Neuronal Hyperexcitability: A Substrate for Central Neuropathic Pain After Spinal Cord Injury

Neuronal hyperexcitability produces enhanced pain transmission in the spinal dorsal horn after spinal cord injury (SCI). Spontaneous and evoked neuronal excitability normally are well controlled by neural circuits. However, SCI produces maladaptive synaptic circuits in the spinal dorsal horn that result in neuronal hyperexcitability. After SCI, activated primary afferent neurons produce enhanced release of glutamate, neuropeptides, adenosine triphosphate, and proinflammatory cytokines, which are known to be major components for pain transmission in the spinal dorsal horn. Enhanced neurochemical events contribute to neuronal hyperexcitability, and neuroanatomical changes also contribute to maladaptive synaptic circuits and neuronal hyperexcitability. These neurochemical and neuroanatomical changes produce enhanced cellular signaling cascades that ensure persistently enhanced pain transmission. This review describes altered neurochemical and neuroanatomical contributions on neuronal hyperexcitability in the spinal dorsal horn, which serve as substrates for central neuropathic pain after SCI.     

Unilateral Focal Burn Injury Is Followed by Long-Lasting Bilateral Allodynia and Neuronal Hyperexcitability in Spinal Cord Dorsal Horn

Pain after burn injury can be intense and long lasting. Treatment is often ineffective, and there is a need for increased knowledge of the underlying pain mechanisms. In the present study, we established a unilateral partial-thickness burn injury model, which produces ipsilateral mechanical allodynia soon after injury, followed by contralateral allodynia. Chronic bilateral allodynia lasts up to 8 weeks postinjury in this model. In addition to the change in pain behavior, electrophysiological analyses showed that dorsal horn neurons become hyperexcitable and display significantly increased evoked activity with enlarged receptive fields, initially on the side ipsilateral to the injury, and subsequently on both sides of the spinal cord. It is known that, following nerve injury, activation of p38 mitogen-activated protein kinase (MAPK) pathways within spinal microglia contributes to the pathogenesis of pain. In our burn injury model, rapid and prolonged activation of phospho-p38-expressing microglia occurs bilaterally in the spinal cord dorsal horn. Taken together, these data demonstrate that a unilateral peripheral burn injury can produce long-lasting allodynia that can spread to the contralateral limb, together with dorsal horn neuronal hyperexcitability and microglial activation on both ipsilateral and contralateral sides of the spinal cord. Our results suggest that central neuropathic mechanisms can contribute to pain after burn injury.PerspectiveMechanisms contributing to pain following burn injury are incompletely understood. In a novel animal model of burn injury, we have demonstrated hyperexcitability of second-order sensory neurons, activation of microglia, and chronic bilateral pain following the burn injury. This work identifies potential therapeutic targets to alleviate pain after burn injury.     

Minocycline Attenuates Mechanical Allodynia and Central Sensitization Following Peripheral Second-Degree Burn Injury

Burn injury induces severe pain that can be refractory to existing pharmacotherapies. The underlying mechanism of burn pain remains unclear. We previously established an animal model and reported that unilateral burn injury induces chronic and bilateral mechanical allodynia, which is associated with central sensitization and microglial activation in the spinal cord dorsal horn. Modulation of the activity of microglia and p38 mitogen-activated protein kinase (MAPK) has been shown to ameliorate neuropathic pain in several nerve-injury pain models. In the present study, we show in this rat model that daily treatment with the microglial inhibitor minocycline (10 mg/kg), administered at the time of burn injury and for 7 days thereafter, significantly attenuates ipsilateral and contralateral allodynia as assessed up to 1 month following burn injury. These sensory changes are paralleled by significant suppression of evoked hyperexcitability of dorsal-horn neurons and of the expression of phosphorylated p38 (phospho-p38) in OX42+ microglial cells within the dorsal horn. Our results suggest that modulation of inflammation at early times after burn injury may have long-lasting effects, attenuating central neuropathic mechanisms which contribute to pain after burn injury.     

