Activation of ERK／CREB pathway in spinal cord contributes to chronic constrictive injury-induced neuropathic pain in rats

Aim: To investigate whether activation and translocation of extracellular signalregulated kinase (ERK) is involved in the induction and maintenance of neuropathic pain, and effects of activation and translocation of ERK on expression of pCREB and Fos in the chronic neuropathic pain. Methods: Lumbar intrathecal catheters were chronically implanted in male Sprague-Dawley rats. The left sciatic nerve was loosely ligated proximal to the sciatica‘s trifurcation at approximately 1.0 mm intervals with 4-0 silk sutures. The mitogen-activated protein kinase kinase (MEK) inhibitor U0126 or phosphorothioate-modified antisense oligonucleotides (ODN) were intrathecally administered every 12 h, 1 d pre-chronic constriction injury (CCI) and 3 d post-CCI. Thermal and mechanical nociceptive thresholds were assessed with the paw withdrawal latency (PWL) to radiant heat and von Frey filaments. The expression of pERK, pCREB, and Fos were assessed by both Western blotting and immunohistochemical analysis. Results: Intrathecal injection of U0126 or ERK antisense ODN significantly attenuated CCI-induced mechanical allodynia and thermal hyperalgesia. CCI significantly increased the expression of p-ERK-IR neurons in the ipsilateral spinal dorsal horn to injury, not in the contralateral spinal dorsal horn. The time courses of pERK expression showed that the levels of both cytosol and nuclear pERK, but not total ERK, were increased at all points after CCI and reached a peak level on postoperative d 5. CCI also significantly increased the expression of pCREB and Fos. Phospho-CREB-positive neurons were distributed in all laminae of the bilateral spinal cord and Fos was expressed in laminae I and II of the ipsilateral spinal dorsal horn. Intrathecal injection of U0126 or ERK antisense ODN markedly suppressed the increase of CCI-induced pERK, pCREB and c-Fos expression in the spinal cord. Conclusion:The activation of ERK pathways contributes to neuropathic pain in CCI rats, and the function of pERK may partly be accomplished via the cAMP response element binding protein (CREB)-dependent gene expression.     

Decreased expression of glial cell line-derived neurotrophic factor signaling in rat models of neuropathic pain

1. In an attempt to clarify whether glial cell line-derived neurotrophic factor (GDNF), a survival factor for subpopulations of primary afferent neurons, is involved in the states of neuropathic pain, we observed changes in the expressions of GDNF and its signal-transducing receptor Ret after nerve injury in two rat models of neuropathic pain. 2. In the rats treated with sciatic nerve ligation (chronic constrictive injury (CCI) model) or spinal nerve ligation at L5 (SNL model), the thresholds of paw withdrawal in response to mechanical or heat stimuli began to decrease on the injured side within the first week after the operation and the decreases in the thresholds persisted for more than 2 weeks. 3. In CCI-treated rats, the GDNF contents in L4 and L5 dorsal root ganglia (DRGs) on the injured side were markedly decreased at day 7 after the operation and stayed at low levels at day 14. In SNL-treated rats, comparable reductions of GDNF levels in L4 and L5 DRGs on the injured side were observed at 14 postoperative days. 4. Significant decreases of the percentages of DRG neurons expressing Ret were also observed at L4 DRGs in CCI-treated rats at 7 and 14 postoperative days and in SNL-treated rats at 14 days. 5. In CCI- or SNL-treated rats, continuous intrathecal administration of GDNF (12 microg day-1) using an osmotic pump suppressed the increased sensitivities to nociceptive stimuli to control levels. 6. The present results suggested that the dysfunction of GDNF signaling in the nociceptive afferent system may contribute to the development and/or maintenance of neuropathic pain states.     

Glial cell line-derived neurotrophic factormediated enhancement of noradrenergic descending inhibition in the locus coeruleus exerts prolonged analgesia in neuropathic pain

Background and Purpose

The locus coeruleus (LC) is the principal nucleus containing the noradrenergic neurons and is a major endogenous source of pain modulation in the brain. Glial cell line-derived neurotrophic factor (GDNF), a well-established neurotrophic factor for noradrenergic neurons, is a major pain modulator in the spinal cord and primary sensory neurons. However, it is unknown whether GDNF is involved in pain modulation in the LC.

