Spinal Nociceptive Reflex Research Articles

Pituitary adenylate cyclase activating polypeptide expression in sensory neurons

Pituitary adenylate cyclase activating polypeptide (PACAP) is a novel vasoactive intestinal peptide (VIP)-like peptide isolated from ovine hypothalami. The presence of PACAP-like immunoreactivity was recently demonstrated in nerve cell bodies of sensory ganglia in the rat. Since PACAP belongs to a large family of chemically related neuropeptides, we have, in the present study, tried to establish the synthesis of PACAP in neurons of sensory ganglia, using in situ hybridization with a 35S-labelled oligonucleotide probe complementary to PACAP mRNA. The expression of PACAP was compared to that of calcitonin gene-related peptide (CGRP) using a radiolabelled CGRP oligonucleotide probe. The PACAP probe labelled small to medium-sized neurons in the trigeminal ganglion and dorsal root ganglia at different levels, indicating the presence of PACAP mRNA. The CGRP probe labelled nerve cell bodies of varying size, outnumbering those labelled by the PACAP probe. In dorsal root ganglia, cells expressing PACAP constituted c. 10% and those expressing CGRP 46% of the total number of nerve cell bodies. Expression of PACAP was seen in a small subpopulation of cells expressing CGRP. We conclude that PACAP is synthesized in a subpopulation of neurons of sensory anglia in the rat. Therefore, the recently described effects of PACAP—cutaneous vasodilation, potentiation of oedema formation and depression of nociceptive spinal reflexes—may be physiological and related to neurogenic inflammation and modulation of pain transmission.

Inhibition of a somatic nociceptive reflex by gastric distention in humans

Background/Aims: The spinal nociceptive RIII reflex, an exteroceptive cutaneous-muscular flexion reflex, is powerfully and specifically inhibited by painful heterotopic somatic stimuli. The aim of the present study was to evaluate the effects of nonpainful and painful visceral stimuli on this reflex.Methods: In nine healthy volunteers, the effects of five levels of gastric distention were tested on the RIII reflex, recorded from the biceps femoris, and elicited by electrical stimulation of the ipsilateral sural nerve. Distentions were performed by means of a balloon that was placed in the proximal part of the stomach and connected to an electronic barostat. The sensations evoked by gastric distention were scored using a graded (0–6) questionnaire. Results: The 200- and 400-mL distention levels elicited no significant modifications of the RIII reflex; the 600-, 800-, and 1000-mL levels inhibited the RIII reflex by 25%, 35%, and 55%, respectively. The magnitude of this inhibition correlated significantly (P < 0.0001) with both the level of distention and the intensity of visceral perception. Conclusions: Gastric distention produces volume-dependent inhibition of the somatic RIII reflex in humans. This model may provide an interesting tool for objective and quantitative evaluation of normal and disturbed visceral sensations in humans.

Intrathecal excitatory amino acid (EAA) agonists increase tail flick latencies (TFLs) of spinal rats

The facilitation of spinal nociceptive reflexes that occurs after spinal transection reveals the existence of descending, supraspinally mediated inhibition. Substantial evidence indicates that the excitatory amino acids (EAAs) are involved in these spinal circuits. Therefore, it was hypothesized that reflex facilitation in the spinal animal might be due to the removal of inhibitory input normally exerted on the spinal action of EAAs. If so, the facilitatory decrease in reflex latency, observed in the spinal preparation, might be potentiated by intrathecal (IT) administration of EAA agonists. This was tested by comparing the effect of IT injections of N- methyl- D-aspartate (NMDA) and α-amino-3-hydroxy-5-methyl-4-isoxazole-propionic acid (AMPA) on the thermally elicited tail flick (TF) response of Intact and acute spinal rats. In intact rats, a low (0.25 nM) dose of NMDA produced a hyperalgesic decrease in latency, relative to saline, whereas higher doses produced an overall increase in latency. A large dose (0.5 μM) produced overt signs of toxicity (crippling, self-mutilation, and loss of the reflex). Only the highest (1.0 nM) dose of AMPA affected the response, resulting in a significant increase. After spinal transection, the hyperalgesic reaction to 0.25 nM of NMDA was absent, and latencies were significantly increased by 1.0 nM. The toxic reaction to 0.5 μM appeared to be potentiated. Tail flick responses to AMPA were also significantly increased in spinal rats. Contrary to the prediction, reflex latencies were significantly increased by these drugs after spinal transection. It was suggested that, although the spinal action of EAAs appears to be supraspinally modulated, this influence may be facilitatory rather than inhibitory.

