基于背根神经节 P2X3受体调控的电针干预糖尿病神经痛作用研究

Diabetic neuropathic pain (DNP) is one of the most common complications of diabetes. It is characterized by spontaneous burning pain and hyperalgesia, and severely affects the patients' life quality. Currently, there is no satisfied drug treatment. Acupuncture including electroacupuncture (EA) is now gradually being applied to treat DNP, with a phenomenon that the effect of EA with low frequency is better than that of EA with high frequency. Studies show that dorsal root ganglion (DRG) P2X3 receptor plays an important role in DNP, which is regulated by PKC. The project is to further determine the superiority of EA with low frequency in treating DNP by comparing the effects of EA with high frequency and low frequency. P2X3-mediated fast-inactivating current of DRG neuron, DRG P2X3 total protein level and its expression in cell membrane are measured to evaluate the effects of EA with low frequency on DRG P2X3 function, expression and its shift from cytoplasm to cell membrane. In addition, the regulation of EA with low frequency of DRG PKC, and the effects of P2X3 and PKC agonists on the action of EA with low frequency are also studied, so as to explore the mechanism of DRG P2X3 regulation through PKC pathway by EA with low frequency in the treatment of DNP, which will provide scientific evidence and explanation for DNP treatment by EA and its frequency selection, to promote EA more widely and more effectively treating DNP.

糖尿病神经痛(DNP)是糖尿病最常见的并发症之一，主要表现为自发性烧灼样疼痛、痛觉敏化，严重影响患者生存质量。目前药物治疗效果不佳。针灸(包括电针)已逐步用于治疗DNP，初步发现电针治疗DNP存在低频作用好于高频的现象。研究表明背根神经节(DRG) P2X3受体在DNP中起着重要作用，并受PKC调制。本项目拟通过高低频率电针效应对比，进一步明确低频电针干预DNP的优势作用；在此基础上，检测DRG神经元P2X3受体介导的快反应电流强度，和P2X3受体总蛋白和细胞膜表达水平，观察低频电针对DRG P2X3受体功能、表达和从胞质向胞膜转移的影响，结合低频电针对DRG PKC的调节，以及P2X3受体、PKC激动剂对低频电针作用的影响，阐明低频电针干预DNP的DRG P2X3受体调控机制及其通过PKC的干预途径，为电针治疗DNP及其频率选择提供科学依据和阐释，以期推动电针更广泛、更好地治疗DNP。

糖尿病神经痛(DNP)是糖尿病最常见的并发症之一，主要表现为自发性烧灼样疼痛、痛觉敏化，严重影响患者生存质量。针灸(包括电针)已逐步用于治疗DNP，初步发现电针治疗DNP存在低频作用好于高频的现象，但有待进一步明确。研究表明背根神经节(DRG) P2X3受体在DNP中起着重要作用，并受PKC调制。本项目拟通过高低频率电针效应对比，进一步明确低频电针干预DNP的优势作用；在此基础上，检测DRG神经元P2X3受体总蛋白和细胞膜表达水平，观察低频电针对DRG P2X3受体功能、表达和从胞质向胞膜转移的影响，结合低频电针对DRG PKC的调节，以及P2X3受体、PKC激动剂对低频电针作用的影响。主要研究结果分三部分论述：第一部分主要研究结果如下：不同频率EA均可改善DNP大鼠神经痛，且低频EA的镇痛作用优于高频。不同频率EA对DNP DRG神经元CGRP表达均有抑制作用，不同程度上抑制P2X3受体表达，其中低频EA对L4 DRG神经元P2X3受体表达的调控作用优于高频。第二部分主要研究结果如下：免疫印迹法结果显示：各组大鼠的P2X3受体总蛋白水平无明显差异。而与Normal组比较，DNP组大鼠L4-L6 DRG P2X3受体膜蛋白表达均明显升高；与DNP组比较，DNP + EA组大鼠L4-L6 DRG P2X3受体膜蛋白表达均明显下降；免疫荧光法结果显示：与Normal组比较，DNP组大鼠L4-L6 DRG p-PKC阳性细胞率明显升高；与DNP组比较，DNP + EA组大鼠L4-L6 DRG p-PKC阳性细胞率均明显下降。第三部分主要研究结果如下：与EA + PBS组比较，EA + αβ-meATP组和EA + PMA组双侧PWTs均明显降低；免疫印迹法结果显示：各组大鼠的P2X3受体总蛋白水平无明显差异。而与EA + PBS组比较，注射PKC激动剂PMA使EA + PMA组大鼠L4-L6 DRG P2X3受体膜蛋白表达明显升高。通过本项目研究阐明低频电针改善糖尿病神经痛可能与其抑制PKC的活化，进而调控L4-L6 DRG P2X3受体的膜转位有关，为电针治疗DNP及其频率选择提供科学依据和阐释，以期推动电针更广泛、更好地治疗DNP。