缝隙连接蛋白36在脑缺血诱导的纹状体神经元损伤中的作用及其活化的分子机制研究

项目名称： 缝隙连接蛋白36在脑缺血诱导的纹状体神经元损伤中的作用及其活化的分子机制研究

项目编号： No.31502025

项目类型： 青年科学基金项目

立项/批准年度： 2016

项目学科： 农业科学

项目作者： 马云飞

作者单位： 中国农业大学

项目金额： 20万元

中文摘要： 在成年哺乳动物的中枢神经系统中，缝隙连接蛋白Cx36维持在低水平并局限于特异类型神经元中表达，然而在缺血性神经元损伤时，Cx36的表达水平迅速升高。但迄今为止Cx36的活化在缺血诱导的神经元损伤中的具体作用和机制尚不清楚。纹状体是缺血性脑损伤的主要部位，本项目拟用脑缺血大鼠模型，以纹状体为靶点，结合应用Cx36阻断剂，检测缺血诱导的神经元死亡数量、Cx36表达和磷酸化水平，研究Cx36在缺血性神经元损伤中的具体功能。最近体外研究发现Cx36能够被CaMKII绑定并被其磷酸化，而CaMKII在活动依赖的电突触可塑性中起着关键作用。因此我们对Cx36作用机制的研究拟从CaMKII对Cx36的调控入手，通过体内缺血模型和体外CaMKII的激动实验及缺糖乏氧实验，采用分子生物学等手段，阐明缺血诱导的Cx36上调表达和活化的分子机制。本项目的实施将为缺血性神经元损伤的治疗提供新的靶点及理论依据。

中文关键词： 缝隙连接蛋白36；钙/钙调素依赖性蛋白激酶II；磷酸化；缺血性脑损伤；大鼠纹状体

英文摘要： In the mammalian central nervous system (CNS), the level of gap junction protein connexin 36 (Cx36) remains low in the adult, confined to specific subsets of neurons. However, following neuronal injury such as ischemia, the coupling and expression of Cx36 rise. Further, until recently, the role of Cx36 in the neuronal injury induced by cerebral ischemia and its specific molecular mechanisms were not understood. The striatum is the major area of ischemic brain injury. Therefore, in the present study we investigate thoroughly the numbers of ischemic neuronal death, the level of Cx36 expression and phosphorylation by using photothrombotic focal cerebral ischemia in rat striatum combined with pharmacological blockade of Cx36 neuronal gap junctions, to determine the function of Cx36 activation during ischemic neuronal injury. More recently, it has been reported that Cx36 binds to and is phosphorylated by Ca/Calmodulin kinase II alpha (CaMKII), and CaMKII has been implicated in electrical synaptic plasticity and learning and memory. Therefore, we tested the hypothesis that CaMKII control the injury-mediated increase and activation in Cx36. Hyposic experiment and CaMKII-activating Cx36 experiment in cultured neuronal cell lines in vitro, and several molecular biology techniques were taken, to clarify the molecular mechanism of upregulation and activation Cx36 expression in neuronal injury by ischemia. The present study would contribute to revealing organization principles of Cx36-based gap junctions in the striatum circuitry and potentially open up novel approaches for neuroprotection in the acute period following ischemic brain injury.

英文关键词： connexin36;CaMKII;phosphorylation;ischemic brain injury;rat striatum