Comparative study of cold hyperalgesia and mechanical allodynia in two animal models of neuropathic pain: different etiologies and distinct pathophysiological mechanisms

Abstract Neuropathic pain (NP) affects more than 8% of the global population. The proposed action of the transient receptor potential ankyrin 1 (TRPA1) as a mechanosensor and the characterization of the transient receptor potential melastatin 8 (TRPM8) as a cold thermosensor raises the question of whether these receptors are implicated in NP. Our study aimed to evaluate the involvement of TRPA1 and TRPM8 in cold and mechanical signal transduction to obtain a comparative view in rat models of streptozotocin-induced diabetes (STZ) and chronic constriction injury of the sciatic nerve (CCI). The electronic von Frey test showed that STZ rats presented mechanical allodynia that was first evidenced on the 14th day after diabetes confirmation, and four days after CCI. This phenomenon was reduced by the intraplantar (ipl) administration of a TRPA1 receptor antagonist (HC-030031; 40 µL/300 µg/paw) in both NP models. Only CCI rats displayed cold hyperalgesia based on the cold plate test. The pharmacological blocking of TRPA1 through the injection of the antagonist attenuated cold hyperalgesia in this NP model. STZ animals showed a reduction in the number of flinches induced by the intraplantar injection of mustard oil (MO; TRPA1 agonist; 0.1%/50 µL/paw), or intraplantar injection of menthol (MT; TRPM8 agonist; 0.5% and 1%/50 µL/paw). The response induced by the ipl administration of MT (1%/50 µL/paw) was significantly different between the CCI and SHAM groups. Together, these data suggest a different pattern in nociceptive behavior associated with different models of NP, suggesting a variant involvement of TRPA1 and TRPM8 in both conditions

The profile of dexamethasone combined with transcranial direct current stimulation in rats submitted to an arthritis model / O perfil da dexametasona combinada com estimulação transcraniana por corrente contínua em ratos submetidos a um modelo de artrite

ABSTRACT BACKGROUND AND OBJECTIVES: To pursue safer and more effective treatments for rheumatoid arthritis, the effect of dexamethasone treatment (DEX, 0.25mg/kg) combined with transcranial direct current stimulation (tDCS) in the behavior and neurochemical parameters of arthritic rats was evaluated. METHODS: Thirty-six Wistar rats were divided into four groups: control+DEX (CTRL+DEX), arthritis+DEX (RA+DEX), arthritis+DEX+sham-tDCS (RA+DEX+sham-tDCS) and arthritis+DEX+tDCS (RA+DEX+tDCS). The arthritic model (RA) was induced by complete Freund's adjuvant (CFA) paw administration. Paw edema and mechanical allodynia were assessed by plethysmometer and von Frey apparatus, respectively. Fourteen days after the CFA injection, rats received the treatment for eight days (DEX and/or tDCS). Behavioral parameters were measured with the Open-Field test. ELISA was used to evaluate hippocampal and spinal cord tumor necrosis factor (TNF-α) levels, cerebral cortex and brainstem BDNF levels. RESULTS: In pre-treatment measurements, arthritic rats presented an increase in joint swelling and mechanical allodynia when compared to the control group, confirming chronic pain establishment. A slight antinociceptive effect of dexamethasone combined with tDCS in the pain model was observed. The pain model significantly induced an increase in the grooming behavior and a reduction in the spinal cord and hippocampal TNF-α levels; these effects were reverted in the sham- and active-tDCS-treated rats. However, no effects of DEX or tDCS were observed in the BDNF levels in the cerebral cortex and brainstem. CONCLUSION: Despite the small effect observed, tDCS treatment cannot be discarded as a non-pharmacological adjuvant technique for inflammatory chronic pain treatment.

