M1毒蕈碱型胆碱信号途径对AMPA受体GluA2运输的调控及其在突触可塑性中的作用

M1 Muscarinic acetylcholine receptor plays an important role in learning and memory. Activation of M1 receptor modulates synaptic plasticity of hippocampal neurons directly or by enhancing the function of NMDA receptors. AMPA receptor trafficking plays a key role in the regulation of the synaptic strength. However, the regulation of AMPA receptor by muscarinic receptors has not been fully clarified. In our preliminary study, we demonstrated that activation of M1 receptor could modulate the trafficking of GluA1 subunits of AMPA receptors in cells of cultured primary hippocampal neurons, which contributes to the secretion of GluA1 to the postsynaptic density.Meanwhile we found that the number and phosphorylation level of GluA2 on the surface of neurons were modulated by M1 receptors as well. The research results of GluA2 help to clarify the modulation mechanism and significance of AMPA receptors by M1 receptors, therefore we will further clarify the details of the regulation of GluA2 trafficking by activation of M1 receptor and elucidate the underlying mechanism through live cell imaging andsubcellular fractionation of postsynaptic densities. Moreover, we will investigate the role of such regulation in synaptic plasticity in hippocampal slices from the mutant mice lack of the phosphorylation sites of GluA2 and in learning and memory in memory-impaired and Alzheimer’s disease mouse models and the phosphomutant mice of GluA2. The study will provide new insight in the role of M1 receptor in enhancement of cognition and it may give a chance to the development of novel methods to combat the impairment of cognitive function such as in Alzheimer’s disease.

M1毒蕈碱型胆碱受体(M1受体)在学习记忆等活动中作用显著。研究表明M1受体可直接或通过增强NMDA受体的功能影响海马神经元的突触可塑性，但对AMPA受体的影响尚未完全阐明，而AMPA受体在神经元内的运输对突触功能起关键作用。前期研究我们证明M1受体激动有助于海马神经元中的GluA1向突触后致密区分泌；同时发现神经元表面GluA2的数量与磷酸化水平亦受其影响。因此我们拟通过活细胞动态观察、突触后致密物蛋白提取等方法深入研究M1受体对AMPA受体GluA2运输的调控作用及其机制；应用点突变小鼠海马脑片研究GluA2的运输在M1受体调控突触可塑性中的作用；应用整体动物模型解析该作用在M1受体调控整体学习记忆中的影响。本课题有助于进一步揭示M1受体激动促进学习记忆的分子机制，有望在阿尔茨海默病等认知功能障碍疾病防治的基础研究中获得新突破。

认知灵活性在功能执行过程中发挥重要作用，胆碱能系统是认知的重要组成部分，主要通过纹状体和前额叶皮质调节认知灵活性。M1毒蕈碱型乙酰胆碱受体(M1mAChRs)是胆碱能系统中一个重要的治疗靶点，其在海马依赖的认知灵活性中的作用尚不清楚。本研究发现，M1mAChRs通过PKC磷酸化Ser880增加GluA2内吞作用。M1mAChR激活引起的海马CA1区长时程抑制(LTD)依赖于AMPA受体GluA2亚基，而不依赖于GluA1。选择性激活M1mAChRs促进了最初习得性反应的消退，并促进了在Morris水迷宫中获得逆转学习，这是一种主要依赖于海马的行为测试。然而，在蛋白激酶C(PKC)磷酸化Ser880缺失的GluA2突变小鼠中，上述作用被取消。抑制PKC可阻断M1mAChR介导的LTD、记忆转换和易化学习。此外，在GluA2突变小鼠和PKC抑制剂处理的小鼠中观察到的缓慢记忆消退似乎影响了逆转学习的巩固和恢复。这些结果表明M1mAChRs主要促进空间反转学习的习得，并进一步阐明这种作用依赖于PKC对GluA2的磷酸化。这项研究有助于阐明M1mAChRs在认知灵活性中的作用，并可能促使认知灵活性的早期预防。