Experimental Approach

Rats with chronic constriction injury (CCI) of the left sciatic nerve were used as a model of neuropathic pain. GDNF was injected into the left LC of rats with CCI for 3 consecutive days and changes in mechanical allodynia and thermal hyperalgesia were assessed. The α₂-adrenoceptor antagonist yohimbine was injected intrathecally to assess the involvement of descending inhibition in GDNF-mediated analgesia. The MEK inhibitor U0126 was used to investigate whether the ERK signalling pathway plays a role in the analgesic effects of GDNF.

Key Results

Both mechanical allodynia and thermal hyperalgesia were attenuated 24 h after the first GDNF injection. GDNF increased the noradrenaline content in the dorsal spinal cord. The analgesic effects continued for at least 3 days after the last injection. Yohimbine abolished these effects of GDNF. The analgesic effects of GDNF were partly, but significantly, inhibited by prior injection of U0126 into the LC.

Conclusions and Implications

GDNF injection into the LC exerts prolonged analgesic effects on neuropathic pain in rats by enhancing descending noradrenergic inhibition.     

川芎嗪对大鼠坐骨神经慢性压迫性损伤L4/L5段背根神经节P2X3受体表达的影响

目的 探讨川芎嗪抑制P2X₃受体介导慢性神经病理痛的作用途径。方法 制备大鼠坐骨神经慢性压迫性损伤(CCI)神经病理痛模型，于d 2起ip川芎嗪100 mg·kg^-1，每天1次，共14 d。免疫组织化学法观察CCI大鼠L₄/L₅段背根神经节P2X₃受体的表达，全细胞膜片钳技术测定新鲜分离的L₄/L₅段背根神经节三磷酸腺苷（ATP）和α,β-亚甲基三磷酸腺苷（α,β-meATP）激活的电流。结果 与正常对照组比较，正常大鼠ip川芎嗪14 d，L₄/L₅段背根神经节P2X₃受体表达、ATP激活电流和α,β-meATP激活电流无明显变化，假手术组亦无明显变化。与假手术组比较，CCI模型组大鼠L₄/L₅段背根神经节P2X₃受体的表达、ATP和α,β-meATP激活电流明显增强。CCI大鼠ip川芎嗪14 d，L₄/L₅段背根神经节P2X₃受体表达、ATP和α,β-meATP激活电流较CCI模型组明显降低。结论 川芎嗪可抑制CCI大鼠L₄/L₅段背根神经节P2X₃受体的表达，从而对P2X₃受体介导的神经病理痛产生抑制作用。     

Effects of intrathecal epigallocatechin gallate, an inhibitor of Toll-like receptor 4, on chronic neuropathic pain in rats

Numerous studies revealed that spinal inflammation and immune response play an important role in neuropathic pain. In this study, we investigated the effects of intrathecal injection of a Toll-like receptor (TLR4) inhibitor epigallocatechin gallate (EGCG) on neuropathic pain induced by chronic constriction injury of the sciatic nerve (CCI). A total of 120 rats were randomly assigned into 4 groups: sham-operated group, CCI group, CCI plus normal saline group and CCI plus EGCG group. CCI and sham surgeries were performed and both thermal hyperalgesia and mechanical allodynia were tested. Lumbar spinal cord was sampled and the mRNA and protein expressions of TLR4 and High Mobility Group 1 protein (HMGB1) were detected, the contents of tumor necrosis factor alpha (TNF-α), interleukin-1 beta (IL-1β) and interleukin-10 (IL-10) were measured by ELISA, and immunohistochemistry for nuclear factor kappa B (NF-κB) was also carried out. When compared with the sham group, both mechanical and heat pain thresholds were significantly decreased, and the mRNA and protein expressions of TLR4 and HMGB1, the contents of TNF-α, IL-1β and IL-10 in the spinal cords and NF-κB expression in the spinal dorsal horn were markedly increased in CCI rats (P<0.05). After intrathecal injection of EGCG (1mg/kg) once daily from 1day before to 3days after CCI surgery, the expressions of TLR4, NF-κB, HMGB1, TNF-α and IL-1β were markedly decreased while the content of IL-10 in the spinal cord increased significantly accompanied by dramatical improvement of pain behaviors in CCI rats (P<0.05). These results show that the TLR4 signaling pathway plays an important role in the occurrence and development of neuropathic pain, and the therapy targeting TLR4 might be a novel strategy in the treatment of neuropathic pain.     

