基于多肽探针的Nav1.2调节NMDA受体功能增强诱发神经毒性的机制研究

The excitotoxicity induced damage caused by the excessive activation of NMDA receptor is the crucial mechanism involving in the pathogenesis of various nervous system diseases, including ischemic stroke and Parkinson's disease. However, the role of voltage-gated sodium channel subtypes in excitotoxicity induced by NMDA receptor is still unknown. Peptide toxins from animal venoms with high selectivity are invaluable probes for investigation of the structure and function of ion channels. Our previous research revealed that scorpion toxin BmK NT1 activated NMDA receptor by selectively activating voltage-gated sodium channel Nav1.2, and produced overloading of intracellular calcium in cerebellar granule cells, ultimately leading to excitotoxic neuronal death. Furthermore, preliminary studies on neurotoxic mechanism showed that activation of Nav1.2 augmented the expression levels and regulated the phosphorylation of NMDA receptors, which is the key regulation mechanism of the enhancement function of NMDA receptor in producing neurotoxicity. Therefore, firstly using BmK NT1 as a selective peptide probe by aberrant activation of Nav1.2, the aim of the project is to elucidate the activation of Nav1.2 by BmK NT1 in the regulation of NMDA receptor opening, the molecular mechanism of enhancement function of NMDA receptor, and the regulation of NMDA receptor mediated neurotoxicity, with combination of electrophysiology, immunohistochemistry and tyrosine-point mutation and siRNA interference. The significance of our research is to elucidate the novel molecular and cellular mechanism of BmK NT1 in the regulation of the enhancement function of NMDA receptor inducing neurotoxicity by activating Nav1.2. Our research will provide novel targets for intervention and strategies for the clinical treatment of excitotoxic-related nervous system diseases.

NMDA受体过度激活引起兴奋性神经毒性是脑卒中等神经疾病的关键病理机制，但钠通道亚型在NMDA受体介导兴奋性毒性中的作用和机制尚无报道。多肽毒素是离子通道功能研究的重要探针。我们前期研究发现，蝎毒多肽BmK NT1专一性激活Nav1.2，诱导NMDA受体异常开放，导致小脑颗粒细胞死亡的神经毒性；进一步研究表明，Nav1.2激活调控NMDA受体上膜表达增强与磷酸化修饰，是调控NMDA受体功能增强从而介导神经毒性的关键机制。本课题利用BmK NT1为探针，建立Nav1.2异常激活诱导神经毒性的细胞病理模型，联合电生理、免疫组化、酪氨酸点突变、siRNA干扰等方法，研究Nav1.2异常激活调控NMDA受体的电生理特性、调控NMDA受体功能增强的分子机制、调控NMDA受体介导的神经毒性三层内容，阐明Nav1.2调控NMDA受体介导神经毒性的机制，为临床兴奋性神经损伤疾病治疗提供新的干预靶点和思路。

NMDA受体过度激活引起兴奋性神经毒性是脑卒中等多种神经系统疾病的关键病理机制，但钠通道亚型在NMDA受体介导兴奋性毒性中的作用和机制尚不清楚。动物多肽毒素是离子通道结构和功能研究的重要工具探针。我们的前期研究发现：蝎毒多肽BmK NT1专一性激活Nav1.2，诱导NMDA受体异常开放，导致小脑颗粒细胞死亡的神经毒性；激活Nav1.2调控NMDA受体上膜表达增强与磷酸化修饰是介导神经毒性的关键机制。本项目在以下几个方面开展了深入探索：① BmK NT1专一性激活Nav1.2的分子机制。② Nav1.2激活调控NMDA受体功能增强的机制。③ 激活Nav1.2增强NNDA功能介导小脑颗粒细胞死亡的机制。本项目的开展揭示了激活Nav1.2调控NMDA受体功能增强介导神经毒性的机制，为理解兴奋性神经损伤疾病机理及制定防治策略提供新的视角和分子靶标。