RESUMO JUSTIFICATIVA E OBJETIVOS: Para investigar métodos mais seguros e eficazes para o manejo da artrite reumatoide, avaliou-se o efeito do tratamento com dexametasona (DEX, 0,25mg/kg) combinado com estimulação transcraniana por corrente contínua (ETCC) sobre parâmetros comportamentais e bioquímicos de ratos submetidos a um modelo de artrite reumatoide. MÉTODOS: Trinta e seis ratos Wistar foram alocados em 4 grupos: controle+DEX (CTRL+DEX), artrite+DEX (AR+DEX), artrite+DEX+sham-ETCC (AR+DEX+sham-ETCC) e artrite+DEX+ETCC (AR+DEX+ETCC). O modelo de artrite foi induzido pela administração de complete Freund's adjuvant (CFA) na pata. Edema na pata e a alodínia mecânica foram avaliadas por pletismômetro e teste de von Frey, respectivamente. 14 dias após injeção de CFA, ratos foram tratados por 8 dias (DEX e/ou ETCC). Atividade locomotora foi avaliada pelo teste do campo aberto. TNF-alfa (hipocampo e medula espinal) e BDNF (córtex e tronco) foram mensurados por ELISA. RESULTADOS: Nas medições pré-tratamento, ratos com artrite exibiram aumento de o inchaço articular e alodínia mecânica comparados ao grupo controle, confirmando o estabelecimento de modelo de dor crônica. Também se observou discreto efeito antinociceptivo da dexametasona combinada com ETCC no modelo de artrite. O modelo de dor induziu um aumento no comportamento de grooming e reduziu os níveis de TNF-alfa no hipocampo; estes efeitos foram revertidos nos grupos sham- e ETCC ativo. Entretanto, não foram observados efeitos da DEX ou ETCC nos níveis de BDNF no córtex cerebral ou no tronco encefálico. CONCLUSÃO: Apesar dos discretos efeitos observados, não se pode descartar a ETCC como uma abordagem terapêutica não farmacológica para o manejo da dor crônica inflamatória na artrite reumatoide.

Indomethacin attenuates mechanical allodynia during the organization but not the maintenance of the peripheral neuropathic pain induced by nervus ischiadicus chronic constriction injury

The neurochemical mechanisms underlying neuropathic pain (NP) are related to peripheral and central sensitization caused by the release of inflammatory mediators in the peripheral damaged tissue and ectopic discharges from the injured nerve, leading to a hyperexcitable state of spinal dorsal horn neurons. The aim of this work was to clarify the role played by cyclooxygenase (COX) in the lesioned peripheral nerve in the development and maintenance of NP by evaluating at which moment the non-steroidal anti-inflammatory drug indomethacin, a non-selective COX inhibitor, attenuated mechanical allodynia after placing one loose ligature around the nervus ischiadicus, an adaptation of Bennett and Xie's model in rodents. NP was induced in male Wistar rats by subjecting them to chronic constriction injury (CCI) of the nervus ischiadicus, placing one loose ligature around the peripheral nerve, and a sham surgery (without CCI) was used as control. Indomethacin (2 mg/kg) or vehicle was intraperitoneally and acutely administered in each group of rats and at different time windows (1, 2, 4, 7, 14, 21, and 28 days) after the CCI or sham surgical procedures, followed by von Frey's test for 30 min. The data showed that indomethacin decreased the mechanical allodynia threshold of rats on the first, second, and fourth days after CCI (P<0.05). These findings suggested that inflammatory mechanisms are involved in the induction of NP and that COX-1 and COX-2 are involved in the induction but not in the maintenance of NP.

Spinal P2X7R contributes to streptozotocin-induced mechanical allodynia in mice / 浙江大学学报（英文版）（B辑：生物医学和生物技术）

Painful diabetic neuropathy (PDN) is a diabetes mellitus complication. Unfortunately, the mechanisms underlying PDN are still poorly understood. Adenosine triphosphate (ATP)-gated P2X7 receptor (P2X7R) plays a pivotal role in non-diabetic neuropathic pain, but little is known about its effects on streptozotocin (STZ)-induced peripheral neuropathy. Here, we explored whether spinal cord P2X7R was correlated with the generation of mechanical allodynia (MA) in STZ-induced type 1 diabetic neuropathy in mice. MA was assessed by measuring paw withdrawal thresholds and western blotting. Immunohistochemistry was applied to analyze the protein expression levels and localization of P2X7R. STZ-induced mice expressed increased P2X7R in the dorsal horn of the lumbar spinal cord during MA. Mice injected intrathecally with a selective antagonist of P2X7R and P2X7R knockout (KO) mice both presented attenuated progression of MA. Double-immunofluorescent labeling demonstrated that P2X7R-positive cells were mostly co-expressed with Iba1 (a microglia marker). Our results suggest that P2X7R plays an important role in the development of MA and could be used as a cellular target for treating PDN.