Cytokine activin C ameliorates chronic neuropathic pain in peripheral nerve injury rodents by modulating the TRPV1 channel

Background and PurposeThe cytokine activin C is mainly expressed in small‐diameter dorsal root ganglion (DRG) neurons and suppresses inflammatory pain. However, the effects of activin C in neuropathic pain remain elusive.Experimental ApproachMale rats and wild‐type and TRPV1 knockout mice with peripheral nerve injury ‐ sciatic nerve axotomy and spinal nerve ligation in rats; chronic constriction injury (CCI) in mice – provided models of chronic neuropathic pain. Ipsilateral lumbar (L)4–5 DRGs were assayed for activin C expression. Chronic neuropathic pain animals were treated with intrathecal or locally pre‐administered activin C or the vehicle. Nociceptive behaviours and pain‐related markers in L4–5 DRGs and spinal cord were evaluated. TRPV1 channel modulation by activin C was measured.Key ResultsFollowing peripheral nerve injury, expression of activin βC subunit mRNA and activin C protein was markedly up‐regulated in L4–5 DRGs of animals with axotomy, SNL or CCI. [Correction added on 26 November 2020, after first online publication: The preceding sentence has been corrected in this current version.] Intrathecal activin C dose‐dependently inhibited neuropathic pain in spinal nerve ligated rats. Local pre‐administration of activin C decreased neuropathic pain, macrophage infiltration into ipsilateral L4–5 DRGs and microglial reaction in L4–5 spinal cords of mice with CCI. In rat DRG neurons, activin C enhanced capsaicin‐induced TRPV1 currents. Pre‐treatment with activin C reduced capsaicin‐evoked acute hyperalgesia and normalized capsaicin‐evoked persistent hypothermia in mice. Finally, the analgesic effect of activin C was abolished in TRPV1 knockout mice with CCI.Conclusion and ImplicationsActivin C inhibits neuropathic pain by modulating TRPV1 channels, revealing potential analgesic applications in chronic neuropathic pain therapy.     

神经病理性疼痛大鼠背根神经节TRPM8表达变化的研究

目的: 观察慢性神经病理性疼痛大鼠背根神经节TRPM8表达的变化.方法: 健康成年雄性SD大鼠72只,体质量250~280 g,随机分为慢性坐骨神经束缚性损伤(CCI)组和假手术组(n = 36), CCI或假手术前1 d,术后1、4、7、10、14 d每组各随机取6只大鼠,测定冷痛阈值、热痛阈值和机械痛阈值后立即处死大鼠,取L5背根神经节行免疫组织化学染色,观察瞬时受体电位melastatin 8(TRPM8)在慢性神经病理性疼痛下的表达变化.结果: CCI组于术后4 d术侧冷痛阈值、机械痛阈值和热痛阈值开始下降,至术后14 d仍处于较低水平,冷痛阈值和热痛阈值于术后10 d降至最低,机械痛阈值术后14 d降至最低(P ＜ 0.05或P ＜ 0.01);CCI组术侧L5背根神经节TRPM8的表达于术后4 d开始增加,术后10 d达到高峰,至术后14 d仍维持在高水平(P ＜ 0.05或P ＜ 0.01).结论: 背根神经节TRPM8受体表达的上调参与神经病理性疼痛的发生和维持.     

The effects of galanin on neuropathic pain in streptozotocin-induced diabetic rats

Diabetic neuropathy is a common complication associated with diabetes and is frequently painful. However, mechanisms responsible for diabetic neuropathic pain are still unclear. Experimental evidence has shown that the galanin and its receptor are involved in pain sensitization. The objective of the present study was to investigate the role of galanin and its receptor antagonist or agonist on neuropathic pain in streptozotocin-induced diabetic rats. The expression of galanin, galanin receptors 1 and 2 in dorsal root ganglion (DRG) and spinal dorsal horn (SDH) in diabetic rats were detected by Western blot assay. The effects of galanin, galanin receptor antagonist M35, galanin receptor 1 agonist M617, and galanin receptor 2 agonist AR-M1896 on neuropathic pain were evaluated by mechanical stimuli. The results showed that (1) the diabetic rats showed a significant mechanical hyperalgesia between 4 and 12weeks; (2) galanin receptor 1 expression decreased in SDH in diabetic rats; (3) galanin receptor 2 expression decreased in DRG and SDH in diabetic rats; (4) intrathecal administration of exogenous galanin attenuated diabetic neuropathic pain, this effect could be blocked by pre-treatment with galanin receptor antagonist M35; and (5) intrathecal administration of galanin receptor 1 agonist M617, but not galanin receptor 2 agonist AR-M1896, attenuated diabetic neuropathic pain. These results imply that galanin acts through receptor 1, but not galanin receptor 2, to exert analgesic effect in diabetic neuropathic pain and is one of the potential therapeutic targets on diabetic neuropathic pain sensitization.     

