Zeʼev Seltzer

A novel behavioral model of neuropathic pain disorders produced in rats by partial sciatic nerve injury

&NA; Partial nerve injury is the main cause of causalgiform pain disorders in humans. We present here a novel animal model of this condition. In rats we unilaterally ligated about half of the sciatic nerve high in the thigh. Within a few hours after the operation, and for several months thereafter, the rats developed guarding behavior of the ipsilateral hind paw and licked it often, suggesting the possibility of spontaneous pain. The plantar surface of the foot was evenly hyperesthetic to non‐noxious and noxious stimuli. None of the rats autotomized. There was a sharp decrease in the withdrawal thresholds bilaterally in response to repetitive Von Frey hair stimulation at the plantar side. After a series of such stimuli in the operated side, light touch elicited aversive responses, suggesting allodynia to touch. The withdrawal thresholds to CO2 laser heat pulses were markedly lowered bilaterally. Suprathreshold noxious heat pulses elicited exaggerated responses unilaterally, suggesting thermal hyperalgesia. Pin‐prick evoked such exaggerated responses bilaterally (mechanical hyperalgesia). In a companion report, we show that these abnormalities critically depend on the sympathetic outflow. Based on the immediate onset and long‐lasting perpetuation of similar symptoms, such as touch‐evoked allodynia and hyperalgesia, and the resemblance of the contralateral phenomena to ‘mirror image’ pains in some humans with causalgia, we suggest that this preparation may serve as a model for syndromes of the causalgiform variety that are triggered by partial nerve injury and maintained by sympathetic activity.

Modulation of neuropathic pain behavior in rats by spinal disinhibition and NMDA receptor blockade of injury discharge

&NA; When a peripheral nerve is severed, damaged sensory fibers emit a barrage of impulses that lasts for many seconds, or even several minutes (‘injury discharge’). We have shown in rats that local anesthetic blockade of this discharge suppresses autotomy (a behavioral model of neuropathic pain). Correspondingly, mimicking prolonged injury discharge with electrical stimulation, especially of C‐fibers, increased autotomy. These data support the hypothesis that injury discharge plays a role in the triggering of neuropathic pain. The mechanism of triggering autotomy was investigated using intrathecal injection of agents affecting glutamatergic transmission. A single intrathecal injection at the lumbar enlargement of the NMDA receptor blockers MK‐801 and 5‐APV, just prior to neurectomy, significantly suppressed autotomy. Blocking glycinergic inhibition just prior to neurectomy with a single strychnine injection strikingly enhanced autotomy. Strychnine enhancement of autotomy was prevented by prior injection of MK‐801 or 5‐APV. These results suggest that the expression of autotomy in rats, and by inference neuropathic pain in humans, is affected by injury discharge, possibly mediated by long‐lasting, NMDA receptor‐related, spinal disinhibition.

Effects of sympathectomy in a model of causalgiform pain produced by partial sciatic nerve injury in rats

&NA; In a previous report we presented a novel behavioral model of neuropathic pain disorders, produced in rat by a unilateral ligation of about half of the sciatic nerve. The model is characterized by rapid onset of behaviors suggesting spontaneous pain and disordered responses to non‐noxious and noxious stimuli. These include reduced withdrawal thresholds to repetitive touch in the partially deafferented skin (“touched‐evoked hyperesthesia”), touch‐evoked allodynia, reduced withdrawal thresholds to noxious thermal stimuli and exaggerated responses to noxious heat and mechanical stimuli (‘thermal hyperalgesia’). Some of these disorders are seen at mirror image sites on the hind limb opposite the lesion. These disorders start within hours after partial nerve injury, last many months and are very similar to causalgia in humans following partial nerve injury. Since sympathetic efferent activity is known to aggravate causalgia in humans and sympathectomy is known to relieve it, we studied the effect of changing sympathetic outflow in the rat model. Reversible sympathectomy was carried out using guanethidine injected intraperitoneally in 3 experiments, each at a different time in relation to the partial nerve injury. We found that: (1) sympathectomy performed several months postoperatively alleviated the sensory disorders bilaterally; (2) sympathectomy prior to nerve injury partially prevented the appearance of thermal hyperalgesia but did not affect hyperesthesia to repetitive touch; and (3) sympathectomy at the time of nerve injury aggravated the sensory disorders during the first few days. As maintenance and production of the sensory disorders in this animal model depended on sympathetic nervous outflow, we conclude that the rats were suffering from a syndrome analogous to sympathetically maintained causalgia in man.

