钾通道在糖尿病神经性疼痛发生中的作用及机制研究

Decreased expression or functional inhibiton of KCNQ potassium channel leads to increased neuronal excitability , accordingly causing excitable diseases such as epilepsy and pain.Activation of KCNQ potassium channels decreased neuronal excitability, which might be a potential drug target for pain releaf. KCNQ potassium channels are often studied in neuron-injury model with decreased expression and function. However it is still not known how KCNQ potassium channels are altered and modulated during the development of diaabetic neuropathy. Here we propose to apply both type 1 diebetes model (streptozotocin(STZ)-induced)and type 2(db/db mice) model to generate diabetic neuropathy model in combination with the following approaches, dorsal root ganglion (DRG) neuronal isolation and culture, whole-cell patch clamp recording, molecular biology and cell biology, to study the role of potassium channels in the disease model. We mainly study the time course of expression and function for DRG neuronal KCNQ channel and the corresponding time course of pharmacological response to the newly indentifed KCNQ2 activator ML213 (published). Further determine the modulation mechanism of channel expression and dysfunction. It would be possible to build the basis for confirmation of the studied channels as drug targets and drug development for diabetic neuropathy treatment.

神经系统表达的KCNQ钾通道表达降低或通道被抑制，神经元兴奋性升高，导致疼痛或癫痫等兴奋性疾病的发生，通道开放剂可有效降低神经元兴奋性，可能是治疗疼痛的有效药物。KCNQ2钾通道多被研究于神经损伤的疼痛模型，表现为表达和功能下降，但在糖尿病神经性疼痛病理性改变过程中是如何改变和被调控的，尚未见报道。本项目拟应用1型(STZ诱发) DN模型，联合采用DRG神经元分离培养、全细胞膜片钳、分子生物学等方法研究DN病理过程中DRG神经元KCNQ2钾通道表达和功能的时间动态关系，以及应用我们发现的KCNQ开放剂ML213（已发表）研究通道活性-药效学动态关系；在此基础上，通过分子生物学和电生理等方法和技术，研究通道表达和功能异常的调控机制，为确证KCNQ2钾通道作为DN药物靶点的可能性以及为抗糖尿病神经性疼痛药物的研发奠定科学的理论基础。

KCNQ钾通道主要表达于外周神经系统的背根神经节神经元和中枢神经系统的海马神经元，负责控制神经元的静息膜电位，对于神经元兴奋性的维持具有重要的作用，是治疗癫痫和疼痛的重要治疗靶点。该通道在多种疼痛模型，表现为表达和功能下降，但在糖尿病神经性疼痛中是如何改变和被调控的，尚未见报道。本研究的目的是探索KCNQ钾离子通道在糖尿病神经性疼痛发生中的机制，最终确证KCNQ钾通道作为糖尿病疼痛治疗靶点的可行性。应用链脲霉素（STZ）诱发的大鼠糖尿病神经性疼痛模型（Diabetic neuropathic pain, DPN），发现DPN可显著诱发大鼠背根神经节神经元（DRG）钾离子通道KCNQ2和KCNQ3表达的下降，KCNQ钾通道开放剂瑞替加滨（Retigabine）可显著缓解糖尿病疼痛的症状。在STZ诱发的糖尿病模型中，糖尿病糖基代谢产物甲基乙二醛（Methylglyoxal，MG）的血浆水平显著上升；MG腹腔注射后，不但可诱发大鼠脚掌对疼痛的刺激阈值显著下降，而且还可诱发体内和体外培养的DRG神经元KCNQ通道表达均下降。进一步的药理学实验表明，醛基清除剂氨基胍（Aminoguanidine, AG）可显著缓解STZ诱发的糖尿病神经痛行为，对KCNQ2和KCNQ3的mRNA水平无显著作用，但是可逆转KCNQ2蛋白表达的下降。所以，本研究证实了KCNQ钾通道可作为治疗糖尿病神经性疼痛的治疗靶点，糖尿病糖基代谢产物甲基乙二醛可能导致了KCNQ钾通道功能活性的下降。本研究为KCNQ钾通道为靶点的新药发现奠定了科学的理论基础。