Effects of neonatal capsaicin treatment on descending modulation of spinal nociception from the rostral, medial medulla in adult rat

Stimulation-produced modulation from the rostral, medial medulla (RMM) on the spinal nociceptive tail-flick (TF) reflex and on lumbar spinal dorsal horn neuron reponses to noxious cutaneous stimulti was studied in adult rats treated as neonates with capsaicin or vehicle. In vehicle-treated rats ( n = 7), both descending facilitatory and inhibitory influences on the TF reflex were produced from the RMM. At 11/23 sites in the RMM, electrical stimulation produced biphasic modulatory effects. Electrical stimulation facilitated the spinal nociceptive TF reflex at low intensities (5–25 μA) and inhibited the TF reflex at greater intensities (50–200 μA). The mean threshold intensity of stimulation to inhibit the TF reflex (cut-off time= 7.0s) was 66 μA ( n = 11). At 11 of 23 sites, electrical stimulation only inhibited the TF reflex; the mean threshold intensity of stimulation to inhibit the TF reflex was 50 μA ( n = 11). At one stimulation site, electrical stimulation only facilitated the TF reflex at the intensities tested (5–100 μA). In capsaicin-treated rats ( n = 6), the proportion of sites from which electrical stimulation only inhibited the TF reflex was significantly less (3/27sites= 11%) than in vehicle-treated rats (11/23= 48%). The threshold intensity of stimulation to inhibit the TF reflex from these three sites was 50 μA. The number of sites RMM fronm which electrical stimulation only facilitated the TF reflex was significantly greater in capaicin-treated rats (15/27= 56%) than in vehicle-treated rats (1/23= 4%). Neither the number of sites in RMM from which electrical stimulation produced biphasic modulatory effects on the TF reflex (48% and 33%, respectively) nor the intensities of stimulation or magnitudes of facilitation or inhibition of the TF reflex significantly differed between vehicle- and capsaicin-treated rats. In electrophysiological experiments, all units studied responded to non-noxious and noxious intensities of mechanical stimulation applied to the glabrous skin of the plantar surface of the ipsilateral hind foot and also to noxious heating of the skin (50°C). The number of sites where electrical stimulation produced only facilitatory effects on responses of spinal dorsal horn neurons to noxious stimulation (thermal or mechanical) of the skin was significantly increased from 13% of the total sites in vehicle-treated rats to 40% in capsaicin-treated rats. The number of sites where stimulation only produced inhibitory effects on responses of spinal dorsal horn neurons to noxious stimulation was decreased from 47% of the total sites in vehicle-treated rats to 35% in capsaicin-treated rats. The intensity of stimulation for producing inhibitory effects and the magnitude of inhibitory effects produced were not significantly different between vehicle- and capsaicin-treated rats. Immunocytochemical evaluation of the lumbar spinal dorsal horn verified that there were substantial reductions in substance-P and calcitonin gene-related peptide-like immunoreactivity, confirming the effectiveness of neonatal capsaicin treatment. The present study, in support of anatomical studies documenting central effects of capsaicin, suggests that neonatal capsaicin treatment also has significant effects on bulbospinal systems important to the modulation of spinal nociceptive transmission. Specifically, capsaicin treatment of neonates leads to a reduction in inhibitory and an increase in facilitatory influences on spinal nociception descending from the caudal brainstem.