A role for peripheral afferents in the pathophysiology and treatment of at-level neuropathic pain in spinal cord injury? A case report

At-level neuropathic pain is a frequent symptom following spinal cord injury, but the underlying pathophysiology is not completely understood. We report a patient suffering from treatment-resistant at-level pain characterized by ongoing pain and mechanical allodynia for three years after an incomplete spinal lesion. Quantitative sensory testing revealed severe thermosensory deficits in the neuropathic pain area. However, topical application of capsaicin in the neuropathic pain area induced a burning pain sensation, a marked decrease in heat pain threshold and an increase in mechanical allodynia. Treatment with topical lidocaine patches (5%) led to considerable pain relief. These results indicate a functional connection between peripheral, spinal and supraspinal nociceptive pathways and that peripheral afferents may contribute to at-level neuropathic pain after spinal cord injury in this patient. Lesioned peripheral afferents in combination with central neuronal hyperexcitability are discussed as a likely underlying pain mechanism.     

Distinct roles of group III metabotropic glutamate receptors in control of nociception and dorsal horn neurons in normal and nerve-injured Rats

Increased glutamatergic input to spinal dorsal horn neurons constitutes an important mechanism for neuropathic pain. However, the role of group III metabotropic glutamate receptors (mGluRs) in regulation of nociception and dorsal horn neurons in normal and neuropathic pain conditions is not fully known. In this study, we determined the effect of the group III mGluR specific agonist L(+)-2-amino-4-phosphonobutyric acid (L-AP4) on nociception and dorsal horn projection neurons in normal rats and a rat model of neuropathic pain. Tactile allodynia was induced by ligation of L5/L6 left spinal nerves in rats. Allodynia was determined by von Frey filaments in nerve-injured rats. The nociceptive threshold was tested using a radiant heat and a Randall-Selitto pressure device in normal rats. Single-unit activity of ascending dorsal horn neurons was recorded from the lumbar spinal cord in anesthetized rats. An intrathecal (5-30 microg) L-AP4 dose-dependently attenuated allodynia in nerve-injured rats but had no antinociceptive effect in normal rats. Topical spinal application of 5 to 50 microM L-AP4 also significantly inhibited the evoked responses of ascending dorsal horn neurons in nerve-ligated but not normal rats. Furthermore, blockade of spinal group III mGluRs significantly decreased the withdrawal threshold and increased the evoked responses of dorsal horn neurons in normal but not nerve-injured rats. These data suggest that group III mGluRs play distinct roles in regulation of nociception and dorsal horn neurons in normal and neuropathic pain states. Activation of spinal group III mGluRs suppresses allodynia and inhibits the hypersensitivity of dorsal horn projection neurons associated with neuropathic pain.     

Functional Characterization of At-Level Hypersensitivity in Patients With Spinal Cord Injury

At-level and above-level hypersensitivity was assessed in patients with chronic complete thoracic spinal cord injury (SCI). Patients were classified using somatosensory mapping (brush, cold, pinprick) and assigned into 2 groups (ie, patients with at-level hypersensitivity [SCIHs, n = 8] and without at-level hypersensitivity [SCINHs, n = 7]). Gender and age-matched healthy subjects served as controls. Quantitative sensory testing (QST), electrically- and histamine-induced pain and itch, laser Doppler imaging, and laser-evoked potentials (LEP) were recorded at-level and above-level in SCI-patients. Six of 8 SCIHs, but 0 of 7 SCINHs patients suffered from neuropathic below-level pain. Clinical sensory mapping revealed spreading of hypersensitivity to more cranial areas (above-level) in 3 SCIHs. Cold pain threshold measures confirmed clinical hypersensitivity at-level in SCIHs. At-level and above-level hypersensitivity to electrical stimulation did not differ significantly between SCIHs and SCINHs. Mechanical allodynia, cold, and pin-prick hypersensitivity did not relate to impaired sensory function (QST), axon reflex flare, or LEPs. Clinically assessed at-level hypersensitivity was linked to below-level neuropathic pain, suggesting neuronal hyperexcitability contributes to the development of neuropathic pain. However, electrically evoked pain was not significantly different between SCI patients. Thus, SCI-induced enhanced excitability of nociceptive processing does not necessarily lead to neuropathic pain. QST and LEP revealed no crucial role of deafferentation for hypersensitivity development after SCI.

