中脑导水管周围灰质腹外侧区P2X7受体在曲马多减轻大鼠神经病理性痛中的作用

摘要

Objective To evaluate the role of P2X7 receptor in the ventrolateral periaqueductal gray (vlPAG) in tramadol-induced reduction of neuropathic pain (NP) in rats.Methods Fifty-four male clean-grade healthy Sprague-Dawley rats,aged 7 days,weighing 190-230 g,were studied.NP was induced by chronic constrictive injury (CCI) to sciatic nerve.Experiment Ⅰ Thirty-six rats were divided into 3 groups (n =12 each) using a random number table method:sham operation group (group S),group NP1 and NP plus tramadol group (group NP1 +T).Tramadol 15 mg/kg was intraperitoneally injected once a day from day 7 to day 14 after CCI in group NP1+T.The mechanical and thermal pain thresholds of the nerve-injured hindlimb were measured before CCI and on 1,5,7,10,12 and 14 days after CCI.Rats were sacrificed after measurement of pain threshold on day 14 after CCI,and the expression of P2X7 receptor in vlPAG was detected by immunohistochemistry and Western blot assay.Experiment Ⅱ Eighteen rats were divided into 3 groups (n =6 each) using a random number table method:group NP2,NP plus tramadol group (group NP2+T) and NP plus tramadol plus a specific P2X7 receptor antagonist A-438079 group (group NP2+T+A).In NP2+T+A group,a catheter was implanted in vlPAG,and the NP model was established on 5th day after successful catheterization.Tramadol 15 mg/kg was intraperitoneally injected once a day from day 7 to day 14 after CCI in group NP2+T.In group NP2+T+A,tramadol 15 mg/kg was intraperitoneally injected once a day from day 7 to day 14 after CCI,followed by a microinjection of A-438079 100 pmol (0.3 μl) via vlPAG before giving tramadol on day 14.The mechanical and thermal pain thresholds were measured at the end of the last tramadol administration and within 1 h after the end of the last tramadol administration.Results Experiment Ⅰ Compared with group S,the mechanical and thermal pain thresholds were significantly decreased at each time point after CCI,the number of P2X7 receptor positive cells was increased,and the expression of P2X7 receptor was up-regulated in the other two groups (P＜0.01).Compared with group NP1,the mechanical and thermal pain thresholds were significantly increased at days 7-14 after CCI,the number of P2X7 receptor positive cells was increased,and the expression of P2X7 receptor was up-regulated in group NP1 +T (P＜0.01).Experiment Ⅱ Compared with group NP2,the mechanical and thermal pain threshold were significantly increased at each time point after CCI in NP2+T and NP2 +T+A groups (P＜0.01).Compared with group NP2 +T,the mechanical and thermal pain thresholds were significantly decreased at each time point after CCI in group NP2+T+A (P＜ 0.01).Conclusion The mechanism by which tramadol mitigates NP is partially related to enhanced function of P2X7 receptors in vlPAG of rats.%目的 评价中脑导水管周围灰质腹外侧区(vlPAG) P2X7受体在曲马多减轻大鼠神经病理性痛中的作用.方法 清洁级健康雄性SD大鼠54只,体重190～230 g,7周龄,采用慢性坐骨神经压迫性损伤(CCI)法制备大鼠神经病理性痛模型.实验Ⅰ 取大鼠36只,采用随机数字表法分成3组(n=12):假手术组(S组)、神经病理性痛组(NP1组)和神经病理性痛+曲马多组(NP1+T组).NP1+T组于CCI后第7-14天腹腔注射曲马多15 mg/kg,1次/d.于CCI前、CCI后1、5、7、10、12、14 d时测定机械痛阈与热痛阈.于CCI后14 d痛阈测定结束后处死大鼠,采用免疫组化法和Westernblot法测定vlPAG P2X7受体表达水平.实验Ⅱ 取大鼠18只,采用随机数字表法分成3组(n=6):神经病理性痛组(NP2组)、神经病理性痛+曲马多组(NP2+T组)和神经病理性痛+曲马多+P2X7受体特异性拮抗剂A-438079组(NP2+T+A组).NP2 +T+A组行vlPAG置管,置管成功后第5天建立神经病理性痛模型.NP2+T组于CCI后第7-14天腹腔注射曲马多15 mg/kg,1次/d;NP2 +T+A组于CCI后第7-14天腹腔注射曲马多15 mg/kg,1次/d,第14天曲马多给药前经vlPAG微量注射A-438079100 pmol(0.3μl).于曲马多最后一次给药结束即刻、结束后1h内每10 min测定机械痛阈与热痛阈.结果 实验Ⅰ与S组比较,其余2组各时点机械痛阈和热痛阈降低,vlPAG P2X7受体阳性细胞数增多,表达上调(P＜0.01);与NP1组比较,NP1+T组CCI后7-14 d机械痛阈和热痛阈升高,vlPAGP2X7受体阳性细胞数增多,表达上调(P＜0.01).实验Ⅱ 与NP2组比较,NP2 +T组和NP2+T+A组各时点热痛阈和机械痛阈升高(P＜0.01);与NP2+T组比较,NP2+T+A组各时点热痛阈和机械痛阈降低(P＜0.01).结论 曲马多减轻大鼠神经病理性痛的部分机制与vlPAG P2X7受体功能增强有关.