Evidence of a role for [formula omitted] (NMDA) receptors in the facilitation of tail withdrawal after spinal transection

Peripheral injury produces a characteristic excitation of spinal cord dorsal horn cells (wind-up) which is associated with a facilitation of spinal nociceptive reflexes (hyperalgesia). These phenomena are believed to be mediated by a trauma-induced increase in the release of excitatory amino acids (EAAs). A similar increase in the activity of dorsal horn neurons and spinal reflexes occurs after spinal transection. Therefore, the present studies examined the possibility that EAAs, acting through the NMDA receptor, might also be involved in behavioral hyperalgesia produced by central injury. The first experiment assessed the effect of pretreatment with the NMDA antagonist, ketamine, on the facilitated tail flick (TF) response of spinally transected rats. Separate groups of animals were spinalized under isoflurane anesthesia alone, intramuscular ketamine anesthesia alone, or a combination of isoflurance and intrathecal ketamine. The TF was examined 24 h later, before and 30 min after an intrathecal injection of morphine. In the second experiment, the effect of intraperitoneal or intrathecal ketamine on the TF was assessed in separate groups of rats that underwent spinal transection or sham surgery under isoflurane anesthesia. Pretreatment with either systemic or intrathecal ketamine did not alter TF facilitation of morphine-induced antinociception in spinal rats. However, both systemic and intrathecal ketamine significantly increased TF latencies in spinal, relative to intact rats. These results indicate that ketamine did not prevent the development of spinal reflex facilitation, but it selectively reduced this reaction once it was established in spinal rats. The data support an involvement of EAAs in reflex facilitation produced by spinal transection.

Spinal antinociception mediated by a cocaine-sensitive dopaminergic supraspinal mechanism

The role of dopaminergic descending supraspinal processes in mediating the antinociceptive action of cocaine was studied in the rat using a combination of extracellular neuronal recording and behavioral techniques. Neurons in the superficial laminae (I-II) of the spinal dorsal horn with receptive fields on the tail were recorded in anesthetized rats using insulated metal microelectrodes. Stimulation of the receptive field with either high intensity transcutaneous electrical pulses or with an infrared CO 2 laser beam produced a biphasic increase in dorsal horn unit discharge. Conduction velocity estimates indicated that the early discharge corresponded to activity in Aδ whereas the late response corresponded to activity in C afferent fibers. Cumulative doses of cocaine (0.1–3.1 mg/kg i.v.) inhibited the late response to either electrical or laser stimulation in a dose-related manner. The early response to laser, but not electrical, stimulation was also suppressed by cocaine. Neurons in the spinal dorsal horn with receptive fields on the ipsilateral hindpaw were activated by natural noxious (pinch) or innocuous (tap) somatic stimulation. Cocaine selectively suppressed nociceptively evoked dorsal horn unit discharge. This antinociceptive effect was dose-related (0.3–3.1 mg/kg, i.v.) and antagonized by eticlopride (0.05–0.1 mg/kg, i.v.), a selective D2 dopamine receptor blocker. The same doses of cocaine failed to inhibit the responses of dorsal horn neurons to low threshold innocuous stimulation. Complete thoracic spinal cord transection eliminated the antinociceptive effect of cocaine on dorsal horn neurons and also eliminated the cocaine-induced attenuation of the tail-flick reflex. These data demonstrte that cocaine selectively inhibits nociceptive spinal reflexes and the nociceptive responses of dorsal horn neurons primarily by means of a D2 dopaminergic receptor mechanism. This antinociceptive effect of cocaine is independent of its local anesthetic activity and requires the integrity of the thoracic spinal cord, suggesting that the drug potentiates or activates supraspinal dopaminergic projections to the dorsal horn.

Intrathecal oxytocin facilitates the spinal nociceptive flexor reflex in the rat.

We examined the effects of intrathecal oxytocin over a wide dose range (10 ng to 10 micrograms) on the spinal nociceptive flexor reflex in decerebrate, spinalized, unanaesthetized rats. Oxytocin facilitated the flexor reflex at all doses. The duration, but not magnitude, of facilitation was dose-related. No reflex depression was observed with intrathecal oxytocin at any dose. It is suggested that activation of spinal oxytocin receptors excites the spinal cord and therefore intrathecal oxytocin is unlikely to be an analgesic agent.