Injury discharges regulate calcium channel alpha-2-delta-1 subunit upregulation in the dorsal horn that contributes to initiation of neuropathic pain

Previous studies have shown that peripheral nerve injury in rats induces increased expression of the voltage gated calcium channel (VGCC) alpha-2-delta-1 subunit (Ca v alpha2 delta1) in spinal dorsal horn and sensory neurons in dorsal root ganglia (DRG) that correlates to established neuropathic pain states. To determine if injury discharges trigger Ca v alpha2 delta1 induction that contributes to neuropathic pain initiation, we examined allodynia onset and Ca v alpha2 delta1 levels in DRG and spinal dorsal horn of spinal nerve ligated rats after blocking injury induced neural activity with a local brief application of lidocaine on spinal nerves before the ligation. The lidocaine pretreatment blocked ligation-induced discharges in a dose-dependent manner. Similar pretreatment with the effective concentration of lidocaine diminished injury-induced increases of the Ca v alpha2 delta1 in DRG and abolished that in spinal dorsal horn specifically, and resulted in a delayed onset of tactile allodynia post-injury. Both dorsal horn Ca v alpha2 delta1 upregulation and tactile allodynia in the lidocaine pretreated rats returned to levels similar to that in saline pretreated controls 2 weeks post the ligation injury. In addition, preemptive intrathecal Ca v alpha2 delta1 antisense treatments blocked concurrently injury-induced allodynia onset and Ca v alpha2 delta1 upregulation in dorsal spinal cord. These findings indicate that injury induced discharges regulate Ca v alpha2 delta1 expression in the spinal dorsal horn that is critical for neuropathic allodynia initiation. Thus, preemptive blockade of injury-induced neural activity or Ca v alpha2 delta1 upregulation may be a beneficial option in neuropathic pain management.     

The effect of electroacupuncture on pain behaviors and noxious stimulus-evoked Fos expression in a rat model of neuropathic pain.

Chronic-constriction injury (CCI) of the sciatic nerve causes mechanical and heat hyperalgesia and mechanical allodynia in the plantar surface of the hindpaw. The underlying mechanism thought to account for these phenomena include central sensitization induced by peripheral nerve injury, ie, the increase in neuronal activity of spinal dorsal horn neurons. As a marker of neuronal activation of the central nervous system, Fos expression has been used widely to monitor the change in neuronal activity evoked by peripheral input. In this study, we examined the antinociceptive effect of electroacupuncture (EA) on pain behavior and noxious stimulus-evoked Fos expression in dorsal horn neurons of the spinal cord in CCI rats 14 days after injury. Male Sprague-Dawley rats (180 to 200 g) received loose ligation of the left sciatic nerve. Heat and mechanical hyperalgesia and mechanical allodynia were examined by the plantar foot test, the pin-prick test, and the von Frey test before and after the EA treatment (100 Hz, 0.3 millisecond, 3 or 1 mA, 20 minutes) into the Zusanli point (S36). When EA stimulation to the Zusanli point was applied, the mechanical and heat hyperalgesia were significantly suppressed; however, mechanical allodynia was not affected. The EA stimulation to nonacupuncture point did not show any significant effect. Next, pinch stimulation was applied to the plantar surface of the operated hindpaw of the CCI rats for 10 minutes, and the stimulus-evoked Fos expression in dorsal horn neurons in L4-L6 spinal cord levels was then examined by using immunohistochemistry. The number of noxious stimulus-evoked Fos-labeled neurons in both the superficial and deep laminae of the dorsal horn in the CCI rats was increased significantly compared with those in sham-operated rats, suggesting an increased excitability of dorsal horn neurons to noxious stimuli. Concurrent EA treatment to the Zusanli point with the pinch stimulus suppressed the increase in the number of Fos-labeled cells in the spinal dorsal horn in the CCI rats. The present results show that EA treatment has antinociceptive effects on both pain behavior and neuronal activation of the spinal dorsal horn neurons in CCI rats.     