Glial Cell Line-derived Neurotrophic Factor-overexpressing Human Neural Stem/Progenitor Cells Enhance Therapeutic Efficiency in Rat with Traumatic Spinal Cord Injury

Spinal cord injury (SCI) causes axonal damage and demyelination, neural cell death, and comprehensive tissue loss, resulting in devastating neurological dysfunction. Neural stem/progenitor cell (NSPCs) transplantation provides therapeutic benefits for neural repair in SCI, and glial cell line-derived neurotrophic factor (GDNF) has been uncovered to have capability of stimulating axonal regeneration and remyelination after SCI. In this study, to evaluate whether GDNF would augment therapeutic effects of NSPCs for SCI, GDNF-encoding or mock adenoviral vector-transduced human NSPCs (GDNF-or Mock-hNSPCs) were transplanted into the injured thoracic spinal cords of rats at 7 days after SCI. Grafted GDNF-hNSPCs showed robust engraftment, long-term survival, an extensive distribution, and increased differentiation into neurons and oligodendroglial cells. Compared with Mock-hNSPC- and vehicle-injected groups, transplantation of GDNF-hNSPCs significantly reduced lesion volume and glial scar formation, promoted neurite outgrowth, axonal regeneration and myelination, increased Schwann cell migration that contributed to the myelin repair, and improved locomotor recovery. In addition, tract tracing demonstrated that transplantation of GDNF-hNSPCs reduced significantly axonal dieback of the dorsal corticospinal tract (dCST), and increased the levels of dCST collaterals, propriospinal neurons (PSNs), and contacts between dCST collaterals and PSNs in the cervical enlargement over that of the controls. Finally grafted GDNF-hNSPCs substantially reversed the increased expression of voltage-gated sodium channels and neuropeptide Y, and elevated expression of GABA in the injured spinal cord, which are involved in the attenuation of neuropathic pain after SCI. These findings suggest that implantation of GDNF-hNSPCs enhances therapeutic efficiency of hNSPCs-based cell therapy for SCI.

Effect of exchange protein directly activated by cyclic adenosine monophosphate 1 on mechanical pain of diabetic rats / 中华内分泌代谢杂志

Objective To investigate effect of exchange protein directly activated by cyclic adenosine monophosphate 1 on mechanical pain in diabetic rats.Methods Male SD rats were randomly divided into CON group and STZ group. CON group were further randomly divided into CONshRNA group,and CONEpac1shRNA group. STZ group induced diabetic mechanical pain (DMA) model were randomly divided into DMA shRNA group, DMA Epac1shRNA group,DMANS group,and DMA 8-pCPT group. The Epac1shRNA lentivirus vector is used to inhibit the expression of Epac1, and the shRNA lentivirus vector is negative control, and 8-pCPT is the activator of Epac1. Group CONshRNA and group DMAshRNA were given intrathecal injection of control shRNA lentivirus vector. Group CONEpac1shRNA and group DMAEpac1shRNA were injected with Epac1shRNA lentivirus carrier, DMANS group was injected into the plantar saline,and 8-pCPT in group DMA8-pCPT was injected into the foot. The changes of hind paw retraction threshold(PWT) were observed and the expression of guanine nucleotide transforming factor 1(Epac1) mRNA was detected by Real Time-PCR and Western blot in rat dorsal root ganglion(DRG) And protein expression changes.Results Compared with CON rats,the mechanical pain threshold of STZ rats decreased (P=0.035). Compared with saline group,the pain of injection Epac1 activator 8-pCPT group was prolonged(2h, P=0.012;4h,P=0.020). The expression of Epac1 mRNA and protein was significantly higher in the DMA group than in the CON group(both P<0.01). Intrathecal injection of shRNA reduced the expression of Epac1 mRNA and protein(P<0.01,P=0.020),and the PWT of the DMA group was significantly lower than that of the CON group (P=0.006).Conclusion Epac1 expression in diabetic rats with increased pain,and down-regulation of Epac1 may relieve pain.

Application of three artemisinin derivatives in neuropathic pain: evaluating co-curation of nociceptive and emotional syndromes in spinal cord ligation mice / 中国中药杂志