A selective suppression of human pain sensitivity by carbon dioxide: central mechanisms implicated

The effect of breathing CO2 on somatic sensitivity was studied in human subjects. Healthy humans breathed room air, 100% O2, or CO2 (5% or 8% in O2). Thresholds to heat pain, mechanical pain, electrically evoked pain, and electrically evoked perception were measured using psychophysical techniques. Also, the effects on sensory and affective components of experimental ischaemic pain, alertness and on cardiovascular parameters were observed. In addition, the effect of CO2 on heat pain threshold was determined in the ischaemic limb to exclude the possibility that the threshold elevation was due to an action on primary afferent fibres. Naloxone (0.8 mg), dexamethasone (0.1 mg) or placebo (0.9% NaCl) were applied intravenously in double blind tests to reverse the threshold elevations. In an electrophysiological experiment the effect of CO2 on a spinal nociceptive flexion reflex evoked by an electric stimulus was measured. The CO2 produced a dose-dependent elevation of the heat pain threshold. Similarly, the sensory and affective components of experimental ischaemic pain were attenuated by CO2. The heat pain threshold was significantly elevated by CO2 in the ischaemic limb, also. However, there was no effect on thresholds to mechanically or electrically induced pain or perception thresholds to electrically evoked sensations. One hundred percent O, did not elevate the heat pain threshold. In double blind tests the heat pain threshold elevation was not significant when the naloxone or dexamethasone was administered. The threshold to electrically evoked spinal flexion reflex was not elevated by CO2. The CO2 at the current dose produced sedation, an increase in blood pressure but no change in heart rate.(ABSTRACT TRUNCATED AT 250 WORDS)

Neither cholecystokinin nor galanin modulate intrathecal clonidine-induced depression of the nociceptive flexor reflex in the rat

Previous studies have established that the antinociceptive effect of morphine was subjected to peptidergic modulation at spinal level. Intrathecal (i.t.) galanin (GAL) potentiated morphine-induced analgesia, whereas i.t. cholecystokinin (CCK) antagonized morphine's hypoalgesic effect. In the present study, we examined the possible interaction between GAL, CCK and clonidine, an α 2 adrenoceptor agonist and potent spinal antinociceptive agent, in the spinal nociceptive flexor reflex. I.t. clonidine dose-dependently depressed the flexor reflex similarly to i.t. morphine However, unlike morphine, the reflex depressive effect of i.t. clonidine was neither potentiated by i.t. GAL nor blocked by i.t. CCK. The present results suggested that the analgesia elicited by activation of spinal α 2 adrenoceptors is not subjected to the modulatory effect of CCK and GAL and therefore may be mediated through different mechanisms than opioids.

Functional evidence for multiple receptor activation by κ‐ligands in the inhibition of spinal nociceptive reflexes in the rat

1. The evidence for kappa-receptor heterogeneity is equivocal. We have now investigated this question by comparing the effects of five putatively selective kappa-agonists. The parameters examined were: the relative potencies in depressing hindlimb flexor muscle reflexes to noxious pinch stimuli in both spinalized and sham-spinalized rats; the reversibility of these effects by naloxone; and the effects on blood pressure. 2. Two types of drug effect was discriminated. One drug group, represented by U-50,488, U-69,593 and PD-117,302, had a potency ratio between sham and spinalized rats approximately 10 fold lower than the other group, which comprised GR103545 and CI-977. 3. Under sham-spinalized conditions, CI-977 and GR103545 at high doses caused only sub-maximal reductions of spinal reflexes. U-50,488 was still active when superimposed on these high doses of GR103545. 4. Naloxone reversed all effects, but different doses were required between compounds, with GR103545 taking some 20 times higher doses of naloxone to cause reversal than did U-50,488. 5. The effects on mean arterial pressure were opposite between groups. 6. The results imply that more than one type of naloxone-sensitive non-mu opioid receptor must be involved in mediating these complex actions of ligands that have been claimed to be selective for kappa-receptors.

