а-Klotho在记忆形成和海马突触可塑性中的作用机制

The project addresses the question whether cellular mechanisms of memory formation can be long-lasting modified by a cell and brain region specific overexpression of the putative anti-aging protein Klotho in mice. .To this end we conducted preliminary experiments showing that the endogenous alpha-Klotho expression is brain region and age dependently regulated. In addition, we observed that overexpression of the secreted form of alpha Klotho in the hippocampal CA1 region alters basal synaptic transmission, induction of activity-dependent synaptic plasticity and neurogenesis in the subgranular zone of the dentate gyrus. Moreover, the nesting behavior of Klotho overexpression mice was altered as well as object recognition and objection location memory. To follow up these preliminary results, we propose a research methodology that includes in vivo behavioral experiments using memory tasks that depend on distinct brain regions, such as the hippocampus, the prefrontal and perirhinal cortices after transduction of neuronal cells with the secreted form of human alpha-Klotho. Further, we propose to study the cellular mechanisms of Klotho’s potential effects on memory formation by in vitro experiments utilizing activity-dependent synaptic plasticity combined with pharmacology. We propose to complement the study by protein biochemical analysis of protein markers for synaptic plasticity and neurogenesis..The proposed study will help to understand the involvement of the Klotho gene family in mechanisms of memory formation and related neurogenesis and has the potential to gain knowledge helping to develop gene therapies that could prevent memory decline.

抗衰老基因Klotho对记忆形成和脑疾病的影响受到关注，但在特定脑区中过表达该基因是否可以长时程调节记忆的形成还有待探究。我们前期工作发现,在小鼠大脑中内源性分泌型Klotho的表达具有区域性以及年龄依赖性。同时在老年小鼠海马区过表达人源分泌型Klotho可以改变基础突触传递，增强突触可塑性,并影响小鼠筑巢行为,物体识别以及位置识别行为。此外在过表达该蛋白的齿状回颗粒下层区域还发现新生神经元数量增加。为进一步追踪这些结果,本项目将联合使用电生理及药理学和行为学实验等技术手段进一步探究它参与记忆形成的分子机制,并通过对神经元再生的特异性相关蛋白质进行生物化学分析补充研究,揭示其是否参与齿状回颗粒下层区域的神经元再生。我们的研究有望阐明Klotho基因家族在记忆形成中的作用机制,并进一步揭示其是否通过影响神经元再生参与学习记忆过程，从而为防止记忆衰退提供一种可行的新型基因治疗策略。

Klotho基因家族由α-、β-、和γ-Klotho组成，可以编码具有胞外结构域的I型单跨膜蛋白。其中α-Klotho作为该家族的主要成员，其表达产物有膜结合型、可溶型和分泌型三种蛋白异构体形式，主要在肾脏和脑中表达，这提示其表达产物在肾脏和脑中可能具有的重要功能。α-Klotho基因的缺失导致小鼠表现出类似人类衰老的症状，而在小鼠体内过表达Klotho蛋白则会延长小鼠寿命，同时，在前期实验中，我们发现，在C57BL/6野生型小鼠海马区，内源性分泌型α-Klotho蛋白的表达量随年龄增加而减少。过表达人源分泌型α-Klotho蛋白可以影响小鼠的筑巢行为和增强小鼠对新物体识别和物体新位置识别的能力、可以提升小鼠在被动回避任务中的表现、还可以调控α-Klotho蛋白参与的信号通路蛋白和突触传递相关蛋白的表达和改变突触的基础传递和突触可塑性。总之，我们的研究表明人源分泌型α-Klotho蛋白的过表达可能有助于记忆的形成。