C纤维“传导丢峰”调控糖尿病神经病理痛信号的低阈值持续钾通道机制研究

Recentlly numerous studies have accumulated that unmyelinated fibers (i.e, C fibers) play a crucial role in the conduction of pain signals and sensitization of which following injury or inflammation leads to marked hyperalgesia. However, how the C fibers play a role in operation and process of information underlying hyperalgesia have remained elusive. Our previous study found that reduced conduction failure of C-fibers contributes to painful diabetic neuropathy in rat and decrease in potassium channel expression involves in the reduction of conduction failure of C fibers. Moreover, our preliminary data demonstrated that α-DTX, a blocker of low threshold sustained potassium channels can inhibit the conduction failure reversibly in the main axon of C-fibers. It suggest that this potassium channel may play an importatnt role in painful diabetic neuropathy regulated by the conduction failure of C-fibers. To test this assumption, we will utilize a combination of single fiber recording in vivo and patch clamp techinique to exaimne the functional change in the degree of conduction failure of coccygeal C-fibers and the density of low threshold suatained potassium currents in small dorsal root ganglion neurons. Moreover, we will use immunostaining methods and biochemical assays to investigate the difference in the expression and distribution of this potassium channels subunits between the control and diabetic C-fibers and DRGs.To further study the effect of change in conduction failure of C-fibers underlying hyperalgesia, we will use behvioral pharmacological experiments to estimate the effect of changes in the function of this potassium channel in the main axon of C-fibers on the animal hyperalgesia. This study might elucidate the potassium channel mechanism for the conduction failure of polymodal nociceptive C-fibers and reveal a novel factor for regulating hyperalgesia. It may be helpful for the identification of a new target for the selective treatment of chronic pain signal on the peripheral axon of C-fibers.

近年研究表明，无髓鞘神经轴突（C纤维）在炎症或损伤导致的痛敏情况下痛信号的传导过程起重要作用，然而C纤维究竟如何发挥信息的加工和处理功能，尚不清楚。我们前期研究发现C纤维传导丢峰的减弱与糖尿病神经病理痛的形成密切相关[Brain,2012]，并且低阈值持续钾通道的阻断可以可逆性地抑制C纤维的这一传导丢峰现象，提示该钾通道可能在C纤维通过传导丢峰调控痛信号的传入从而导致外周痛敏过程发挥重要作用。为了论证这一设想，本项目拟采用在体单纤维和膜片钳记录从功能上检测糖尿病后尾神经C纤维和背根节小细胞上传导丢峰程度的变化及其通道电流机制，同时检测C纤维和背根神经节上该钾通道亚型分布和表达的变化，并结合行为药理学实验对该钾通道功能的调变对动物痛敏行为的影响作出判断。研究可望阐明C纤维传导丢峰发生的特异钾通道机制，确认一个外周痛信号调控的新环节，有助于在外周C纤维轴突上识别选择性抑制慢性痛信号的新靶位。

无髓鞘神经轴突（C纤维）的主要功能是感受与传导痛觉信息。然而，在炎症或损伤导致的病理情况下动物痛敏行为的形成过程中，传入痛信号的C纤维究竟发生了哪些功能变化，尚不清楚。近年，我们在研究C纤维的传导丢峰(conduction failure)和传导编码(conduction coding)等活动规律时，发现C纤维传导丢峰的减弱与糖尿病神经病理痛的形成密切相关，提示C纤维的活动变化可能通过调控痛信号的传导，参与影响痛敏行为的形成。进一步的研究还发现阻断神经轴突上一种低阈值的持续钾通道（α-DTX敏感K通道）可以可逆性地翻转这一传导丢峰现象，那么该钾通道是否可能在C纤维通过传导丢峰调控痛信号的传入，从而导致外周痛敏过程中发挥重要作用呢？为此，本项目分别采用大鼠和转基因小鼠，综合运用电生理学、分子生物学、形态学和行为药理学等方法开展了以下研究：（1）糖尿病或炎症状态下，C纤维的传导功能及活动规律发生了哪些变化？（2）介导C纤维传导丢峰调变的离子通道机制；（3）C纤维传导丢峰的调变与动物痛敏行为之间的关系。取得的重要结果有：（1）糖尿病或炎症状态下，C纤维的传导丢峰程度均明显减弱；（2）低阈值持续钾通道阻断剂α-DTX可以显著减弱C纤维的传导丢峰，并且糖尿病状态下背根节神经节（DRG）小细胞上IDTX明显减小，进一步研究发现DRG小细胞和外周C纤维上Kv1通道亚型（Kv1.2和Kv1.6）分布和表达均明显下调；（3）HCN通道阻断剂ZD7288可以显著增强C纤维的传导丢峰，并且炎症状态下DRG小细胞上Ih明显增强，进一步研究发现DRG小细胞和外周C纤维上HCN通道亚型（HCN2）的分布和表达均明显上调；（4）外周坐骨神经干周围注射ZD7288明显减弱了动物的机械触痛和热痛过敏行为但不影响运动和心血管功能。该研究揭示了C纤维传导丢峰的变化在痛敏形成过程中的作用及其活动规律，阐明了C纤维传导丢峰的特异K通道和HCN通道机制。确认了一个外周痛信号调控的新环节，为研发在外周C纤维轴突上选择性抑制慢性痛的治疗药物提供新的特异靶位。