Effect of RP 67580, a non‐peptide neurokinin1 receptor antagonist, on facilitation of a nociceptive spinal flexion reflex in the rat

1. In order to examine the contribution of neurokinin1 (NK1) receptors to facilitation of a spinal nociceptive reflex in the rat, we have investigated the effects of RP 67580 (3aR, 7aR)-7,7-diphenyl-2(1-imino-2-(2-methoxyphenyl/ethyl)perhydrois oindole)), a non-peptide neurokinin1 (NK1) receptor antagonist, selective for the rodent receptor sub-type, on the activity of individual motorunits. These results were compared with the effects of RP 68651, the inactive 3aS, 7aS enantiomer of RP 67580, as a control for non-specific activity. 2. Experiments were performed on 15 rats anaesthetized with a continuous i.v. infusion of alphaxalone/alphadalone and spinalized at T9-10. Single unit recordings of motorunit activity from biceps femoris/semitendinosus were made with a concentric needle electrode. In each experiment, a vehicle dose followed by 4 sequential rising doses of either RP 67580 or RP 68651 were given at 15 min intervals. High intensity electrical stimuli were applied to the hindlimb receptive field of the motorunit at a rate of 1 per 60 s throughout the experiment to establish a baseline. A conditioning stimulus (20 of these stimuli at 1 Hz) was delivered 5 min after each dose and the effect of the size of the baseline response examined. 3. The conditioning stimulus evoked a facilitation of the baseline at the start of all experiments (mean increase +/- s.e. mean = 151 +/- 20%). RP 67580 attenuated this facilitation, with an ID50 (+/- s.e. mean) of 2.5 +/- 4.2 micrograms kg-1, i.v., whereas RP 68651 at doses of up to 3 mg kg-1, i.v. did not. There was no statistically significant effect of drug on the baseline reflex, nor on the response to the conditioning stimulus. Doses of 300 and 3000 microg kg-1 of both RP 67580 and RP 68651 evoked small depressor effects on systemic arterial blood pressure.4. We conclude that the facilitation of a spinal flexor reflex by noxious conditioning stimuli in the rat is mediated by NK1 receptors whereas the baseline reflex is not. The results suggest that brain penetrantNK1 receptor antagonists may have central anti-nociceptive effects.

Use of non-peptide tachykinin receptor antagonists to substantiate the involvement of NK 1 and NK 2 receptors in a spinal nociceptive reflex in the rat

Non-peptide antagonists of the NK 1 and NK 2 receptors were tested as inhibitors of the reaction time in the rat tail-flick and on the decrease of reaction time induced by the intrathecal injection of the NK 1 receptor selective agonist [Sar 9,Met(O 2) 11]SP or of the NK 2 selective agonist NKA-(4–10). The decrease in reaction time produced by the NK 1 agonist lasted less than 11 min while that evoked by the NK 2 agonist persisted 26 min after injection. When given intrathecally, CP-96,345 and its chloro analog, Cl-CP, blocked dose-dependently both the behavioral responses and the decrease of reaction time induced by 6.5 nmol of [Sar 9,Met(O 2) 11]SP while they failed to modify the hyperalgesic response to 6.5 nmol NKA-(4–10); CP-96,345 was found more potent than Cl-CP and was also active as an antagonist when given intravenously. In contrast, SR 48968 (6.5 and 65 nmol) blocked the NKA-(4–10)-induced decreases in reaction time and was inactive against the hyperalgesic effect of [Sar 9,Met(O 2) 11]SP. The three antagonists blocked in a reversible manner, were inactive on their own on reaction time and non-toxic. The results indicate that the non-peptide CP-96,345 readily crosses the blood brain barrier and acts as a selective antagonist on spinal NK 1 receptors, while SR 48968 is selective on NK 2 receptors in the rat spinal cord. Hence, CP-96,345 and SR 48968 highlight a functional role for NK 1 and NK 2 receptors in spinal sensory neurotransmission.