Role of reactive oxygen species and spinal cord apoptotic genes in the development of neuropathic pain.

A mouse model of neuropathic pain consisting of chronic constriction injury (CCI) of the sciatic nerve was used to examine the involvement of reactive oxygen species (ROS) in early spinal cord pro-apoptotic gene over-expression during the development of neuropathic pain. RT-PCR analysis showed increased expression of bax, apoptotic protease-activating factor-1 (apaf-1), and caspase-9 in the dorsal horn spinal cord 3 days after chronic constriction injury of sciatic nerve. Consistent with biomolecular data, a marked increase in TUNEL-positive and caspase-3 active form was observed by 3 days CCI. Administration of phenyl-N-tert-butylnitrone (PBN), a potent ROS scavenger, reduced the development of thermal hyperalgesia and mechanical allodynia at 1 and 3 days post-CCI, and decreased the mRNA levels of bax, apaf-1, and caspase-9. PBN also reduced apoptotic and active Caspase-3 positive profiles in the superficial laminae (I-III) of the spinal cord. This study provides evidence that PBN inhibits over-expression of pro-apoptotic genes and neural apoptosis in the spinal cord dorsal horn induced by early-CCI of the sciatic nerve. These findings suggest that ROS regulate expression of some apoptotic genes which might play a role in the onset of neuropathic pain.     

蛛网膜下隙注射胶质细胞源性神经营养因子抑制脊神经结扎大鼠脊髓背角胶质原纤维酸性蛋白表达

目的观察蛛网膜下隙注射胶质细胞源性神经营养因子(GDNF)对脊神经结扎(SNL)大鼠脊髓背角胶质原纤维酸性蛋白(GFAP)的影响。方法采用结扎SD大鼠第5~第6腰脊神经(L5~L6)制备SNL模型。术后隔日一次性蛛网膜下隙注射10μl GDNF2 g·L-1组。术后第3,7和14天采用免疫组织化学和Western免疫印迹法测定脊髓背角处GFAP蛋白表达。结果免疫组化结果显示,SNL组在术后第3天、第7天和第14天脊髓GFAP阳性细胞数目明显增加,可持续至第14天,而正常对照组与假手术组的星形胶质细胞未发生此种改变。GDNF组的阳性细胞显著减少。Western印迹结果显示,正常对照组和假手术组脊髓背角均出现GFAP免疫阳性条带,灰度值较低;SNL在术后第3天即可诱导脊髓背角GFAP蛋白的表达,随着时间的延长,GFAP蛋白的灰度值逐渐升高,术后第3,7,14天分别为2.55±0.33,2.88±0.79和3.12±0.75(P<0.01)。与SNL模型组比较,GDNF可显著降低GFAP的表达水平,术后第3,7,14天分别为1.61±0.38,1.65±0.64和1.57±0.41(P<0.01)。结论蛛网膜下隙注射GDNF减轻大鼠神经病理性疼痛机制可能与其抑制脊髓背角GFAP蛋白表达有关。     

Intrathecal administration of Cav3.2 and Cav3.3 antisense oligonucleotide reverses tactile allodynia and thermal hyperalgesia in rats following chronic compression of dorsal root of ganglion

AIM: The present study aimed to elucidate the role of T-subtype calcium channels (Cav3.1, Cav3.2, and Cav3.3) in the pathogenesis of neuropathic pain at spinal level. METHODS: The chronic compression of the dorsal root ganglion (CCD) rat model was adopted. The antisense oligonucleotide of Cav3.1, Cav3.2, and Cav3.3 or normal saline (NS) were intrathecally administered twice per day from the first day to the fourth day after operation. Paw mechanical withdrawal threshold and paw thermal withdrawal latency were measured to evaluate the tactile allodynia and thermal hyperalgesia, respectively. RESULTS: CCD rats developed reliable tactile allodynia and thermal hyperalgesia after operation. Intrathecal administration of antisense oligonucleotide of Cav3.2 and Cav3.3 significantly relieved tactile allodynia and thermal hyperalgesia in CCD rats, but not Cav3.1. CONCLUSION: Cav3.2 and Cav3.3 subtype calcium channels in the spinal cord may play an important role in the pathogenesis of neuropathic pain, which may contribute to the management of the neuropathic pain.     

