Camilla Ultenius

Cholinergic mechanisms involved in the pain relieving effect of spinal cord stimulation in a model of neuropathy

Abstract The mechanisms underlying the pain relieving effect of spinal cord stimulation (SCS) on neuropathic pain remain unclear. We have previously demonstrated that suppression of tactile hypersensitivity produced by SCS may be potentiated by i.t. clonidine in a rat model of mononeuropathy. Since the analgesic effect of this drug is mediated mainly via cholinergic mechanisms, a study exploring the possible involvement of the spinal cholinergic system in SCS was undertaken. The effect of SCS was assessed with von Frey filaments in rats displaying tactile hypersensitivity after partial ligation of the sciatic nerve and both SCS‐responding and non‐responding as well as normal rats were subjected to microdialysis in the dorsal horn. Acetylcholine (ACh) was analyzed with HPLC before, during and after SCS. SCS produced significantly increased release of ACh in the dorsal horn in rats responding to SCS whereas the release was unaffected in the non‐responding animals. Furthermore, the basal release of ACh was significantly lower in nerve lesioned than in normal rats. In another group of rats it was found that the response to SCS was completely eliminated by i.t. atropine and a muscarinic M4 receptor antagonist while a partial attenuation was produced by M1 and M2 antagonists. Blocking of nicotinic receptors did not influence the SCS effect. In conclusion, the attenuating effect of SCS on pain related behavior is associated with the activation of the cholinergic system in the dorsal horn and mediated via muscarinic receptors, particularly M4, while nicotinic receptors appear not to be involved.

Pain relief by spinal cord stimulation involves serotonergic mechanisms: an experimental study in a rat model of mononeuropathy.

ABSTRACT The aim of the present study was to examine the role of the spinal serotonergic system in the pain relieving effect of spinal cord stimulation (SCS) using a rat model of mononeuropathy. Tactile withdrawal thresholds, cold responses and heat withdrawal latencies were assessed before and after SCS. In some rats, SCS produced an attenuation of the hypersensitivity following nerve injury (SCS responding rats). When SCS was applied immediately prior to sacrifice, the 5‐HT content in the dorsal quadrant of the spinal cord ipsilateral to the nerve injury was increased in SCS responding rats. But there was no change in responding rats without stimulation, or in SCS non‐responding rats with or without stimulation or in controls. Immunohistochemical examination showed a high density of 5‐HT stained terminals in the dorsal horn superficial laminae (I‐II) in SCS responding rats following stimulation. It was also found that i.t. administration of a sub‐effective dose of serotonin in SCS non‐responding rats markedly enhanced the pain relieving effect of SCS on tactile and cold hypersensitivity, while there was no effect on heat hyperalgesia. This enhanced effect on tactile hypersensitivity could be partially blocked by a GABAB receptor antagonist (CGP 35348) but not by a muscarinic M4 receptor antagonist (Muscarinic toxin 3) administered i.t. shortly before the 5‐HT injection. In conclusion, there is evidence that the spinal 5‐HT system plays an important role in the mode of action of SCS involving the activation of descending serotonergic pathways that may inhibit spinal nociceptive processing partially via a GABAergic link.

Spinal GABAergic mechanisms in the effects of spinal cord stimulation in a rodent model of neuropathic pain: is GABA synthesis involved?

The effects of spinal cord stimulation (SCS) on the spinal γ‐amino butyric acid (GABA) system have previously been studied in animal models of neuropathic pain. These studies, confirming the pivotal role of segmental GABA actions for the efficacy of SCS, have led to the question if the disturbance of the GABA inhibitory system as demonstrated both in basal and clinical studies also encompasses malfunction of the GABA synthesis.