CP-96,345, but not its stereoisomer, CP-96,344, blocks the nociceptive responses to intrathecally administered substance P and to noxious thermal and chemical stimuli in the rat

The effects of subcutaneous administration of the non-peptide NK-1 (substance P) receptor antagonist, CP-96,345, and its stereoisomer, CP-96,344, were tested in three nociceptive paradigms in the rat. In the first paradigm, tail flick responses were monitored before and after intrathecal administration of substance P (6.5 nmol) in rats pretreated subcutaneously with saline, CP-96,344 (5 mg/kg) or CP-96,345 (5 mg/kg). In the control groups, pretreated with saline ( n = 6) or with CP-96,344 ( n = 5), substance P reduced the tail flick reaction time at 1 min after administration to 38.3 ± 5.1 (mean ±S.E.M.) and 32.1 ± 7.7% of the mean baseline value, respectively. In contrast, in the group pretreated with CP-96,345 ( n = 6) the reaction time following administration of substance P was 98.8 ± 3.3% of the baseline reaction time; this value was not significantly different from the baseline value of this group, indicating a block ( P < 0.01) of the substance P-induced facilitation of the tail flick response. In the second paradigm, rats were anesthetized with a mixture of chloral hydrate (120 mg/kg, i.p.) and sodium pentobarbital (20 mg/kg, i.p.), and the effects were determined on tail flick reaction time of a sustained noxious cutaneous stimulation, immersing the tip of the tail in hot water at 55°C. In the groups of rats pretreated with saline ( n = 4) or with CP-96,344 ( n = 7 ), this noxious stimulus produced a transient decrease in reaction time to 62–74% of the baseline value. However, in the group given CP-96,345 ( n = 6), the reaction time was 97.4 ± 13.7% of the baseline reaction time, again indicating a block of the transient response and the involvement of NK-1 receptors. In the third paradigm, nociceptive scores were determined after subcutaneous injection of 50 μl of 2.5% formalin into one hind paw. The first phase of the response to this noxious chemical stimulus was similar in rats pretreated with saline, with CP-96,344 and with CP-96,345. The second phase was also unaffected in rats pretreated with saline ( n = 10), with 20 mg/kg of CP-96,344 ( n = 8) or with 5 mg/kg of CP-96,345 ( n = 9). However, in the groups given 10 ( n = 8) or 20 ( n = 6) mg/kg of CP-96,345, there was a significant reduction in this second phase ( P < 0.05−0.01). indicating an involvement of NK-1 receptors in evoking this second phase. The results indicate the efficacy of a systemically administered NK-1 receptor antagonist in blocking substance P-induced facilitation of a spinal nociceptive reflex and the stereospecificity of the actions of CP-96,345 in nociceptive tests. The data support the involvement of substance P or a related agent, acting via NK-1 receptors, in nociceptive responses to sustained noxious thermal and noxious chemical stimuli. NK-1 receptor antagonists may, therefore, be useful in the clinical treatment of pain originating from burns or lacerations of the skin or from inflamed tissue.

The depressive effect of intrathecal clonidine on the spinal flexor reflex is enhanced after sciatic nerve section in rats

The effect of intrathecal (i.t.) α 2-adrenoceptor agonist, clonidine, on the spinal nociceptive flexor reflex was studied in decerebrate, spinalized, unanesthetized rats with intact sciatic nerves or in rats in which the sciatic nerve had been ipsilaterally sectioned. In rats with intact nerves i.t. clonidine caused a dose-dependent biphasic effect on flexor reflex excitability. At low dose (10 ng) the effect of clonidine was purely facilitatory, whereas with 50–100 ng clonidine the initial facilitation was often followed by reflex depression. Long-lasting, strong reflex depression was observed after i.t. injection of high doses of clonidine (1 and 10 μg). Four to 18 days after sciatic nerve section, the depressive effect of clonidine on the flexor reflex was dramatically enhanced. Depression was frequently observed already with doses of 5 and 10 ng, and maximal depression was reached at 100 ng and 1 μg in axotomized rats. The facilitatory effect of low doses of clonidine on the reflex was also observed, although somewhat less frequently than in normals. The depressive effect of clonidine on the flexor reflex was reversed by the selective α 2-receptor antagonist, atipamezole (20 μg i.t.), in rats with both intact and sectioned sciatic nerves. The present results revealed an increased sensitivity and effectiveness of the depression of spinal reflex mechanisms by i.t. clonidine after sciatic nerve section, which is opposite to the decreased sensitivity to i.t. morphine after axotomy that we observed previously. Since the flexor reflex is evoked by the activation of afférents that may be the source of neuropathic pain after nerve lesion, the results support the usefulness of i.t. clonidine in treating pain associated with peripheral nerve injury.