Enhancement of NMDA receptor phosphorylation of the spinal dorsal horn and nucleus gracilis neurons in neuropathic rats

NR1 is an essential component of functional NMDA receptors and can be activated by phosphorylation. It is suggested that phosphorylation of NR1 (pNR1) contributes to central sensitization after intradermal capsaicin injection. The present study investigates whether increases of spinal pNR1 are correlated to central sensitization and thus pain behaviors in neuropathic pain. Neuropathic rats were produced by L5 spinal nerve ligation, mechanical thresholds of the paw were measured, and then the L4/5 spinal cords and the nucleus gracilis (NG) were removed and immunostained for pNR1. The results showed that the number of pNR1-immunoreactive neurons was significantly increased in the ipsilateral cord, at 3, 7, and 28 days after nerve ligation and these increases coincide with mechanical allodynia. The increase of pNR1-immunoreactive neurons in the NG was observed only at 28 days after the nerve ligation. Western blot analyses confirmed the significant increase of pNR1 protein in spinal dorsal horn after nerve ligation. A protein kinase A inhibitor, H89, moderately reversed mechanical allodynia in 7 day neuropathic rats. Many pNR1-immunoreactive neurons were identified as projection neurons by retrograde tracer. The data suggest that PKA mediated NMDA receptor phosphorylation plays an important role in spinal nerve ligation induced neuropathic pain.     

Calcium channel alpha-2-delta-1 protein upregulation in dorsal spinal cord mediates spinal cord injury-induced neuropathic pain states

Spinal cord injury (SCI) commonly results in the development of neuropathic pain, which can dramatically impair the quality of life for SCI patients. SCI-induced neuropathic pain can be manifested as both tactile allodynia (a painful sensation to a non-noxious stimulus) and hyperalgesia (an enhanced sensation to a painful stimulus). The mechanisms underlying these pain states are poorly understood. Clinical studies have shown that gabapentin, a drug that binds to the voltage-gated calcium channel alpha-2-delta-1 subunit (Ca_vα2δ-1) proteins is effective in the management of SCI-induced neuropathic pain. Accordingly, we hypothesized that tactile allodynia post SCI is mediated by an upregulation of Ca_vα2δ-1 in dorsal spinal cord. To test this hypothesis, we examined whether SCI-induced dysregulation of spinal Ca_vα2δ-1 plays a contributory role in below-level allodynia development in a rat spinal T9 contusion injury model. We found that Ca_vα2δ-1 expression levels were significantly increased in L4-6 dorsal, but not ventral, spinal cord of SCI rats that correlated with tactile allodynia development in the hind paw plantar surface. Furthermore, both intrathecal gabapentin treatment and blocking SCI-induced Ca_vα2δ-1 protein upregulation by intrathecal Ca_vα2δ-1 antisense oligodeoxynucleotides could reverse tactile allodynia in SCI rats. These findings support that SCI-induced Ca_vα2δ-1 upregulation in spinal dorsal horn is a key component in mediating below-level neuropathic pain states, and selectively targeting this pathway may provide effective pain relief for SCI patients.     

Pannexin 1: A novel participant in neuropathic pain signaling in the rat spinal cord

Pannexin 1 (panx1) is a large-pore membrane channel expressed in many tissues of mammals, including neurons and glial cells. Panx1 channels are highly permeable to calcium and adenosine triphosphatase (ATP); on the other hand, they can be opened by ATP and glutamate, two crucial molecules for acute and chronic pain signaling in the spinal cord dorsal horn, thus suggesting that panx1 could be a key component for the generation of central sensitization during persistent pain. In this study, we examined the effect of three panx1 blockers, namely, 10panx peptide, carbenoxolone, and probenecid, on C-reflex wind-up activity and mechanical nociceptive behavior in a spared nerve injury neuropathic rat model involving sural nerve transection. In addition, the expression of panx1 protein in the dorsal horn of the ipsilateral lumbar spinal cord was measured in sural nerve–transected and sham-operated control rats. Sural nerve transection resulted in a lower threshold for C-reflex activation by electric stimulation of the injured hindpaw, together with persistent mechanical hypersensitivity to pressure stimuli applied to the paw. Intrathecal administration of the panx1 blockers significantly depressed the spinal C-reflex wind-up activity in both neuropathic and sham control rats, and decreased mechanical hyperalgesia in neuropathic rats without affecting the nociceptive threshold in sham animals. Western blotting showed that panx1 was similarly expressed in the dorsal horn of lumbar spinal cord from neuropathic and sham rats. The present results constitute the first evidence that panx1 channels play a significant role in the mechanisms underlying central sensitization in neuropathic pain.     

