the periphery (Lynn & Carpenter, 1982; Fleischer, Handwerker & Joukhadar, 1983). In the normal animal, afferent C fibres represent a large proportion of the input to those neurones in the dorsal horn of the spinal cord responsible for the integration of nociceptive information (Brown, 1982). In spite of the large decrease in the numbers of afferent C fibres following neonatal capsaicin, the proportions of mechanoreceptive (Class 1), multireceptive (Class 2) and nocireceptive (Class 3) neurones in the dorsal horn and trigeminal nucleus caudalis of these animals appear to be unchanged (Salt, Crozier & Hill, 1982; Cervero, Schouenborg, Sjolund & Waddell, 1984). However, several reports have described changes in the functional organization of the post-synaptic relay in animals treated at birth with capsaicin including increased thresholds to noxious heating of the skin (Salt et al. 1982), reduced ' wind-up' of neurones on peripheral C-fibre stimulation (Cervero et al. 1984), changes in receptive-field properties (Cervero et al. 1984) and alterations of somatotopic maps (Wall, Fitzgerald, Nussbaumer, Van der Loos & Devor, 1982). Presynaptic inhibition of primary afferent fibres may contribute to the control of receptive-field size of second-order neurones (Eccles, Schmidt & Willis, 1963; Schmidt, 1971). This form of inhibition is the result of the activation of an interneuronal pathway generating primary afferent depolarization (p.a.d.) and thus inhibiting presynaptic impulses. It has been proposed that the pathway that mediates p.a.d. includes neurones in the substantia gelatinosa of the spinal cord as well as other neurones in deeper parts of the dorsal horn (Wall, 1962; Cervero, Iggo & Molony, 1978). The substantia gelatinosa is a major area of termination for afferent C fibres and therefore, presynaptic inhibition from myelinated afferent fibre inputs onto

3 NEONATAL CAPSAICIN AND D.R.P.S myelinated afferent fibres could be disrupted by a life-long loss of the C-fibre input to the p.a.d.-generating system. It could also be argued that some of the changes in dorsal horn organization reported in animals treated at birth with capsaicin are the result of alterations in the neuronal network that generates p.a.d. The present study was designed to determine whether or not p.a.d. of myelinated afferent fibres produced by activity in myelinated afferent fibres was altered in rats treated at birth with capsaicin. In these experiments p.a.d. was quantified by measurement of the time course and magnitude of the dorsal root potential (d.r.p.). This potential is attributed to the electrotonic spread of depolarization along primary afferent fibres in the dorsal root and has long been recognized as a simple measurement of the magnitude of p.a.d. (Schmidt, 1971). A preliminary report of this study has been published (Cervero & Plenderleith, 1984). METHODS Neonatal administration of capsaicin. Sprague-Dawley and Wistar rats were injected on their second or third day of life with either capsaicin (50 mgkg-') or with the same volume of the solvent vehicle (10% (v/v) ethanol, 10% (v/v) Tween 80 in normal saline). All injections were carried out under halothane anaesthesia (1 % in 1:3 02 and 2:3 N20). The animals were returned to their mothers after recovering from the effects of the anaesthetic and the immediate effects of the injection. No further procedures were conducted on these animals until the acute electrophysiological experiments. Electrophy8iological experiments. D.r.p.s were recorded from eight capsaicin-treated rats, two vehicle-injected animals and ten untreated controls. In addition, two capsaicin-treated and two control rats were used in experiments in which p.a.d. was assessed using the technique of excitability testing of primary afferent fibres. At the age of 3-5 months the rats were anaesthetized with sodium pentobarbitone (50 mg kg-' I.P.). Further injections of pentobarbitone (10 mg kg-' h-1 i.v.) were given to maintain anaethesia. The trachea, the right femoral artery and the right femoral vein were cannulated. Arterial blood pressure was continuously recorded and the experiments were terminated if the mean blood pressure fell below 75 mmhg. The left common peroneal, tibial and sural nerves were dissected free in the popliteal fossa and the lumbar enlargement of the spinal cord was exposed by a laminectomy. Stimulating electrodes (bipolar silver electrodes) were placed on each of the peripheral nerves and a recording electrode (bipolar silver electrode) was placed more proximally on the sciatic nerve to allow monitoring