Intrathecal CP-96,345 blocks reflex facilitation induced in rats by substance P and C-fiber-conditioning stimulation

We have examined the effects of intrathecally (i.t.) administered CP-96,345, a non-peptide NK 1 receptor ligand, on the spinal nociceptive flexor reflex and on the facilitation of this reflex evoked by i.t. substance P (SP), neurokinin A (NKA) and electrical conditioning stimulation of cutaneous C-afferents. CP-96,345 i.t. at 24 pmol-2.4 nmol had no significant effect on flexor reflex excitability. At the highest dose tested (24 nmol), CP-96,345 caused a brief facilitation of the flexor reflex, which was similar to the effect of the vehicle used at this drug concentration. CP-96,345 did not depress the flexor reflex at any dose. In rats with chronically implanted i.t. catheters, CP-96,345 at 24 nmol caused neither motor impairment nor morphological damage to the spinal cord. Pretreatment with CP-96,345 dose dependently and similarly antagonized facilitation of the flexor reflex induced by 7 pmol i.t. SP or by a 20-s, 1-Hz conditioning stimulus train applied to cutaneous C-fibers in the sural nerve innervation area. The vehicle had no effect. The antagonistic effect of CP-96,345 on the SP- and C-fiber reflex facilitation induced by conditioning stimulation became maximal only 20–30 min after the i.t. injection and lasted 3–4 h at the highest dose. CP-96,345 did not significantly block the facilitatory effect of 7 pmol i.t. NKA on the flexor reflex. These results demonstrate that CP-96,345 is a potent, long-lasting and selective antagonist of SP in rat spinal cord. Furthermore, facilitation of the flexor reflex (central sensitization) induced by conditioning stimulation of cutaneous C-afferents is mediated by NK 1 tachykinin receptors, but the NK 1 receptor may not be involved in the transmission of the flexor reflex. CP-96,345 is thus useful in experimental studies of the role of SP in the central nervous system.

Interactions between substance P and norepinephrine in the regulation of nociception in mouse spinal cord.

This study examined interactions between effects of the undecapeptide substance P and norepinephrine and the alpha-adrenoceptor agonist clonidine in the mouse spinal cord. All compounds were injected into the lumbar subarachnoid space, and effects on the tail-flick reflex and the behavioural response to intrathecal substance P were evaluated. The tail-flick response latencies were markedly increased 5-20 min. after intrathecal application of norepinephrine (0.0125-0.1 microgram) or clonidine (0.0125-0.1 microgram). The actions of both intrathecal norepinephrine (0.025 microgram) and intrathecal clonidine (0.025 microgram) were significantly attenuated when substance P (5 micrograms) was given intrathecally 55 and 45 min. before the agonists. There was a significant relationship between the tail-flick response latencies and the tail skin temperature. However, the tail-flick results were not due to changes in the skin temperature. Intrathecally applied substance P (10 ng) produced a response consisting of biting of the caudal part of the body and a few hindlimb scratches. The number of bites was significantly reduced 5 min. after injection of norepinephrine (0.1 micrograms) or clonidine (0.05-0.1 microgram), but the number of scratches was unchanged. The data show that increased stimulation of spinal alpha-adrenoceptors inhibits a spinal nociceptive reflex as well as the action of substance P in the spinal cord. Substance P modulates the action of alpha-adrenoceptor agonists on the tail-flick reflex, which may tentatively be explained by downregulation of alpha-adrenoceptors by substance P.