Leukotrienes in nociceptive pathway and neuropathic/inflammatory pain

The purpose of this review is to summarize the recent studies examining the expression of leukotrienes (LTs) and their receptors in nociceptive pathways, and their crucial roles in pathological pain conditions. LTs belong to a large family of lipid mediators, termed eicosanoids, which are derived from arachidonic acids and released from the cell membrane by phospholipases. LTs are known to be important factors in a variety of local and systemic diseases and allergic/inflammatory diseases. We examined whether LTs were implicated in neuropathic pain following peripheral nerve injury. Using the SNI model in rats, we investigated the expression of LT synthases and receptors mRNAs in the spinal cord and the roles on the pain behaviors. We found the expression of 5-lipoxygenase (5-LO), FLAP and the cysteinyl leukotrienes (CysLT1) mRNAs in spinal microglia, LTA4h and LTC4s mRNAs in both spinal neurons and microglia, and BLT1 mRNA in spinal neurons. Administration of the 5-LO inhibitor or the receptor antagonists suppressed mechanical allodynia. Our findings suggest that the increase of LT synthesis in spinal microglia produced via p38 mitogen-activated protein kinase (MAPK) plays a role in the generation of neuropathic pain. We also examined the expression and roles on pain behaviors of LT receptors in the dorsal root ganglion (DRG) using a peripheral inflammation model. The data indicate CysLT2 expressed in DRG neurons may play a role as a modulator of P2X3, and contribute to the potentiation of the neuronal activity following peripheral inflammation. This review summarizes the hypothesis that LTs might work in the spinal cord and primary afferent in pathological pain conditions.     

The antipsychotic drug,fluphenazine, effectively reverses mechanical allodynia in rat models of neuropathic pain

Rationale Fluphenazine is a potent antipsychotic drug used to treat schizophrenia and other psychotic symptoms. Its clinical benefit is mainly mediated by the antagonism of dopamine D2 receptors. We have recently discovered, however, that fluphenazine is also a potent sodium channel blocker, a property that may offer additional therapeutical indications, including analgesia. Objectives The present study sought to determine the analgesic effect of fluphenazine on neuropathic pain in animal models. Methods The effect of fluphenazine on mechanical allodynia was assessed in three animal neuropathic pain models, including spinal nerve ligation, chronic constriction nerve injury (CCI), and sural-spared sciatic nerve injury models. Results Systemic fluphenazine effectively attenuated mechanical allodynia in all three rat neuropathic pain models at doses (0.03–0.3 mg/kg) that approximate those used in rodent models of psychosis. In parallel with its in vivo antiallodynic effect, fluphenazine (3–30 μM) effectively suppressed the ectopic discharges in injured afferent fibers without affecting the propagation of action potentials evoked by electrical nerve stimulation in an ex vivo dorsal root ganglia (DRG)-nerve preparation excised from CCI rats. Furthermore, similar concentrations of fluphenazine significantly blocked sodium channels in DRG neurons. Conclusions The inhibitory action of fluphenazine on ectopic afferent discharges may be due to its ability to block voltage-gated sodium channels, and this may also provide a mechanistic basis for the drug’s antiallodynic effect in animal models of neuropathic pain. In summary, our study demonstrates that the classic antipsychotic drug fluphenazine has antiallodynic properties in multiple rodent models of nerve injury-induced neuropathic pain. Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Increase in hemokinin-1 mRNA in the spinal cord during the early phase of a neuropathic pain state

Background and purpose:

Substance P (SP), a representative member of the tachykinin family, is involved in nociception under physiological and pathological conditions. Recently, hemokinin-1 (HK-1) was identified as a new member of this family. Although HK-1 acts on NK₁ tachykinin receptors that are thought to be innate for SP, the roles of HK-1 in neuropathic pain are still unknown.

Experimental approach:

Using rats that had been subjected to chronic constrictive injury (CCI) of the sciatic nerve as a neuropathic pain model, we examined the changes in expression of SP- and HK-1-encoding genes (TAC1 and TAC4, respectively) in the L4/L5 spinal cord and L4/L5 dorsal root ganglia (DRGs) in association with changes in pain-related behaviours in this neuropathic pain state.