Calcium channel alpha2delta1 subunit mediates spinal hyperexcitability in pain modulation

Mechanisms of chronic pain, including neuropathic pain, are poorly understood. Upregulation of voltage-gated calcium channel (VGCC) alpha2delta1 subunit (Ca(v)alpha2delta1) in sensory neurons and dorsal spinal cord by peripheral nerve injury has been suggested to contribute to neuropathic pain. To investigate the mechanisms without the influence of other injury factors, we have created transgenic mice that constitutively overexpress Ca(v)alpha2delta1 in neuronal tissues. Ca(v)alpha2delta1 overexpression resulted in enhanced currents, altered kinetics and voltage-dependence of VGCC activation in sensory neurons; exaggerated and prolonged dorsal horn neuronal responses to mechanical and thermal stimulations at the periphery; and pain behaviors. However, the transgenic mice showed normal dorsal horn neuronal responses to windup stimulation, and behavioral responses to tissue-injury/inflammatory stimuli. The pain behaviors in the transgenic mice had a pharmacological profile suggesting a selective contribution of elevated Ca(v)alpha2delta1 to the abnormal sensations, at least at the spinal cord level. In addition, gabapentin blocked VGCC currents concentration-dependently in transgenic, but not wild-type, sensory neurons. Thus, elevated neuronal Ca(v)alpha2delta1 contributes to specific pain states through a mechanism mediated at least partially by enhanced VGCC activity in sensory neurons and hyperexcitability in dorsal horn neurons in response to peripheral stimulation. Modulation of enhanced VGCC activity by gabapentin may underlie at least partially its antihyperalgesic actions.     

Peripheral axonal injury results in reduced mu opioid receptor pre- and post-synaptic action in the spinal cord

In both the spared nerve injury (SNI) and spinal nerve ligation (SNL) rat peripheral neuropathic pain models the presynaptic inhibitory effect of the mu opioid receptor (MOR) agonist (DAMGO) on primary afferent-evoked excitatory postsynaptic currents (EPSCs) and miniature EPSCs in superficial dorsal horn neurons is substantially reduced, but only in those spinal cord segments innervated by injured primary afferents. The two nerve injury models also reduce the postsynaptic potassium channel opening action of DAMGO on lamina II spinal cord neurons, but again only in segments receiving injured afferent input. The inhibitory action of DAMGO on ERK (extracellular signal-regulated kinase) activation in dorsal horn neurons is also reduced in affected segments following nerve injury. MOR expression decreases substantially in injured dorsal root ganglion neurons (DRG), while intact neighboring DRGs are unaffected. Decreased activation of MOR on injured primary afferent central terminals and the second order neurons they innervate may minimize any reduction by opioids of the spontaneous pain mediated by ectopic input from axotomized small diameter afferents. Retention of MOR expression and activity in nearby non-injured afferents will enable, however, an opioid-mediated reduction of stimulus-evoked and spontaneous pain carried by intact nociceptor afferents and we find that intrathecal DAMGO (1000 ng) reduces mechanical hypersensitivity in rats with SNL. Axotomy-induced changes in MOR may contribute to opioid- insensitive components of neuropathic pain while the absence of these changes in intact afferents may contribute to the opioid sensitive components.     

Perturbing PSD-95 Interactions With NR2B-subtype Receptors Attenuates Spinal Nociceptive Plasticity and Neuropathic Pain

Peripheral inflammation or nerve injury induces a primary afferent barrage into the spinal cord, which can cause N-methyl -aspartate (NMDA) receptor-dependent alterations in the responses of dorsal horn sensory neurons to subsequent afferent inputs. This plasticity, such as “wind-up” and central sensitization, contributes to the hyperexcitability of dorsal horn neurons and increased pain-related behavior in animal models, as well as clinical signs of chronic pain in humans, hyperalgesia and allodynia. Binding of NMDA receptor subunits by the scaffolding protein postsynaptic density protein-95 (PSD-95) can facilitate downstream intracellular signaling and modulate receptor stability, contributing to synaptic plasticity. Here, we show that spinal delivery of the mimetic peptide Tat-NR2B9c disrupts the interaction between PSD-95 and NR2B subunits in the dorsal horn and selectively reduces NMDA receptor-dependent events including wind-up of spinal sensory neurons, and both persistent formalin-induced neuronal activity and pain-related behaviors, attributed to central sensitization. Furthermore, a single intrathecal injection of Tat-NR2B9c in rats with established nerve injury-induced pain attenuates behavioral signs of mechanical and cold hypersensitivity, with no effect on locomotor performance. Thus, uncoupling of PSD-95 from spinal NR2B-containing NMDA receptors may prevent the neuronal plasticity involved in chronic pain and may be a successful analgesic therapy, reducing side effects associated with receptor blockade.     

