星形胶质细胞缝隙连接蛋白Cx43在PSNL引起的神经病理性疼痛中的作用及机制

Accumulating evidence suggests that glial cells play an important role in the development and maintenance of neuropathic pain. However, the detail mechanism remains unclear. Astrocytes possess the largest gap junctions composed of Connexin43. Our study and others have showed that inhibition of gap junction significantly suppressed neuropathic pain. The fact that ATP is one of the vital signals involved in neuropathic pain and that ATP can be regulated by Cx43 channels lead us to postulate that astrocyte Cx43 is involved in the development of neuropathic pain via regulating ATP. In support of this hypothesis, we construct neuropathic pain rat model caused by partial sciatic nerve ligation (PSNL) and observe the alteration of Cx43 and channels in the astrocytes of the spinal cord dorsal horn. To examine the influence of Cx43 on ATP release and neuropathic pain, Cx43 knockout mice will be used in this study in combination with the manipulation of Cx43 channel by chemicals and molecular biological methods in wide type. Moreover, whether or not and how ATP mediate the effect of Cx43 channel on neuropathic pain will be investigated in vitro and in vivo rat models. Finally, the effects of commonly-used analgesics ( morphine and tramadol) on Cx43 channel and ATP release, and the role of Cx43 in analgesics-induced acitivity on neuropathic pain will be assessed. Our study is aimed to determine the role of Cx43 in neuropathic pain, thereby explore their potential role as a preventive and therapeutic target for pharmacological and biochemical strategies against neuropathic pain.

胶质细胞在神经病理性疼痛的发生和维持中具有重要作用，但机理未明。星形胶质细胞间富含缝隙连接蛋白Cx43组成的连接通道（gap junction,GJ）。最近研究（包括我们研究）表明：抑制GJ能抑制病理性疼痛。结合Cx43通道能调节ATP以及ATP是神经病理性疼痛发生和维持中的重要分子，我们提出：星形胶质细胞Cx43可能通过调节ATP在神经病理性疼痛中发挥重要作用。本研究拟在坐骨神经部分损伤（PSNL）病理性疼痛大鼠模型上观察脊髓背角星形胶质细胞Cx43及所组成通道的变化；应用分子生物学方法改变Cx43通道功能探讨Cx43对ATP释放和病理性疼痛的影响，在体内外模型上分析ATP在Cx43影响病理性疼痛中的作用；在此基础上观察曲马多和吗啡对星形胶质细胞Cx43及ATP释放的影响，探讨Cx43在椎管内镇痛中的作用。本研究旨在阐明Cx43在神经病理性疼痛中的作用及机理，为其防治和药物开发提供新思路

神经病理性疼痛是临床上常见的难治的慢性疼痛，严重危害患者的生存质量，临床上缺乏满意的治疗方法。神经胶质细胞对神经元不仅起支持和营养作用，在慢性疼痛的产生和维持中具有重要作用，其中广泛存在于脊髓的星形胶质细胞被认为在病理性疼痛的长期维持阶段具有主动和不可替代的作用，但其机理不明。 我们的研究发现，相较于正常组大鼠，SNI引起的神经病理性疼痛大鼠同侧机械缩足阈值和热刺激回缩潜伏期减低，脊髓背角Cx43表达明显增加，而应用Cx组成的缝隙连接通道抑制剂carbenoxolone和半通道抑制剂INI-0602后，Cx组成的通道功能明显降低，大鼠机械缩足阈值和热刺激回缩潜伏期增加。这说明抑制Cx通道功能，能够明显减弱神经病理性疼痛的痛觉敏感。同时我们还发现，相较于正常组，神经病理性疼痛大鼠脊髓背角星形胶质细胞标志性蛋白GFAP和小胶质细胞标志性蛋白Iba-1蛋白明显增加，同时伴随着脑脊液ATP释放和炎症因子IL-6，IL-1β和TNF-α的表达增加，神经元NMDA受体Nr2b表达明显增强；而鞘内注射Cx通道抑制剂后，SNI神经病理性疼痛大鼠的脑脊液ATP和炎症因子IL-6，IL-1β和TNF-α的表达明显下降，标志性蛋白GFAP和小胶质细胞标志性蛋白Iba-1蛋白表达下降，同时神经元Nr2b受体及磷酸化表达受抑制。我们还通过动物研究发现，镇痛药曲马多能够抑制SNI神经病理性疼痛大鼠的机械痛，抑制脊髓背角Cx43蛋白的表达和星形胶质细胞GFAP的表达。我们的研究结果表明，Cx43介导的信号通路参与介导星形胶质细胞的激活和炎症因子释放，从而调控脊髓神经元的兴奋性，在SNI引起的神经病理性疼痛中发挥作用。非阿片类镇痛药曲马多能够显著抑制SNI引起的神经病理性疼痛，该抑制作用与其抑制Cx43及其通道功能及星形胶质细胞的激活有关。