In this study, on aspects of the nociceptive, anxiety and depressive syndromes in neuropathic pain (NP), the effects of dihydroartemisinine (DHA), artesunate (ART) and artemether (ARTN) (40 mg·kg⁻¹) were analyzed in the spinal cord ligation (SNL) mice. Clinical equivalent dose of the first-line drug for NP, pregabalin (PGB, 25 mg·kg⁻¹) and amitriptyline (ARP, 20 mg·kg⁻¹), were used as positive controls. General, from day 7 to 14, significant remissions of the nociceptive, anxiety and depressive behaviors were achieved by DHA, ART and ARTN separately. Moreover, on day 14, on aspects of the nociceptive behaviors, analyzed 1.5 h after the gavage administration, no significant difference between the shamed mice and mice administrated with DHA, ART and ARTN was detected; analyzed 3 h after the gavage, significant decreases of pain thresholds in ARTN, but not in DHA nor ART group, were detected as compared with thresholds measured 1.5 h; analyzed 24 h after gavage, pain thresholds in DHA, ART and ARTN were still higher than PGB, in spite of the significant decreases as compared to Sham group. On aspects of the anxiety and depressive behaviors, no significant difference was detected between the shamed mice and mice administrated with DHA nor ART. However, differences still remained between the shamed ones and ones administrated with ARTN. Preliminarily, the effects of DHA, ART and ARTN were consolidated in SNL mice. On aspects of the duration of analgesic effects and the control of negative emotion, ART and ARTN were proven more favorable than ARTN.

Involvement of NF-κB and the CX3CR1 Signaling Network in Mechanical Allodynia Induced by Tetanic Sciatic Stimulation / 神经科学通报·英文版

Tetanic stimulation of the sciatic nerve (TSS) triggers long-term potentiation in the dorsal horn of the spinal cord and long-lasting pain hypersensitivity. CX3CL1-CX3CR1 signaling is an important pathway in neuronal-microglial activation. Nuclear factor κB (NF-κB) is a key signal transduction molecule that regulates neuroinflammation and neuropathic pain. Here, we set out to determine whether and how NF-κB and CX3CR1 are involved in the mechanism underlying the pathological changes induced by TSS. After unilateral TSS, significant bilateral mechanical allodynia was induced, as assessed by the von Frey test. The expression of phosphorylated NF-κB (pNF-κB) and CX3CR1 was significantly up-regulated in the bilateral dorsal horn. Immunofluorescence staining demonstrated that pNF-κB and NeuN co-existed, implying that the NF-κB pathway is predominantly activated in neurons following TSS. Administration of either the NF-κB inhibitor ammonium pyrrolidine dithiocarbamate or a CX3CR1-neutralizing antibody blocked the development and maintenance of neuropathic pain. In addition, blockade of NF-κB down-regulated the expression of CX3CL1-CX3CR1 signaling, and conversely the CX3CR1-neutralizing antibody also down-regulated pNF-κB. These findings suggest an involvement of NF-κB and the CX3CR1 signaling network in the development and maintenance of TSS-induced mechanical allodynia. Our work suggests the potential clinical application of NF-κB inhibitors or CX3CR1-neutralizing antibodies in treating pathological pain.

Allopregnanolone suppresses mechanical allodynia and internalization of neurokinin-1 receptors at the spinal dorsal horn in a rat postoperative pain model

BACKGROUND: To identify a new strategy for postoperative pain management, we investigated the analgesic effects of allopregnanolone (Allo) in an incisional pain model, and also assessed its effects on the activities of the primary afferent fibers at the dorsal horn. METHODS: In experiment 1, 45 rats were assigned to Control, Allo small-dose (0.16 mg/kg), and Allo large-dose (1.6 mg/kg) groups (n = 15 in each). The weight bearing and mechanical withdrawal thresholds of the hind limb were measured before and at 2, 24, 48, and 168 h after Brennan's surgery. In experiment 2, 16 rats were assigned to Control and Allo (0.16 mg/kg) groups (n = 8 in each). The degree of spontaneous pain was measured using the grimace scale after the surgery. Activities of the primary afferent fibers in the spinal cord (L6) were evaluated using immunohistochemical staining. RESULTS: In experiment 1, the withdrawal threshold of the Allo small-dose group was significantly higher than that of the Control group at 2 h after surgery. Intergroup differences in weight bearing were not significant. In experiment 2, intergroup differences in the grimace scale scores were not significant. Substance P release in the Allo (0.16 mg/kg) group was significantly lower than that in the Control group. CONCLUSIONS: Systemic administration of Allo inhibited mechanical allodynia and activities of the primary afferent fibers at the dorsal horn in a rat postoperative pain model. Allo was proposed as a candidate for postoperative pain management.