of the input volley evoked by the stimulation of each of the peripheral nerves. The animals were then paralysed with gallamine and artificially ventilated. The end-tidal C02 was continuously measured and kept at around 3 %. Body temperature was recorded by a rectal probe and maintained at 37 C by a thermostatically controlled heating device. The ipsilateral dorsal surface of the lumbar enlargement of the cord was mapped in the rostro-caudal direction with a monopolar ball-tipped electrode while the tibial nerve was stimulated at a maximal A-volley intensity. The first large negative wave of the recorded cord dorsum potential is the result of electrical fields set up within the cord by the depolarization of post-synaptic neurones activated by the afferent volley. The magnitude of this wave was used as an indicator of the area of termination within the spinal cord of afferent A fibres in the tibial nerve (Handwerker, Iggo & Zimmerman, 1975). The results of such a study in one rat are shown in Fig. 1 (top). The dorsal rootlet entering this area of the cord was sectioned and the central end was placed on a bipolar electrode for the recording of d.r.p.s. An electrode with a fixed interpolar distance of 6 mm was used in all experiments and the proximal pole of the electrode was placed as close as possible to but not touching the spinal cord. Fig.1 (bottom) shows a diagram of the experimental arrangement. D.r.p.s were evoked by maximal A-fibre stimulation. Twenty-five sweeps were averaged using a micro-computer and the average was stored on magnetic disks. Peak amplitude, time to peak and area under the negative d.r.p.s were measured 'off-line' using the same computer. Excitability testing of primary afferent fibres. Two capsaicin-treated and two control rats were used in experiments in which p.a.d. was assessed using the technique of excitability testing of 359

4 360 F. CERVERO AND M. B. PLENDERLEITH primary afferent fibres (Wall, 1958). These four rats were prepared as described above. One control and one capsaicin-treated rat were anaesthetized with sodium pentobarbitone (dose as above) and the other two animals were anaesthetized with urethane (1P25 g kg-' i.p.). A glass-coated tungsten micro-electrode (30-40,sm exposed tip) was inserted into the spinal cord 400 pv 3 ms n. alra Peroneal n. Fig. 1. Top, cord dorsum potentials (negativity downwards) evoked by maximal A-fibre volleys in the tibial nerve. Centre record, potential recorded at the location where the amplitude of the negative wave was maximal. Potentials recorded 2-5 mm rostral (left) and 2-5 mm caudal (right) to the location of maximal amplitude are also shown. Bottom, diagram of the experimental arrangement. See text for details. at a point half-way between the mid line of the cord and the dorsal root entry zone and in the area of termination of the tibial nerve. The location of this area was assessed by recording the cord dorsum potential evoked by a maximal A volley in the tibial nerve. The spinal cord micro-electrode was used to stimulate antidromically primary afferent fibres at 1 Hz (pulse duration 0-1 ms). The intensity of the stimulus was adjusted until the amplitude of the A wave in the antidromic compound action potential recorded in the tibial nerve was half the maximum that could be evoked in this nerve by intraspinal stimulation through the micro-electrode. These stimulus intensities were of the order of sA. Micro-electrode stimulations were applied at depths of 250 and 500,um from the cord surface and at several points within 1 mm rostral and 1 mm caudal to the spot from which the largest cord dorsum potential evoked from the tibial nerve could be recorded. Electrical stimulation of the A fibres in the common peroneal nerve (1 Hz, 01 ms duration) was used as a conditioning stimulus. The intensity of this stimulus was adjusted to evoke a maximal A volley in the nerve. Conditioning stimuli were applied at time of between 0 and 100 ms before the delivery of the test stimulation from the cord micro-electrode. The generation of p.a.d. by these conditioning stimuli was estimated as a percentage increase in the area under the antidromic compound action potential evoked in the tibial nerve by the intraspinal stimulation. Areas were measured off-line from averaged recordings of twenty-five sweeps. At the end of the excitability tests, d.r.p.s were recorded, as described before, from a rootlet entering the cord in the area of termination of the tibial nerve. D.r.p.s were evoked by electrical stimulation of A fibres in the tibial and common peroneal nerves.