Possible startle response contamination of the spinal nociceptive withdrawal reflex

The objective of this study was to examine the possibility that the spinal nociceptive withdrawal reflex, otherwise known as the RIII reflex, is contaminated by the startle response, which is a non-pain-related supraspinal response. Startle response contamination of the RIII reflex would seriously compromise the RIIIs ability to measure spinal nociceptive processes in man, since a change in the startle response affecting EMG amplitude in the RIII latency range would be erroneously interpreted as a change in a spinal nociceptive process. EMG responses evoked by electrical stimulation of the sural nerve were recorded from the orbicularis oculi, neck, biceps, and biceps femoris muscles in 31 healthy human volunteers. The startle response was elicited under conditions often used to record the RIII reflex. Procedures are described that will completely eliminate the startle response. Comparisons between subjects that did and did not elicit a startle response revealed that the startle does not appear to significantly contaminate the biceps femoris RIII reflex, at least when performing group comparisons. There are, however, situations not dealt with in this study in which the startle might significantly contaminate the RIII reflex, such as patients with pre-existing negative emotional states, experimental procedures that induce fear and/or anxiety, and single case studies. It is important, therefore, that investigators using the RIII reflex be cognizant of the startle response and take appropriate precautions to monitor and if necessary eliminate the startle before attributing a change in the RIII reflex to a spinal nociceptive process.

Vagal modulation of nociception

The role of vagal afferents in cardiopulmonary and gastrointestinal function is well-established. An appreciation of their role in the modulation of pain, however, is relatively recent. The authors present studies that investigated the hypothesis that physiological stimuli activate vagal afferents and engage CNS structures integral to the modulation of pain. Electrical stimulation of vagal afferents produces biphasic modulation of spinal nociceptive reflexes and of spinal nociceptive transmission, effects that descend from the brainstem and are mediated by spinal monoamine and opioid receptors. The studies also have revealed that intravenous morphine-produced antinociception and inhibition of spinal nociceptive transmission is mediated in part by vagal afferents. In the aggregate, the studies reviewed document a role for vagal afferents in pain modulation and suggest that these “visceral” afferents are an integral component of endogenous pain control systems.

Intrathecal neurokinin A facilitates the spinal nociceptive flexor reflex evoked by thermal and mechanical stimuli and synergistically interacts with substance P.

The neuropeptide neurokinin A was injected intrathecally and its effect on the spinal nociceptive flexor reflex was examined. The reflex, which was evoked by electrical, thermal or mechanical stimulation of the foot and was recorded from the ipsilateral hamstring muscles, was substantially facilitated by 7 pmol intrathecally injected neurokinin A. The facilitatory effect of neurokinin A to thermal stimulation was, however, significantly stronger than to electrical or mechanical stimuli. Furthermore, co-administration of neurokinin A with substance P induced a significant synergistic facilitation of the reflex. It is suggested that neurokinin A, like substance P, may be released in association with activation of polymodal C-nociceptors.

GABA-mediated inhibition in rostral ventromedial medulla: role in nociceptive modulation in the lightly anesthetized rat

Local microinjection of GABA A receptor agonists and antagonists was used to characterize the role of GABA-mediated inhibitory processes in the nociceptive modulatory functions of the rostral ventromedial medulla (RVM) in the lightly anesthetized rat. Microinjection of selective GABA A receptor antagonists bicuculline methiodide and SR95531 produced a significant increase in tail-flick (TF) latency. This antinociception was dose related, showed recovery and was attenuated by prior injection of the GABA A receptor agonist THIP at the same site. Microinjection of saline or the glycine receptor antagonist strychnine did not significantly affect TF latency. In contrast, administration of GABA A receptor agonists THIP and muscimol resulted in a significant decrease in TF latency. Microinjections at sites surrounding the RVM did not significantly affect TF latency. These results demonstrate that a GABA-mediated process within the RVM is crucial in permitting execution of the TF and, presumably, other spinal nociceptive reflexes.