Preemptive application of QX-314 attenuates trigeminal neuropathic mechanical allodynia in rats

The aim of the present study was to examine the effects of preemptive analgesia on the development of trigeminal neuropathic pain. For this purpose, mechanical allodynia was evaluated in male Sprague-Dawley rats using chronic constriction injury of the infraorbital nerve (CCI-ION) and perineural application of 2% QX-314 to the infraorbital nerve. CCI-ION produced severe mechanical allodynia, which was maintained until postoperative day (POD) 30. An immediate single application of 2% QX-314 to the infraorbital nerve following CCI-ION significantly reduced neuropathic mechanical allodynia. Immediate double application of QX-314 produced a greater attenuation of mechanical allodynia than a single application of QX-314. Immediate double application of 2% QX-314 reduced the CCI-ION-induced upregulation of GFAP and p-p38 expression in the trigeminal ganglion. The upregulated p-p38 expression was co-localized with NeuN, a neuronal cell marker. We also investigated the role of voltage-gated sodium channels (Navs) in the antinociception produced by preemptive application of QX-314 through analysis of the changes in Nav expression in the trigeminal ganglion following CCI-ION. Preemptive application of QX-314 significantly reduced the upregulation of Nav1.3, 1.7, and 1.9 produced by CCI-ION. These results suggest that long-lasting blockade of the transmission of pain signaling inhibits the development of neuropathic pain through the regulation of Nav isoform expression in the trigeminal ganglion. Importantly, these results provide a potential preemptive therapeutic strategy for the treatment of neuropathic pain after nerve injury.

Participation of central GABAA receptors in the trigeminal processing of mechanical allodynia in rats

Here we investigated the central processing mechanisms of mechanical allodynia and found a direct excitatory link with low-threshold input to nociceptive neurons. Experiments were performed on male Sprague-Dawley rats weighing 230-280 g. Subcutaneous injection of interleukin 1 beta (IL-1β) (1 ng/10 µL) was used to produce mechanical allodynia and thermal hyperalgesia. Intracisternal administration of bicuculline, a gamma aminobutyric acid A (GABAA) receptor antagonist, produced mechanical allodynia in the orofacial area under normal conditions. However, intracisternal administration of bicuculline (50 ng) produced a paradoxical anti-allodynic effect under inflammatory pain conditions. Pretreatment with resiniferatoxin (RTX), which depletes capsaicin receptor protein in primary afferent fibers, did not alter the paradoxical anti-allodynic effects produced by the intracisternal injection of bicuculline. Intracisternal injection of bumetanide, an Na-K-Cl cotransporter (NKCC 1) inhibitor, reversed the IL-1β-induced mechanical allodynia. In the control group, application of GABA (100 µM) or muscimol (3 µM) led to membrane hyperpolarization in gramicidin perforated current clamp mode. However, in some neurons, application of GABA or muscimol led to membrane depolarization in the IL-1β-treated rats. These results suggest that some large myelinated Aβ fibers gain access to the nociceptive system and elicit pain sensation via GABA(A) receptors under inflammatory pain conditions.

The efficacy of combination treatment of gabapentin and electro-acupuncture on paclitaxel-induced neuropathic pain

Paclitaxel, a chemotherapeutic drug, induces severe peripheral neuropathy. Gabapentin (GBT) is a first line agent used to treat neuropathic pain, and its effect is mediated by spinal noradrenergic and muscarinic cholinergic receptors. Electro-acupuncture (EA) is used for treating various types of pain via its action through spinal opioidergic and noradrenergic receptors. Here, we investigated whether combined treatment of these two agents could exert a synergistic effect on paclitaxel-induced cold and mechanical allodynia, which were assessed by the acetone drop test and von Frey filament assay, respectively. Significant signs of allodynia were observed after four paclitaxel injections (a cumulative dose of 8 mg/kg, i.p.). GBT (3, 30, and 100 mg/kg, i.p.) or EA (ST36, Zusanli) alone produced dose-dependent anti-allodynic effects. The medium and highest doses of GBT (30 and 100 mg/kg) provided a strong analgesic effect, but they induced motor dysfunction in Rota-rod tests. On the contrary, the lowest dose of GBT (3 mg/kg) did not induce motor weakness, but it provided a brief analgesic effect. The combination of the lowest dose of GBT and EA resulted in a greater and longer effect, without inducing motor dysfunction. This effect on mechanical allodynia was blocked by spinal opioidergic (naloxone, 20 μg), or noradrenergic (idazoxan, 10 μg) receptor antagonist, whereas on cold allodynia, only opioidergic receptor antagonist blocked the effect. In conclusion, the combination of the lowest dose of GBT and EA has a robust and enduring analgesic action against paclitaxel-induced neuropathic pain, and it should be considered as an alternative treatment method.