5 NEONATAL CAPSAICIN AND D.R.P.S C-fibre lose. The effectiveness of the neonatal capsaicin treatment in causing a loss of afferent C fibres was assessed at the end of each experiment by recording the antidromic C wave of the compound action potential evoked in the hind-limb peripheral nerves by stimulation of the entire L4 or L5 dorsal root at an intensity appropriate for non-myelinated fibres (10-15 V, 0-5 ms pulse width, at 0-33 Hz). The dorsal roots used were cut and their peripheral ends were stimulated. This method avoided the contamination of the evoked C waves with the component due to activity in unaffected efferent C fibres. 361 Control 130 pvl, 30 ms Peroneal n. A Capsaicin Tibial n. A Sural n. A Fig. 2. Dorsal root potentials (average of twenty-five sweeps) evoked by maximal A-fibre volleys in the common peroneal, tibial and sural nerves in control and capsaicin-treated rats. The time of application of the stimulus has been indicated (arrows) in each recording. RESULTS D.r.p.s evoked in the two vehicle-injected rats studied were found to be similar to those evoked in untreated control animals. Therefore data from control and vehicle-injected rats have been pooled in a single control group. As shown in Fig. 2, the d.r.p.s evoked in the area of termination of the tibial nerve by stimulation of the tibial, sural or common peroneal nerves at a maximal intensity for afferent A fibres were qualitatively similar in control and capsaicin-treated animals. D.r.p.s reached maximal amplitude within 25 ms of their onset and declined exponentially over ms. Maximal amplitude. The maximal amplitude of the d.r.p. is the most commonly used indicator of p.a.d. and in this study was calculated as the voltage between the base line before the stimulus was applied and the d.r.p. peak (Fig. 3). No statistically significant difference at the 5 % level was found between the maximal amplitudes of the d.r.p.s evoked by each nerve in control and in capsaicin-treated animals (two-tailed Student's t test; Fig. 3). Area of d.r.p. A more accurate index of the magnitude of p.a.d. is the value of the area under the negative d.r.p. Measurement of the d.r.p. area takes into account the magnitude of the potential with respect to time. D.r.p. areas were calculated using a computer program which calculated the time-voltage integral between the two

6 362 F. CERVERO AND M. B. PLENDERLEITH 500 n =5 n =9 n 5 n=5 400 n=8 0L00 M (U0 Peroneal n. Tibial n. Sural n. Fig. 3. Peak amplitude (inset) of the d.r.p.s evoked by maximal A volleys in the common peroneal, tibial and sural nerves in control (El) and capsaicin-treated rats (U). n indicates number of animals in each group. Data presented as mean peak amplitude with standard errors of the mean. -id~ ~ ~ ~ ~ = > n =9. b25 L'l 0 Peroneal n. Tibialn. Sural n. Fig. 4. Area of the d.r.p.s (inset) evoked by maximal A volleys in the common peroneal, tibial and sural nerves in control (El) and capsaicin-treated rats ( M). indicates n number of animals in each group. Data presented as mean d.r.p. area with standard errors of the mean.