Key results:

The TAC4 mRNA level was increased on the ipsilateral side of the dorsal spinal cord, but not in DRGs, at day 3 after CCI. In contrast, the TAC1 mRNA level was significantly increased in the DRGs at day 3 after CCI without any changes in the dorsal spinal cord. Analysis of a cultured microglial cell line revealed the presence of TAC4 mRNA in microglial cells. Minocycline, an inhibitor of microglial activation, blocked the increased expression of TAC4 mRNA after CCI and inhibited the associated pain-related behaviours and microglial activation in the spinal cord.

Conclusions and implications:

The present results suggest that HK-1 expression is increased at least partly in activated microglial cells after nerve injury and is clearly involved in the early phase of neuropathic pain.     

Activation of NK₁ receptors in the locus coeruleus induces analgesia through noradrenergic-mediated descending inhibition in a rat model of neuropathic pain

BACKGROUND AND PURPOSE

The locus coeruleus (LC) is a major source of noradrenergic projections to the dorsal spinal cord, and thereby plays an important role in the modulation of nociceptive information. The LC receives inputs from substance P (SP)-containing fibres from other regions, and expresses the NK₁ tachykinin receptor, a functional receptor for SP. In the present study, we investigated the roles of SP in the LC in neuropathic pain.

EXPERIMENTAL APPROACH

Chronic constriction injury (CCI) of the left sciatic nerve was performed in rats to induce neuropathic pain. After development of neuropathic pain, SP was injected into the LC and the nocifensive behaviours were assessed. The involvement of noradrenergic descending inhibition in SP-induced analgesia was examined by i.t. administration of yohimbine, an α₂-adrenoceptor antagonist. NK₁ receptor expression in the LC was examined by immunohistochemistry.

KEY RESULTS

In CCI rats, mechanical allodynia was alleviated by SP injection into the LC. These effects were abolished by prior injection of WIN 51708, an NK₁ receptor antagonist, into the LC or i.t. treatment with yohimbine. NK₁ receptor-like immunoreactivity was observed in noradrenergic neurons throughout the LC in intact rats, and remained unchanged after CCI.

CONCLUSION AND IMPLICATIONS

SP in the LC exerted analgesic effects on neuropathic pain through NK₁ receptor activation and resulted in facilitation of spinal noradrenergic transmission. Accordingly, manipulation of the SP/NK₁ receptor signalling pathway in the LC may be a promising strategy for effective treatment of neuropathic pain.     

Inhibition of SNL-induced upregulation of CGRP and NPY in the spinal cord and dorsal root ganglia by the 5-HT(2A) receptor antagonist ketanserin in rats

Our previous study has demonstrated that topical and systemic administration of the 5-HT_2A receptor antagonist ketanserin attenuates neuropathic pain. To explore the mechanisms involved, we examined whether ketanserin reversed the plasticity changes associated with calcitonin gene-related peptides (CGRP) and neuropeptide Y (NPY) which may reflect distinct mechanisms: involvement and compensatory protection. Behavioral responses to thermal and tactile stimuli after spinal nerve ligation (SNL) at L5 demonstrated neuropathic pain and its attenuation in the vehicle- and ketanserin-treated groups, respectively. SNL surgery induced an increase in CGRP and NPY immunoreactivity (IR) in laminae I-II of the spinal cord. L5 SNL produced an expression of NPY-IR in large, medium and small diameter neurons in dorsal root ganglion (DRG) only at L5, but not adjacent L4 and L6. Daily injection of ketanserin (0.3 mg/kg, s.c.) for two weeks suppressed the increase in CGRP-IR and NPY-IR in the spinal cord or DRG. The present study demonstrated that: (1) the expression of CGRP was enhanced in the spinal dorsal horn and NPY was expressed in the DRG containing injured neurons, but not in the adjacent DRG containing intact neurons, following L5 SNL; (2) the maladaptive changes in CGRP and NPY expression in the spinal cord and DRG mediated the bioactivity of 5-HT/5-HT_2A receptors in neuropathic pain and (3) the blockade of 5-HT_2A receptors by ketanserin reversed the evoked upregulation of both CGRP and NPY in the spinal cord and DRG contributing to the inhibition of neuropathic pain.