Intravenous Administration of Substance P Attenuates Mechanical Allodynia Following Nerve Injury by Regulating Neuropathic Pain-Related Factors

This study aimed to investigate the analgesic effect of substance P (SP) in an animal model of neuropathic pain. An experimental model of neuropathic pain, the chronic constriction injury (CCI) model, was established using ICR mice. An intravenous (i.v.) injection of SP (1 nmole/kg) was administered to the mice to examine the analgesic effects of systemic SP on neuropathic pain. Behavioral testing and immunostaining was performed following treatment of the CCI model with SP. SP attenuated mechanical allodynia in a time-dependent manner, beginning at 1 h following administration, peaking at 1 day post-injection, and decaying by 3 days post-injection. The second injection of SP also increased the threshold of mechanical allodynia, with the effects peaking on day 1 and decaying by day 3. A reduction in phospho-ERK and glial fibrillary acidic protein (GFAP) accompanied the attenuation of mechanical allodynia. We have shown for the first time that i.v. administration of substance P attenuated mechanical allodynia in the maintenance phase of neuropathic pain using von Frey’s test, and simultaneously reduced levels of phospho-ERK and GFAP, which are representative biochemical markers of neuropathic pain. Importantly, glial cells in the dorsal horn of the spinal cord (L4–L5) of SP-treated CCI mice, expressed the anti-inflammatory cytokine, IL-10, which was not seen in vehicle saline-treated mice. Thus, i.v. administration of substance P may be beneficial for improving the treatment of patients with neuropathic pain, since it decreases the activity of nociceptive factors and increases the expression of anti-nociceptive factors.

Differential Role of Central GABA Receptors in Nociception of Orofacial Area in Rats

The present study investigated the role of central GABA(A) and GABA(B) receptors in orofacial pain in rats. Experiments were conducted on Sprague-Dawley rats weighing between 230 and 280 g. Intracisternal catheterization was performed for intracisternal injection, under ketamine anesthesia. Complete Freund's Adjuvant (CFA)-induced thermal hyperalgesia and inferior alveolar nerve injury-induced mechanical allodynia were employed as orofacial pain models. Intracisternal administration of bicuculline, a GABA(A) receptor antagonist, produced mechanical allodynia in naive rats, but not thermal hyperalgesia. However, CGP35348, a GABA(B) receptor antagonist, did not show any pain behavior in naive rats. Intracisternal administration of muscimol, a GABA(A) receptor agonist, attenuated the thermal hyperalgesia and mechanical allodynia in rats with CFA treatment and inferior alveolar nerve injury, respectively. On the contrary, intracisternal administration of bicuculline also attenuated the mechanical allodynia in rats with inferior alveolar nerve injury. Intracisternal administration of baclofen, a GABA(B) receptor agonist, attenuated the thermal hyperalgesia and mechanical allodynia in rats with CFA treatment and inferior alveolar nerve injury, respectively. In contrast to GABA(A) receptor antagonist, intracisternal administration of CGP35348 did not affect either the thermal hyperalgesia or mechanical allodynia. Our current findings suggest that the GABA(A) receptor, but not the GABA(B) receptor, participates in pain processing under normal conditions. Intracisternal administration of GABA(A) receptor antagonist, but not GABA(B) receptor antagonist, produces paradoxical antinociception under pain conditions. These results suggest that central GABA has differential roles in the processing of orofacial pain, and the blockade of GABA(A) receptor provides new therapeutic targets for the treatment of chronic pain.

Mechanical Antiallodynic Effect of Intrathecal Nefopam in a Rat Neuropathic Pain Model

Nefopam has a pharmacologic profile distinct from that of opioids or other anti-inflammatory drugs. Several recent studies demonstrate that nefopam has a mechanism of action similar to those of anti-depressants and anticonvulsants for treating neuropathic pain. The present study investigates the mechanical antiallodynic effect of nefopam using immunohistochemical study and western blot analysis in a rat neuropathic pain model. Twenty-eight male Sprague-Dawley rats were subjected to left fifth lumbar (L5) spinal nerve ligation and intrathecal catheter implantation, procedures which were not performed on the 7 male Sprague-Dawley rats in the sham surgery group (group S). Nefopam, either 10 or 100 microg/kg (group N10 or N100, respectively), and normal saline (group C) were intrathecally administered into the catheter every day for 14 days. The mechanical allodynic threshold of intrathecal nefopam was measured using a dynamic plantar aesthesiometer. Immunohistochemistry targeting cluster of differentiation molecule 11b (CD11b) and glial fibrillary acidic protein (GFAP) was performed on the harvested spinal cord at the level of L5. Extracellular signal-regulated kinase 1/2 (ERK 1/2) and cyclic adenosine monophosphate response element binding protein (CREB) were measured using western blot analysis. The N10 and N100 groups showed improved mechanical allodynic threshold, reduced CD11b and GFAP expression, and attenuated ERK 1/2 and CREB in the affected L5 spinal cord. In conclusion, intrathecal nefopam reduced mechanical allodynia in a rat neuropathic pain model. Its mechanical antiallodynic effect is associated with inhibition of glial activation and suppression of the transcription factors' mitogen-activated protein kinases in the spinal cord.