7 NEONATAL CAPSAICIN AND D.R.P.S 363 points in the negative d.r.p. that intersected the base line (Fig. 4). A summary of d.r.p. data for each nerve in the control and capsaicin groups is shown in Fig. 4. No statistically significant differences at the 5 % level were found between the values of d.r.p. areas calculated in the two groups (two-tailed Student's t test). 25 n=9 n=5 = n E E Pernel n Tfl9 n=5 ualn 0 Peroneal n. Tibial n. Sural n. Fig. 5. Time to peak amplitude (inset) of the d.r.p.s evoked by maximal A volleys in the common peroneal, tibial and sural nerves in control (El) and capsaicin-treated (U). n indicates number of animals in each group. Data presented as mean time to peak with standard errors of the mean. D.r.p. time to peak. To confirm that the loss of afferent C fibres had not affected the time course of the p.a.d., the values of time to peak amplitude of the d.r.p.s were measured. These values were calculated as the times between the peak of the small positive wave produced by the arrival of the afferent volley into the cord and the point of maximal amplitude of the negative d.r.p. (Fig. 5). As shown in Fig. 5 no statistically significant differences at the 5 % level were observed between the time to peak values in control and capsaicin-injected animals (two-tailed Student's t test). Relation between the A volley and the d.r.p. Although there were no differences between the d.r.p.s evoked by maximal afferent A volleys in control and capsaicintreated rats, it was conceivable that with submaximal volleys there was an effect due to the lack of C fibres. In order to test this possibility, the relation between the afferent A-volley size and the d.r.p. it produces was first studied in normal rats. Fig. 6A shows the data obtained from three animals when the percentage of the maximal A volley evoked in the tibial nerve by stimuli of increasing intensity was plotted against the percentage of the maximal d.r.p. area evoked by it. Linear-regression analysis of this data (slope = 095, r = 098) indicates a 45 deg linear relation between the size of the afferent A volley and the d.r.p. evoked by it. Fig. 6B shows the data from similar experiments carried out on three capsaicin-injected rats. The linear-regression analysis of this data (slope = 1t03, r = 094) showed a similar 45 deg linear relation

8 364 F. CERVERO AND M. B. PLENDERLEITH between the afferent A volley and the evoked d.r.p. No statistically significant difference at the 5 % level was found between the two regression lines. C-wave data. The extent of afferent C-fibre loss produced by neonatal capsaicin was assessed by recording antidromic compound action potentials in all three nerves studied following electrical stimulation of the L4 or L5 dorsal roots. No differences 100 A x E D ~~~~~~~~~~~~~~ A volley (% of max.) Fig. 6. Relation between the area of the incoming A volley and the area of the resulting d.r.p. in three control rats (A) and three capsaicin-treated animals (B). Lines fitted by linear regression. were observed between the A waves of these compound action potentials in control and capsaicin-treated rats. In contrast, the well-defined C waves evoked in the nerves of control animals were absent or very severely reduced in capsaicin-treated rats (Fig. 7). Therefore, the neonatal treatment had been effective in destroying a large proportion of afferent C fibres in these animals. Excitability tests. P.a.d. was also assessed in control and in capsaicin-treated rats by the technique of excitability testing of primary afferent fibres (Wall, 1958). Fig. 8 shows the results obtained in one control (Fig. 8A) and one capsaicin-treated (Fig. 8B) rat anaesthetized with sodium pentobarbitone. Similar excitability curves were obtained in the two rats. The magnitude of the changes in the excitability of tibial afferent fibres varied in both animals depending on the location of the stimulating micro-electrode in the spinal cord, but the range and variability of the changes in excitability were similar in both rats. Smaller excitability changes were obtained by moving the test electrode within 1 mm of the spot from which the largest excitability change was obtained. The time course of the excitability curves was comparable to the time course of the d.r.p.s recorded in the same animals and evoked by A volleys in the common peroneal nerve (Fig. 8, bottom records). P.a.d. (as assessed by excitability testing) and d.r.p.s were also recorded in one control and one capsaicin-treated rat which had been anaesthetized with urethane. No differences were observed between the excitability curves or between the d.r.p.s recorded in these two animals. No C waves could be recorded in the tibial nerves of the two capsaicin-treated rats used in excitability tests, following electrical stimulation of the L4 or L5 dorsal roots at the appropriate intensity for C-fibre activation.