396 DOWNREGULATION OF M4 BUT NOT M2 MUSCARINIC RECEPTORS IN DORSAL HORN AFTER PERIPHERAL NERVE INJURY RELATES TO RESPONSIVENESS TO SCS

Tricyclic antidepressants, given systemically, have been widely used for the treatment of various chronic and neuropathic pain conditions in humans for 40 years. Venlafaxine is a novel antidepressant drug that is chemically unrelated to tricyclic or other available antidepressants. Clinicals trials indicate that venlafaxine is effective in chronic pain patients, some of whom are insensitive to other analgesics. Antidepressants induced analgesia was attributed to central actions within spinal cord and at supraspinal sites. However, recently the local peripheral administration of some antidepressants was demonstrated to produce analgesia in the formalin and neuropathic pain models. In the present study, we determined whether venlafaxine could produce peripheral antinociceptive actions in formalin test, a model for acute and tonic pain. For this purpose we treated different groups of male Sprague Dawley rats with venlafaxine locally and systemically. For checking the effect is local or not, we obtained blood at different times after both the local and systemic application to determine the levels of venlafaxine. Venlafaxine induced antinociception at 100, 200 and 400 lg/paw concentrations with local peripheral and at 20 and 40 mg/kg doses with the systemic application. Our results showed that venlafaxine has antinociceptive effect when applied locally to the periphery. By the local application it may be possible to reach high levels at application site and also to get rid of some systemic side effects. Such an activity may led to trials for to use this drug as a gel and cream formulation for analgesia in clinics in the future.

168 SUSCEPTIBILITY TO NERVE INJURY-INDUCED HYPERSENSITIVITY AND RESPONSIVENESS TO SCS IN DA, PVG, AND PVG-RT1AV1 RATS

Background and aims. We have previously demonstrated that gabapentin supraspinally activates the descending noradrenergic system to alleviate neuropathic pain. In this study, we investigated behaviorally and neurochemically whether pregabalin, an anticonvulsant and analgesic drug that is structurally and pharmacologically related to gabapentin, also exhibits similar analgesic effects involving the descending noradrenergic system. Methods. A chronic pain model was prepared by partially ligating the sciatic nerve in mice. The mice received intraperitoneal (i.p.), intracerebroventricular (i.c.v.) or intrathecal (i.t.) injections of pregabalin combined with either central noradrenaline (NA) depletion by 6hydroxydopamine or pharmacological blockade of spinal a2-adrenoceptors. Concentrations of spinal monoamines were also measured using high-performance liquid chromatography in mice after i.c.v. injection of pregabalin. Results. Systemically administered pregabalin (10 and 30 mg/kg, i.p.) reduced mechanical and thermal hypersensitivity in mice only after peripheral nerve injury. Similar analgesic effects were obtained when pregabalin (10 and 30 lg) was injected i.c.v. or i.t. Depletion of spinal NA or blockade of spinal a2-adrenoceptors with yohimbine (1 and 3 lg, i.t.) reduced the analgesic effects of pregabalin (i.p. or i.c.v.). Moreover, i.c.v. administered pregabalin increased the spinal MHPG concentration and the MHPG/NA ratio only in neuropathic pain mice, suggesting that supraspinal pregabalin resulted in an increase in spinal NA turnover. By contrast, the concentrations of NA, serotonin, 5-hydroxyindoleacetic acid and dopamine were unchanged. Conclusions. These results indicate that pregabalin supraspinally activates the descending noradrenergic pain inhibitory system to ameliorate neuropathic pain.

223 REGULATION OF GABAERGIC ACTIVITY IN THE DORSAL HORN AFTER PERIPHERAL NERVE INJURY AND THE EFFECTS OF SPINAL CORD STIMULATION