The role of injury discharge in the induction of neuropathic pain behavior in rats

&NA; When sensory fibers are damaged, a discharge of impulses is emitted, which can last up to a few minutes. In the present study, we examined whether this injury discharge plays a role in triggering ‘autotomy’ — a behavior involving self‐injury in animals that is induced by total denervation of a hind paw. Sensory input from the sciatic and saphenous neuroma is thought to elicit chronic pain sensations which cause the rat to injure the hind paw. In the present experiments: (1) injury discharge was prevented by using a local anesthetic block (2) injury discharge was artificially prolonged by delivering 150 electrical pulses to the nerve just prior to transection, at a strength sufficient to drive A‐ and C‐fibers. In one group of animals, the nerve was stimulated at 0.5 Hz at which frequency a synchronous, repetitive activity in C‐fibers augments the response of some nociceptive dorsal horn neurons by temporal summation (‘wind‐up’). In 2 other groups, the sciatic nerve was stimulated at 0.1 Hz and 10 Hz. The results show that blocking injury discharge significantly delayed the average time of onset of autotomy and suppressed it in magnitude compared to control rats. In contrast, electrical stimulation, especially at the ‘wind‐up’ frequency, significantly shortened the onset of autotomy and enhanced its severity. Thus, in spite of its short duration, injury discharge affects the subsequent development of neuropathic pain related behavior.

Correlation of intact sensibility and neuropathic pain-related behaviors in eight inbred and outbred rat strains and selection lines

&NA; In some rat strains, total hindpaw denervation triggers autotomy, a behavior of self mutilation presumably related to neuropathic pain. Partial sciatic ligation (PSL) in rats produces tactile allodynia and heat hyperalgesia but not autotomy. Our aims in this study were to examine: (1) whether sensibility of intact rats to noxious and non‐noxious stimuli is strain‐dependent; (2) whether sensibility of intact rats could predict levels of autotomy, or of allodynia and hyperalgesia in the PSL model; and (3) whether autotomy levels are correlated with levels of allodynia or hyperalgesia. Here we report that in two inbred rat strains (Lewis and Fisher 344), two outbred rat strains (Sabra and Sprague–Dawley) and four selection lines of rats (Genetically Epilepsy‐Prone Rats, High Autotomy, Low Autotomy and Flinders Sensitive Line), tactile sensitivity and response duration to noxious heat of intact animals were strain‐dependent. Levels of autotomy following hindpaw denervation and of allodynia and hyperalgesia in the PSL model were also strain‐dependent. Thus, these traits are determined in part by genetic factors. Sensory sensibility of intact rats was not correlated with levels of autotomy following total denervation, or allodynia and hyperalgesia following partial denervation. We suggest that preoperative sensibility of intact rats is not a predictor of levels of neuropathic disorders following nerve injury. Likewise, no correlation was found between autotomy, allodynia and hyperalgesia, suggesting that neuropathic pain behaviors triggered by nerve injury of different etiologies are mediated by differing mechanisms.

Mapping a gene for neuropathic pain-related behavior following peripheral neurectomy in the mouse

&NA; Total hindpaw denervation in rodents elicits an abnormal behavior of licking, scratching and self‐injury of the anesthetic limb (‘autotomy’). Since the same denervation produces phantom limb pain and anesthesia dolorosa in humans, autotomy has been used as a model of human neuropathic pain. Autotomy is an inherited trait in rodents, attributable to a few genes of major effect. Here we used recombinant inbred (RI) mouse lines of the AXB‐BXA RI set to map a gene for autotomy. Autotomy levels following unilateral sciatic and saphenous nerve section were scored daily for 36 days, using a standardized scale, in all 23 RI lines available for this set. We used a genetic map of 395 marker loci and a permutation‐based statistical method for categorical data to assess the statistical significance of mapping results. We identified a marker on chromosome 15 with statistical support (P=0.0003) in the range considered significant for genome‐wide scans in the mouse. Several genes located in this chromosomal region encode for neural functions related to neuropathic pain and may indicate targets for development of novel analgesics.