9 NEONATAL CAPSAICIN AND D.R.P.S 365 Sural n. Tibial n. ~~ Dorsal root ~~~Peroneal n. Control Capsaicin _.4 Peroneal n. I / I.; j Tibial n. I I sj ---- Noft Sural n. 40MVL4 20 ms Fig. 7. Compound action potentials recorded in the common peroneal, tibial and sural nerves of a control rat (left) and of a capsaicin-treated rat (right) and evoked by stimulation of the L4/L5 dorsal root. The diagram shows the experimental arrangement. Note the presence of a conspicuous C wave in the control animal and the absence of such a potential in the capsaicin-treated rat. Arrows indicate the time of application of the stimulus. I , A B O 'U W~~ ~ ~ \1 Time (ms ine(s) 12-20ms1I - Fig. 8. P.a.d. in a control (A) and a capsaicin-treated (B) rat as assessed by the excitability testing technique (top) and by the recording of d.r.p.s (bottom). Each excitability curve was obtained by intraspinal micro-electrode stimulation at a different point within the cord and in the area of termination of the tibial nerve. Conditioning stimuli were applied to the common peroneal nerve ms before the test stimulus. Bottom records are d.r.p.s evoked by maximal A volleys in the common peroneal nerve and recorded from a rootlet entering the area of termination of the tibial nerve.

10 366 F. CERVERO AND M. B. PLENDERLEITH DISCUSSION We have been unable to find any changes in amplitude, time course or latency of d.r.p.s recorded in rats treated at birth with capsaicin. We therefore conclude that a life-long reduction in the number of afferent C fibres does not significantly affect the spinal cord mechanism responsible for the generation of p.a.d. in myelinated afferent fibres. D.r.p.s are an expression of p.a.d., which is generally acknowledged to be an indicator of presynaptic inhibition in the spinal cord. These results suggest that the changes in the functional organization and receptive-field control reported to occur in the spinal cord of the capsaicin-treated rats (Salt et al. 1982; Wall et al. 1982; Cervero et al. 1984) are not due to alterations in presynaptic inhibition. It must be emphasized that d.r.p.s represent the p.a.d. of myelinated afferent fibres only and that p.a.d. in non-myelinated afferent fibres does not contribute to the amplitude and time course of d.r.p.s (Schmidt, 1971). The small value of the length constant of non-myelinated afferent fibres would reduce to very low levels of current any possible antidromic spread of their p.a.d. through the dorsal roots. Hence, our observations based on the recording of d.r.p.s can only apply to the mechanisms of p.a.d. generated on myelinated afferent fibres. Also, care was taken to restrict the afferent volley into the cord to impulses in myelinated fibres. Therefore, the system under study was triggered by impulses in myelinated afferent fibres which generated p.a.d. in myelinated afferent fibres. This system is not altered after neonatal capsaicin since no changes were observed in the relation between the size of the incoming A volley and the size of the evoked d.r.p. It is therefore unlikely that the interneurones in this p.a.d.-generating pathway are, in normal animals, under powerful influences from afferent C fibres. However, it should be noted that p.a.d. in individual fibres is the result of the activation of highly organized systems (Schmidt, Senges & Zimmerman, 1967; Janig, Schmidt & Zimmerman, 1968a, b) whereas the d.r.p.s recorded here were generated by activity in afferent fibres of unknown origin and were recorded from a pool of non-homogeneous dorsal root fibres. Therefore, it is quite possible that a life-long reduction in afferent C fibres could induce subtle changes in the organization of p.a.d., not easily detected by the methods used in our study. Nevertheless, we failed to observe changes in d.r.p.s evoked by nerves of predominantly cutaneous (sural) or mixed (tibial and common peroneal) origin. Also, we have commonly observed dorsal root reflexes in intracellular recordings from identified primary afferent fibres in the dorsal columns of adult rats treated at birth with capsaicin (F. Cervero & M. B. Plenderleith, unpublished observations). The results presented in this paper do not agree with the statement by Wall (1982) that p.a.d. is virtually absent in rats treated at birth with capsaicin. His conclusion was based on observations made on two capsaicin-treated rats using the