Probing the Structure of the Synapse Using Superresolution Light Microscopy

摘要 记忆和其他认知功能部分存在于突触连接的模式和强度中 在神经元之间。对突触强度的分子决定因素的了解由来已久 神经科学的目标，以及该领域的进展将影响我们对 几乎所有的神经疾病，从自闭症到阿尔茨海默病？S病。在过去十年中 生物化学和传统的分子和遗传学方法已经开始拼凑成 神经递质受体与其他突触蛋白的相互作用对突触的调控 强度和可塑性，但一个主要限制是很少或根本没有关于如何 蛋白质在突触处排列成信号复合体。许多信号分子只能 与紧邻的蛋白质相互作用，这种定位本身可能受到经验的调节。 了解功能信号复合体是如何产生的，以及它们如何反过来调节突触 因此，我们需要探索突触后蛋白质的空间排列。 密度(PSD)。传统方法没有足够的分辨率来允许 在这些微小的突触结构中定位突触蛋白。在此，我建议 开发工具来绘制单个突触蛋白(如谷氨酸)的空间排列 )，并确定突触如何影响这些空间排列 可塑性，使用超分辨率光学显微镜。通过映射许多不同的 突触后膜内的蛋白质和PSD我们将能够生成一个三维模型 构成突触后侧的蛋白质晶格。此方法有望实现 将大量关于蛋白质-蛋白质相互作用的生化和分子数据放入一个结构 语境对其解释是必不可少的，并将为突触的分析增加一个强大的新工具 功能。

ABSTRACT Memory and other cognitive functions reside in part in the pattern and strength of synaptic connections between neurons. Understanding the molecular determinants of synaptic strength has been a longstanding goal of neuroscience, and advances in this field stand to influence our understanding of virtually every neurological disorder from Autism to Alzheimer¿s disease. Over the past decade biochemical and conventional molecular and genetic approaches have begun to piece together how interactions between neurotransmitter receptors and other synaptic proteins regulate and control synaptic strength and plasticity, but a major limitation is that there is little or no structural information about how proteins are arranged into signaling complexes at the synapse. Many signaling molecules can only interact with immediately adjacent proteins, and this localization may itself be regulated by experience. Understanding how functional signaling complexes are generated and how they in turn regulate synaptic strength thus requires that we probe the spatial arrangements of proteins within the postsynaptic density (PSD). Conventional approaches do not have sufficient resolution to allow the position of synaptic proteins to be mapped within these tiny (< 1 &#956;m) synaptic structures. Here I propose to develop tools to map the spatial arrangements of individual synaptic proteins (such as glutamate receptors) within the PSD, and to determine how these spatial arrangements are influenced by synaptic plasticity, using super resolution light microscopy. By mapping the relative positions of many different proteins within the postsynaptic membrane and PSD we will be able to generate a 3 dimensional model of the protein lattices that comprise the postsynaptic side of the synapse. This method has the promise to put a vast array of biochemical and molecular data on protein-protein interactions into a structural context that is essential for its interpretation, and will add a powerful new tool to the analysis of synaptic function.