Comparison of autotomy behavior induced in rats by various clinically-used neurectomy methods

&NA; When a peripheral nerve is cut, a neuroma develops at its proximal end. Nerve‐end neuromas are known to be a source of ectopic sensory input. In some humans this input may cause spontaneous and evoked neuropathic pain. There is currently no available animal model for developing better methods of cutting nerves that produce less painful neuromas than those currently in clinical use. Transection of the sciatic and saphenous nerves in rats also produces nerve‐end neuromas. Afferent fibers in such neuromas spontaneously emit ectopic input that coincides with the outbreak of licking, scratching and self‐mutilation of the denervated limb (‘autotomy’). This behavior is considered to be the expression of spontaneous disagreeable sensations such as paresthesias, dysesthesias or neuropathic pain. We propose here that the autotomy model can be used as the first step for development of better neurectomy methods. As a demonstration, in this report we compared the course of autotomy expressed by rats following several methods of cutting peripheral nerves that are currently in clinical use. We found that the lowest extent of autotomy was caused by sciatic and saphenous neurectomy with a CO2 laser. Tight ligation of the nerve, or a simple cut with scissors, also yielded significantly lower autotomy scores compared to cryoneurolysis and electrocut. The differing scores of autotomy caused by these neurectomy methods may derive from different properties of the injury discharge produced by these methods at the time of nerve cut. Our results raise the possibility that a higher incidence of neuropathic pain or related sensory disorders in humans may be expected following cryosurgical and electrocut neurectomies. If validated by further studies, neurectomy methods eliciting lower incidence of autotomy, and sensory disorders in models not based on autotomy may produce lower levels of neuropathic pain in humans.

Suppression of neuropathic pain behavior in rats by a non-psychotropic synthetic cannabinoid with NMDA receptor-blocking properties

&NA; HU211 is a novel synthetic derivative of tetrahydro‐cannabinol (THC), the active marijuana ingredient. The stereochemistry of HU211 is enantiomeric to that of THC. In contrast to THC, HU211 is not psychotropic. This agent exhibits other types of biological activities; it is a non‐competitive NMDA receptor blocker and has antinociceptive activity when injected with cupric chloride. This study examined its effects in autotomy, a behavioral model of neuropathic pain. Autotomy, a behavior of self‐mutilation of denervated areas, was induced in Sabra rats by cutting the sciatic and saphenous nerves. We found that injections of HU211 (2.5 mg/kg) with cupric chloride (0.8 mg/kg) every 2nd day markedly suppressed autotomy during the injection period by delaying its average onset day and reducing the incidence of severe autotomy. Moreover, suppression of autotomy was retained in the postinjection period (for at least 30 days) but only when the drug was injected intraperitoneally. Lesser effects were achieved by subcutaneous injections. Cupric chloride or HU211 alone were ineffective. The general behavior and open field motor activity indicated that the effects of HU211 with Cu++ on autotomy were not due to sedation or ataxia but presumably due to antinociception mediated by NMDA receptor blockade.

Dietary supplementation with the inhibitory amino acid taurine suppresses autotomy in HA rats.

TAURINE is an inhibitory amino acid in the CNS. When supplied to rats it produces analgesia in some acute pain tests. Here we examined the effect of taurine supplementation on sensitivity to pain in intact rats, and whether perioperative dietary supplementation with taurine in rats would suppress autotomy, a behavior produced by peripheral neurectomy and related to neuropathic pain. Thermal pain sensitivity of intact rats consuming 1% taurine in the drinking solution for 2 weeks was not significantly different from that of control rats. Autotomy levels, determined in rats consuming taurine pre-, post- or perioperatively were significantly lower than in matching control groups. We conclude that taurine plays an important role in the autotomy model, presumably by protecting inhibitory neurons in the CNS against an excitotoxic damage triggered by injury discharge and ectopic input from the severed nerves.

Inherited propensity for neuropathic pain is mediated by sensitivity to injury discharge

We reported previously that injury discharge (ID), a burst of impulses fired following nerve injury, plays a role in triggering autotomy, a neuropathic pain-related behavior in rats. Here we affirm this link using two lines of rats, derived by selective breeding from the Sabra strain to express high (HA) or low (LA) levels of autotomy following hindpaw denervation. Blocking ID in HA rats before injury suppressed autotomy. Correspondingly, artificial prolongation of ID in LA rats just prior to neurectomy, increased autotomy. The autotomy in these HA and LA rats was like that of their Sabra ancestors. This suggests that the underlying basis for selection of these lines was differential sensitivity of the CNS to the effects of ID.