The 5-HT3 receptor facilitates at-level mechanical allodynia following spinal cord injury

Spinal cord injury (SCI) results in the development of mechanical allodynia immediately rostral to the lesion site, within the dermatome border of normal sensation and sensory loss (at-level mechanical allodynia). We propose that an observed threefold increase in serotonergic fibre immunoreactivity within spinal segments corresponding to these allodynic dermatomes facilitates the maintenance of chronic neuropathic pain via activation of the 5-HT(3) receptor (5-HT(3)-R). Serotonin (5-HT), the non-selective 5-HT(1)/5-HT(2) receptor antagonist, methysergide, the 5-HT(3)-R agonist, m-chlorophenylbiguanide (m-CPBG) or the 5-HT(3)-R antagonist, ondansetron were intrathecally administered five weeks following SCI in rats. Ondansetron produced a robust, long-term reduction of at-level mechanical allodynia, while m-CPBG exacerbated allodynia. Exogenous 5-HT transiently reduced at-level mechanical allodynia. This effect was opposed by methysergide, which enhanced mechanical allodynia. Co-administration of 5-HT and ondansetron produced a short-lasting partial summation of effects, further decreasing mechanical allodynia while co-administration of methysergide attenuated the anti-allodynic effect of ondansetron. Depletion of spinal 5-HT via 5,7-dihydroxytryptamine (5,7-DHT) resulted in decreased at-level mechanical allodynia. The reduction of allodynia by ondansetron was lost following 5,7-DHT administration, suggesting that reduced allodynia following intrathecal ondansetron is via blockade of 5-HT-induced excitation of the 5-HT(3)-R. These results suggest that increased 5-HT fibre density immediately rostral to the SCI lesion site could have transient effects to reduce mechanical allodynia via actions at 5-HT(1) and/or 5-HT(2) receptors. However, the more long-lasting effects of this enhanced serotonergic input may facilitate chronic, at-level allodynia via the 5-HT(3)-R.     

Dermatomal Scratching After Intramedullary Quisqualate Injection: Correlation With Cutaneous Denervation

Central nervous system lesions cause peripheral dysfunctions currently attributed to central cell death that compromises function of intact peripheral nerves. Injecting quisqualate (QUIS) into the rat spinal cord models spinal cord injury (SCI) and causes at-level scratching and self-injury. Such overgrooming was interpreted to model pain until patients with self-injurious scratching after SCI reported itch motivated scratching that was painless because of sensory loss. Because self-injurious scratching is difficult to explain by central mechanisms alone, we hypothesized that QUIS injections damage peripheral axons of at-level afferents. QUIS was injected into thoracic spinal cords of 18 Long-Evans rats. Animals were killed 3 days after overgrooming began or 14 days after injection. Spinal cord lesions were localized and DRG-immunolabeled for ATF-3. At-level and control skin samples were PGP9.5-immunabeled to quantify axons. Eighty-four percent of QUIS rats overgroomed. Skin in these regions had lost two-thirds of epidermal innervation as compared with controls (P < .001). Rats that overgroomed had 47% less axon-length than nongrooming rats (P = .006). The presence of ATF-3 immunolabeled neurons within diagnosis-related groups of QUIS rats indicated death of afferent cell bodies. Overgrooming after QUIS injections may not be due entirely to central changes. As in humans, self-injurious neuropathic scratching appeared to require loss of protective pain sensations in addition to peripheral denervation.PerspectiveThis study suggests that intramedullary injection of quisqualic acid in rats causes death of at-level peripheral as well as central neurons. Self-injurious dermatomal scratching that develops in spinal-injured rats may reflect neuropathic itch and loss of protective pain sensations.     