Olanzapine Attenuates Mechanical Allodynia in a Rat Model of Partial Sciatic Nerve Ligation

BACKGROUND: Neuropathic pain is a global clinical problem; nevertheless, nerve injury treatment methods remain limited. Olanzapine has antinociceptive and anti-nueropathic properties; however, its preventive effects have not been assessed in nerve injury models. METHODS: We prepared a partial sciatic nerve ligation (Seltzer model) or sham-operated model in male Sprague-Dawley rats under isoflurane anesthesia. In a pre-treatment study, we administered olanzapine (10 mg/kg) intraperitoneally 1 h before nerve ligation. In post-treatment and dose-dependent studies, we injected 3 different doses of olanzapine intraperitoneally 1 h after nerve ligation. Mechanical allodynia was measured before and 7 days after surgery. Immunohistochemical analysis using anti-Iba-1 antibody was used to assess the effect of olanzapine at the spinal level. RESULTS: In the pre-treatment study, median withdrawal thresholds of the normal saline groups were significantly lower than those of the sham-operated groups; however, those of the olanzapine (10 mg/kg) and sham-operated groups were not different. In the post-treatment and dose-dependent studies, the median withdrawal thresholds of the olanzapine (2.5 mg/kg) and normal saline groups were not different; however, those of the olanzapine (10 and 50 mg/kg) groups were significantly higher than those of the normal saline groups. Olanzapine did not have a significant effect on the density of Iba-1 staining. CONCLUSIONS: Olanzapine attenuated mechanical allodynia dose-dependently in the Seltzer model. This anti-allodynic effect of olanzapine was observed even when injected 1 h after nerve ligation. This effect of olanzapine appeared to be unrelated to microglia activation in the ipsilateral dorsal horn of the lumbar spinal cord.

Spatio-temporal expression of P2X3 receptor in rats with diabetic mechanical allodynia / 解剖学报

Objective To investigate the spatio-temporal expression of P2X3 receptor ( P2X3R) in rats with diabetic mechanical allodynia ( DMA ) .Methods DMA model in rats was induced by intraperitoneal injection of streptozocin ( STZ) .The von Frey filaments were applied to identify the changes of the paw withdrawal threshold ( PWT) in DMA rats.Immunofluorescence was employed to detect the spatio-temporal expression of P2X3R in the spinal dorsal horn (SDH), dorsal root ganglion (DRG) and hind paw skin on different time points after intraperitoneal injection of STZ , respectively.The protein expression of P2X3R in SDH and DRG was further semi-quantitatively analyzed by Western blotting.Results Compared with control group , intraperitoneal injection STZ induced significant mechanical allodynia indicated by the reduced PWT from 7 days, and which reached the peak on 14d and maintained to 28days (P<0.05). The expression of P2X3R in DRG neurons was significantly increased on 14 days and 21 days (P<0.05), while that in SDH and skin was markedly increased on 21 days and 28 days, compared with control group (P<0.05).Conclusion With the progress of DMA, the expression of P2X3R was significantly increased in the SDH, DRG and skin, which was almost parallel with the mechanical allodynia , but the changes in SDH and skin were 1 week later than that in DRG .These results suggest that P2X3R may play a key role in the maintenance of the DMA .

