中脑导水管周围灰质OrexinA及其受体OX1R通过大麻素受体1参与针刺缓解内脏痛敏的机制研究

Central sensitization is a key factor in the development of visceral hypersensitivity of irritable bowel syndrome (IBS). Acupuncture mainly activates descending inhibitory pathway to relieve visceral hypersensitivity. Studies show that orexinA is a pivotal peptide that plays an anti-nociceptive role in pain regulation both at spinal and supra-spinal levels, and periaqueductal gray matter (PAG, the major section of descending inhibitory pathway) is the key region where orexinA exerts the anti-nociceptive effect on. Our previous research found that orexinA is involved in acupuncture-induced analgesia through its receptor OX1R to activate descending inhibitory pathway. Thus, the present study is employing behavior tests, neuropharmacological, molecular biological and electrophysiological measures etc, 1) to verify that acupuncture produces the anti-central sensitization effect through regulating PAG orexinA and OX1R; 2) to illuminate that orexinA and OX1R participate in acupuncture-induced analgesia in visceral hypersensitivity through activating CB1; and 3) to further explore that presynaptic inhibition produced by OX1R-CB1 activation is the molecular mechanism of the orexinA involvement in acupuncture-induced analgesia. Results of the study can be expected to provide theoretical and scientific evidence in acupuncture regulation of descending inhibitory pathway to inhibit central sensitization.

中枢敏化是肠易激综合征（IBS）内脏痛敏发生过程中的关键因素，针刺主要通过激活下行抑制系统抑制内脏痛敏。研究显示OrexinA是一种在脊髓及脊髓上水平疼痛调控中具有重要作用的神经肽，而导水管周围灰质（PAG，下行抑制系统关键环节）是OrexinA发挥脊髓上水平抗伤害作用的重要脑区。结合申请人前期工作基础即OrexinA通过其受体OX1R参与针刺对于下行抑制系统的激活效应，本研究拟采用行为学、神经药理学、分子生物学及神经电生理等技术，开展如下研究：1)证实电针通过调控中脑vlPAG的OrexinA及其受体发挥抗中枢敏化效应；2)阐明中脑PAG的OrexinA及OX1R通过激活下游大麻素受体（CB）1参与电针缓解内脏痛敏效应；3)深入阐释OX1R-CB1通路引起的突触前抑制是OrexinA参与电针镇痛的关键分子机制，以期为临床针刺调控下行抑制系统从而抑制中枢敏化提供理论基础和科学依据。

制备IBS内脏痛敏模型并采用行为学、病理观察等方法评估电针的镇痛效应，测定vlPAG中OrexinA mRNA，OX1R mRNA的表达及OXA，CB和GABA的含量。结果显示电针对模型大鼠具有一定镇痛效应，与模型组和假针组均有统计学差异，正常组、模型组、电针组和假针组中脑PAG的OrexinA mRNA及OX1R mRNA没有统计学差异，OXA，CB及GABA含量均没有统计学差异。于是尝试建立另一种内脏痛模型，环磷酰胺（CYP）腹腔注射制备内脏痛敏模型，分别尝试了75mg/kg腹腔注射一次，150mg/kg腹腔注射一次，300mg/kg腹腔注射一次及75mg/kg腹腔注射每天一次、共四次，采用von Frey和病理组织学的方法进行评估不同剂量CYP注射后机械性痛阈和膀胱粘膜变化，选定150mg/kg CYP一次注射与75mg/kg连续注射四天均可作为膀胱痛的模型。采用行为学和病理学观察方法进行电针效应评估，结果D5天起电针对M75*4组及M150组大鼠腹部机械性痛觉过敏状况有明显缓解，并可不同程度缓解膀胱上皮粘膜损伤。该模型大鼠vlPAG中OrexinA mRNA和OX1R及OXA含量测定显示，正常组、模型组、电针组和假针组没有统计学差异。结论为电针对两种模型内脏痛有一定镇痛效应，但vlPAG OrexinA及其受体在这个过程中是否为最为关键的因素有待进一步探讨。