Analgesic effect of vitamin E is mediated by reducing central sensitization in neuropathic pain

Recent studies suggest that reactive oxygen species (ROS) are critically involved in neuropathic pain. Although vitamin E is a well-known antioxidant, its efficacy on chronic pain is not known. This study investigated the efficacy and mechanisms of vitamin E analgesia in a rat model of neuropathic pain produced by spinal nerve ligation. The effects of vitamin E were investigated using behavioral testing, electrophysiological recording of dorsal horn neurons, and determinations of phosphorylated NMDA receptor subunit 1 (pNR1) levels in the spinal dorsal horn. Results showed that a systemic single injection of a high dose or repetitive daily injections of low doses of vitamin E significantly reduced neuropathic pain behaviors. Vitamin E was also effective in producing analgesia by intrathecal injection, suggesting the importance of spinal mechanisms. In spinal dorsal horn neurons, vitamin E reduced evoked responses to mechanical stimuli as well as the sizes of their receptive fields. In addition, levels of pNR1 in neuropathic rats were also reduced by vitamin E injection. These data suggest that vitamin E produces analgesia in neuropathic rats that is, at least in part, mediated by reducing central sensitization which, in turn, is induced by peripheral nerve injury.     

脊神经结扎神经病理痛模型大鼠脊髓星形胶质细胞激活与痛行为的关系

目的：研究L5脊神经结扎模型大鼠脊髓星形胶质细胞的激活与痛行为之间的关系。方法：44只雄性SD大鼠180—220g，随机分成4组，分别为假手术组、脊神经结扎1天、3天和7天组。行为学上使用von Frey Hair测定大鼠在上述各时间点50％缩足阈的变化（n=8），星形胶质细胞的激活使用免疫组织化学方法观察其特异性标志物GFAP的染色情况（n=3）。结果：（1）脊神经结扎后1天动物出现机械性痛超敏，3天和7天痛行为稳定并持续存在；假手术组未见显著变化。（2）结扎侧脊髓背角星形胶质细胞在术后1天发生激活，3天和7天可见星形胶质细胞强烈的激活反应，假手术组亦可见轻微的激活。（3）脊神经结扎后，星形胶质细胞发生了激活，其激活程度和痛行为的产生和维持紧密相关。结论：脊髓背角星形胶质细胞的激活可能在神经病理痛中发挥作用。     

Chemokine contribution to neuropathic pain: Respective induction of CXCL1 and CXCR2 in spinal cord astrocytes and neurons

Recent studies have indicated an important role of chemokines such as CCL2 in the development of chronic pain. However, the distinct roles of different chemokines in the development and maintenance of neuropathic pain and in their interactions with neurons have not been clearly elucidated. We found that spinal nerve ligation (SNL) not only induced persistent neuropathic pain symptoms, including mechanical allodynia and heat hyperalgesia, but also produced sustained CXCL1 upregulation in the spinal cord. Double staining of immunofluorescence and in situ hybridization revealed that CXCL1 was primarily induced in spinal astrocytes. In cultured astrocytes, tumor necrosis factor-α induced robust CXCL1 expression via the activation of the c-jun N-terminal kinase. Intrathecal administration of CXCL1 neutralizing antibody transiently reduced SNL-induced pain hypersensitivity, suggesting an essential role of CXCL1 in neuropathic pain sensitization. In particular, intraspinal delivery of CXCL1 shRNA lentiviral vectors, either before or after SNL, persistently attenuated SNL-induced pain hypersensitivity. Spinal application of CXCL1 not only elicited pain hypersensitivity but also induced rapid neuronal activation, as indicated by the expression of phosphorylated extracellular signal-regulated kinase and cAMP response element binding protein, and c-Fos in spinal cord neurons. Interestingly, CXCR2, the primary receptor of CXCL1, was upregulated in dorsal horn neurons after SNL, and the CXCR2 antagonist SB225002 completely blocked the CXCL1-induced heat hyperalgesia. SB225002 also attenuated SNL-induced pain hypersensitivity. Collectively, our results have demonstrated a novel form of chemokine-mediated glial-neuronal interaction in the spinal cord that can drive neuropathic pain. Inhibition of the CXCL1-CXCR2 signaling may offer a new therapy for neuropathic pain management.