Effect of superoxide on the development and maintenance of mechanical allodynia in a rat model of chronic post-ischemia pain / 대한마취과학회지

BACKGROUND: Reactive oxygen species and inflammatory responses contribute to the development of neuropathic pain. Superoxide serves to mediate cell signaling processes and tissue injury during inflammation. We examined the effects of superoxide on the development and maintenance of mechanical allodynia, as well as its contribution to central sensitization in a superoxide-rich animal model of neuropathic pain. METHODS: Chronic post-ischemia pain (CPIP) was induced via the left hindpaw ischemia for 3 h, followed by reperfusion. Superoxide dismutase (4,000 U/kg, i.p.) was administered either 5 min before ischemia (BI), 5 min before reperfusion (BR), or 3 days after reperfusion (3AR). Withdrawal thresholds of the four paws were measured to assess the mechanical allodynia and the effects of circulating xanthine oxidase (XO)-mediated superoxide production. In addition, we measured the levels of N-methyl D-aspartate receptor subunit 1 phosphorylation (p-NR1) in the ipsilateral and contralateral spinal cord (L4-6), by Western blotting, to examine the superoxide-mediated central sensitization. Superoxide production was assessed by allopurinol-sensitive, XO-mediated lipid peroxidation of the spinal cord and gastrocnemius muscles. RESULTS: Withdrawal thresholds of forepaws did not vary across the 7 days of testing. In the hindpaws, both ipsilateral and contralateral mechanical allodynia was most attenuated in the BR group, followed by the BI and 3AR groups. The degree of NR1 activation was in contrast to the changes in the withdrawal thresholds. CONCLUSIONS: These data suggest that superoxide is involved in the development and maintenance of mechanical allodynia, particularly via central sensitization in the spinal cord.

Gender differences in paclitaxel-induced neuropathic pain behavior and analgesic response in rats / 대한마취과학회지

BACKGROUND: Females show greater sensitivity than males to several modalities of experimental pain. However, the gender differences in paclitaxel-induced neuropathic pain have not been studied. The current study examined the gender differences in neuropathic pain behavior and the effect of analgesics in a paclitaxel-induced neuropathic pain model in rats. METHODS: Neuropathic pain was induced by intraperitoneal injection of paclitaxel (2 mg/kg) on 4 alternate days in Sprague-Dawley rats of both genders. Mechanical allodynia was measured using a von Frey filament. The gender differences in analgesic responses were determined after administration of morphine (2 or 5 mg/kg), ketamine (2 or 5 mg/kg), or combined morphine (2 mg/kg) and ketamine (2 mg/kg). RESULTS: Paclitaxel induced mechanical allodynia, which began to manifest on day 4, peaked within 10 days, and plateaued for at least 2 months after the first paclitaxel injection. No gender difference in the manifestation of mechanical allodynia was observed. A 2 mg/kg dose of ketamine increased the mechanical threshold only in males. The 5 mg/kg dose of ketamine significantly increased the mechanical threshold in both genders. Morphine (2 and 5 mg/kg) dose-dependently increased the mechanical thresholds in both genders. The 2 mg/kg dose of ketamine enhanced the antinociceptive effect of 2 mg/kg morphine only in females. CONCLUSIONS: No gender difference in paclitaxel-induced neuropathic pain or analgesic response to ketamine or morphine was observed in Sprague-Dawley rats. Low dose ketamine enhanced the analgesic effect of morphine on paclitaxel-induced mechanical allodynia but only in female rats.

The Long-Lasting Pattern of the Neuropathic Radicular Pain in An Autologous Nucleus Pulposus Model of Rat

OBJECTIVE: To observe the long-lasting changes of pain progression with time course in an autologous nucleus pulposus model of rat. METHOD: The subjects were 25 Sprague-Dawley (Sprague- Dawley, 250 gm) male rats. They were randomly assigned into either the sham or experimental group. In the experimental group (n=15), autologous nucleus pulposus was harvested from the coccygeal intervertebral disc of the rat and this was grafted on the left L5 dorsal root ganglion. In the sham group (n=10), the left L4 and L5 nerve roots were exposed by laminectomy, but the nucleus pulposus was not grafted. All the rats were evaluated for mechanical allodynia and thermal hyperalgesia at 2 days before surgery, and on days 1, 5, 10, 20, 30, 40 and 50 after surgery. The morphological changes of the spinal nerves were assessed by toluidine blue staining on days 5 after surgery. RESULTS: In the ipsilateral hindpaw of the experimental group, there was a dramatic drop of the mechanical withdrawal threshold and the thermal withdrawal latency on day 1 after surgery, which was maintained at day 50 after surgery. In morphological study, pathological findings such as swelling of the myelin sheath, demyelination, swelling and degeneration of the axoplasm were observed in the spinal nerve at day 5 after surgery. CONCLUSION: The long-lasting pattern of neuropathic radicular pain shown in a rat model of lumbar disc herniations is helpful to understand the natural history of neuropathic radicular pain due to ruptured nucleus pulposus.