There is much evidence that the mechanism behind development of pathological pain following peripheral nerve injury is linked to a disturbance of the GABAergic activity in the dorsal horns (DHs). Whether this loss of inhibition is due to a disappearance of GABAergic interneurons or more to down-regulation of the GABA synthesis is presently unclear. Neuropathic pain may effectively be relieved by electric spinal cord stimulation (SCS) but so far the mechanisms mediating this effect are still poorly understood. Earlier studies have shown that SCS induces release of GABA in the DH, suggesting it to be important for suppression of tactile hypersensitivity, observed in rats after peripheral nerve injury. In this study we investigated changes concerning glutamatic acid decarboxylase (GAD), in allodynic and non-allodynic rats. Subsequently, also the effect of SCS on GAD expression was analysed. Our findings suggest that down-regulation of GAD is related to the presence of allodynia and not to the nerve injury per se. Preliminary results also show that allodynic rats responding to SCS with an increased withdrawal threshold to von Frey filaments appear to have an increased immunoreactivity of GAD as compared with non-responders. Our hypothesis is that the loss of inhibitory transmission in the spinal cord following peripheral nerve injury may in part be due to a down-regulation of the GAD enzyme and GABA synthesis rather than death of these interneurons. SCS appears to counteract processes that lead to a reduced inhibitory activity and to re-establish “near normal” conditions.

249 CHOLINERGIC MECHANISMS IN THE ANALGESIC EFFECT OF SPINAL CORD STIMULATION

Spinal cord stimulation (SCS) is an established treatment for chronic neuropathic pain. However, in recent studies conflicting results regarding the effect of SCS were noted in a selected group of patients suffering from Complex Regional Pain Syndrome and mechanical allodynia. In the present study we investigated the pain relieving effect of SCS in a rat experimental model of neuropathic pain as related to the severity of mechanical allodynia. Adult male rats (n = 45) were submitted to a unilateral sciatic nerve ligation. The level of allodynia was tested using the withdrawal response to tactile stimuli with the von Frey test. A portion of these rats developed marked tactile hypersensitivity in the nerve-lesioned paw (von Frey test), similar to “tactile allodynia” observed after nerve injury in humans. Then prior to SCS treatment the rats were subdivided into three groups based on the level of allodynia: mild, moderate and severe. All allodynic rats (n = 27) were treated with SCS for 30min (f = 50Hz; pulse with 0.2ms and stimulation at 2/3 of motor threshold) at 16 days post-injury. Our data demonstrate a differential effect of SCS related to the severity of the mechanical allodynia. SCS leads to a faster and better pain relief in mildly allodynic rats as compared to the more severely allodynic rats. Thus, we suggest that the selection and subdivision of patient groups similar to those defined in our experimental setting (mild, moderate and severe allodynic) may provide better pre-treatment prediction of possible therapeutic benefits of SCS.

247 EFFECT OF SPINAL CORD STIMULATION IN AN ANIMAL MODEL OF NEUROPATHIC PAIN RELATES TO DEGREE OF TACTILE ALLODYNIA

Spinal cord stimulation (SCS) is an established treatment for chronic neuropathic pain. However, in recent studies conflicting results regarding the effect of SCS were noted in a selected group of patients suffering from Complex Regional Pain Syndrome and mechanical allodynia. In the present study we investigated the pain relieving effect of SCS in a rat experimental model of neuropathic pain as related to the severity of mechanical allodynia. Adult male rats (n = 45) were submitted to a unilateral sciatic nerve ligation. The level of allodynia was tested using the withdrawal response to tactile stimuli with the von Frey test. A portion of these rats developed marked tactile hypersensitivity in the nerve-lesioned paw (von Frey test), similar to “tactile allodynia” observed after nerve injury in humans. Then prior to SCS treatment the rats were subdivided into three groups based on the level of allodynia: mild, moderate and severe. All allodynic rats (n = 27) were treated with SCS for 30min (f = 50Hz; pulse with 0.2ms and stimulation at 2/3 of motor threshold) at 16 days post-injury. Our data demonstrate a differential effect of SCS related to the severity of the mechanical allodynia. SCS leads to a faster and better pain relief in mildly allodynic rats as compared to the more severely allodynic rats. Thus, we suggest that the selection and subdivision of patient groups similar to those defined in our experimental setting (mild, moderate and severe allodynic) may provide better pre-treatment prediction of possible therapeutic benefits of SCS.