method of excitability testing of primary afferent fibres. In our study, p.a.d. was assessed by the recording of d.r.p.s complemented with the use of the excitability testing technique in some animals. No change was detectable in either d.r.p.s or in the excitability curves of control and capsaicin-treated animals. However, we noted that the method of excitability testing depends on the accurate positioning of the stimulating electrode within the spinal cord. Smaller excitability changes were obtained by moving this test electrode within 1 mm of the spot from which the largest excitability change was obtained. This variability may account for the conclusion

11 NEONATAL CAPSAICIN AND D.R.P.S reached by Wall (1982) that p.a.d. was virtually absent in his animals. Our study was largely based on the recording of the d.r.p., a phenomenon whose presence indicates p.a.d. and whose variability can easily be assessed. By recording d.r.p.s in several control and capsaicin-treated animals we were able to establish that the variability of the d.r.p. from animal to animal was similar in both groups of rats. Since the mechanisms considered to mediate presynaptic inhibition in the spinal cord appear to function normally after neonatal capsaicin, other systems have to be responsible for the altered functional organization of the spinal cord in capsaicintreated rats. The early part of this study was conducted in the Physiology Department of Edinburgh University. The financial support of the M.R.C. and the excellent technical assistance of Norma Latham are gratefully acknowledged. Professor Bruce Matthews made very helpful comments on earlier versions of the manuscript. M.B.P. was supported by an M.R.C. post-graduate studentship. 367 REFERENCES BROWN, A. G. (1982). The dorsal horn of the spinal cord. Quarterly Journal of Experimental Physiology 67, CERVERO, F., IGGo, A. & MOLONY, V. (1978). The tract of Lissauer and the dorsal root potential. Jouriwal of Physiology 282, CERVERO, F. & McRITcHIE, M. A. (1982). Neonatal capsaicin does not affect unmyelinated efferent fibres of the autonomic nervous system: functional evidence. Brain Research 239, CERVERO, F. & PLENDERLEITH, M. B. (1984). Dorsal root potentials in adult rats treated at birth with capsaicin. Journal of Physiology 346, 23P. CERVERO, F., SCHOUENBORG, J., SJOLUND, B. H. & WADDELL, P. J. (1984). Cutaneous inputs to dorsal horn neurones in adult rats treated at birth with capsaicin. Brain Research 301, ECCLES, J. C., SCHMIDT, R. F. & WILLIS, W. D. (1963). Depolarization of the central terminals of cutaneous afferent fibres. Journal of Neurophysiology 26, FLEISCHER, E., HANDWERKER, H. 0. & JOUKHADAR, S. (1983). Unmyelinated nociceptive units in two skin areas of the rat. Brain Research 267, HANDWERKER, H. O., IGGo, A., ZIMMERMAN, M. (1975). Segmental and supraspinal actions on dorsal horn neurones responding to noxious and non-noxious skin stimuli. Pain 1, JANCSO, G., KIRALY, E. & JANCSO-GABOR, A. (1977). Pharmacologically induced selective degeneration of chemosensitive primary sensory neurones. Nature 270, JANIG, W., SCHMIDT, R. F. & ZIMMERMAN, M. (1968a). Single unit responses and the total afferent outflow from the cat's foot pad upon mechanical stimulation. Experimental Brain Research 6, 10(-115. JANIG, W., SCHMIDT, R. F. & ZIMMERMAN, M. (1968b). Two specific feedback pathways to the central afferent terminals of phasic and tonic mechanoreceptors. Experimental Brain Research 6, JESSELL, T. M., IVERSEN, L. L. & CUELLO, A. C. (1978). Capsaicin-induced depletion of substance P from primary sensory neurones. Brain Research 152, LAWSON, S. N. (1981). Dorsal root ganglion neurones and dorsal roots: effects of neonatal capsaicin. In Spinal Cord Sensation, ed. BROWN, A. G. & RETHELYI, M., pp Edinburgh: Scottish Academic Press. LYNN, B. & CARPENTER, S. E. (1982). Primary afferent units from the hairy skin of the rat hind limb. Brain Research 238, NAGY, J. I., HUNT, S. P., IVERSEN, L. L. & EMSON, P. C. (1981). Biochemical and anatomical observations on the degeneration of peptide-containing primary afferent neurones after neonatal capsaicin. Neuroscience 6, NAGY, J. I., IVERSEN, L. L., GOEDERT, M., CHAPMAN, D. & HUNT, S. P. (1983). Dose-dependent effects of capsaicin on primary sensory neurones in the neonatal rat. Journal of Neuroscience 3,

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