The role of nitric oxide signaling in synaptic plasticity

一氧化氮信号传导在突触可塑性中的作用

• 批准号: RGPIN-2014-06085
• 负责人: Ryan, Scott
• 金额: $ 2.91万
• 依托单位: University of Guelph
• 依托单位国家: 加拿大
• 项目类别: Discovery Grants Program - Individual
• 财政年份: 2016
• 资助国家: 加拿大
• 起止时间: 2016-01-01 至 2017-12-31
• 项目状态: 已结题
• 来源: https://www.nserc-crsng.gc.ca/ase-oro/Details-Detailles_eng.asp?id=612023
• 关键词: role; nitric; oxide; signaling; synaptic

Learning and memory has at its core modifications to tiny spine like structures that exist on a special subset of neurons in the brain known as “spiny neurons.” These spines must move, adapt, grow and retract in response to chemical neurotransmitters in the brain, a process that as a whole is believed to be the cellular manifestation of learning, known as synaptic plasticity. The spines themselves are thought to serve as basic units of memory storage. While much work has focused directly on the chemical signals that mediate communication in the brain and the ion channels that are opened as a result, little is know of the specific signaling pathways that control the actual generation, shape and loss of individual spines. This proposal aims to shed light on this process by focusing on a specific molecule (Nitric Oxide) that is produced in these spines following ion channel opening. Nitric Oxide is unique in that it can directly alter the proteins responsible for dynamic changes to spine morphology. This proposal is driven by the hypothesis that Nitric Oxide signaling regulates synaptic plasticity by altering the density and morphology of spines through a process called S-nitrosylation. Ion channels link a cell with the environment that surrounds it. These channels allow the cell to interpret its surroundings by responding to neurotransmitters and propagating signals from the environment outside of the cell inward. These signals mediate changes in the structure and shape of the cell itself and facilitate spine growth and retraction. One class of ion channel of interest to this proposal is characterized by its ability to respond to a neurotransmitter called glutamate. Glutamate signaling can increase the efficiency of communication between cells in the brain by both increasing the number and stability of new spines as well by pruning excess or unnecessary spines, leaving a more efficient communication network. Glutamate-mediated opening of the “NMDA” type ion channel results in production of Nitric Oxide that modifies the proteins in spines through a chemical reaction called S-Nitrosylation. This event alters the function of near-by proteins and as a result the dynamic properties of the spines they inhabit. While it is estimated that 50% of proteins in the brain are altered by S-nitrosylation, a comprehensive analysis of the role it plays in synaptic plasticity has never been performed. Using powerful mouse genetics we will control the level of Nitric Oxide synthesized in spines by altering the genetic composition of the NMDA-type ion channel and thus its ability to open and close. Using live imaging microscopy techniques coupled with fluorescent probes, we will monitor the flux of ions through NMDA-type channels as well the amount Nitric Oxide subsequently produced in cultured brain slices and isolated neurons from these mice. This will allow us to determine how the level of Nitric Oxide correlates with dynamic changes to spine morphology. We will then monitor what proteins are S-nitrosylated by Nitric Oxide under conditions of spine growth, spine stabilization and spine retraction using Mass Spectrometry to analyze changes to proteins composition. This will form the basis of a novel molecular pathway that underlies the processes of synaptic plasticity. Moreover, this research program will further our understanding neural architecture and the means by which cells of the brain communicate.

学习和记忆的核心是对微小的脊椎状结构的修改，这些结构存在于大脑中一种特殊的神经元子集上，称为“棘神经元”。这些脊椎必须移动、适应、生长和收缩，以响应大脑中的化学神经递质，这个过程被认为是学习的细胞表现，被称为突触可塑性。脊椎本身被认为是记忆存储的基本单位。虽然很多研究都直接集中在化学信号和离子通道上，这些化学信号在大脑中起中介作用，从而打开了离子通道，但对控制单个脊椎的实际产生、形状和损失的具体信号通路却知之甚少。这项提议旨在通过关注离子通道开放后在这些脊椎中产生的一种特定分子(一氧化氮)来阐明这一过程。一氧化氮的独特之处在于，它可以直接改变导致脊柱形态动态变化的蛋白质。这一假设是基于这样一个假设，即一氧化氮信号通过一种名为S-亚硝化的过程改变脊椎的密度和形态，从而调节突触的可塑性。 离子通道将细胞与其周围的环境联系起来。这些通道允许细胞通过对神经递质的反应来解释其周围环境，并将来自细胞外环境的信号向内传播。这些信号调节细胞本身的结构和形状的变化，并促进脊柱的生长和回缩。对这一提议感兴趣的一类离子通道的特征是它对一种名为谷氨酸的神经递质做出反应的能力。谷氨酸信号可以通过增加新脊椎的数量和稳定性，以及通过修剪多余或不必要的脊椎，从而提高大脑细胞之间的沟通效率，留下更有效的沟通网络。谷氨酸介导的“N-甲基-D-天冬氨酸”型离子通道的开放导致一氧化氮的产生，一氧化氮通过称为S-亚硝化的化学反应改变脊柱中的蛋白质。这一事件改变了邻近蛋白质的功能，从而改变了它们所栖息的脊椎的动态特性。虽然据估计，大脑中50%的蛋白质会被S-亚硝化改变，但对其在突触可塑性中所起作用的全面分析从未进行过。 利用强大的小鼠遗传学，我们将通过改变NMDA型离子通道的遗传组成，从而改变其打开和关闭的能力，来控制脊柱中合成的一氧化氮的水平。使用结合荧光探针的实时成像显微镜技术，我们将监测通过NMDA型通道的离子流量，以及随后在培养的脑片和这些小鼠的分离神经元中产生的一氧化氮的量。这将使我们能够确定一氧化氮水平如何与脊柱形态的动态变化相关。然后，我们将监测在脊柱生长、脊柱稳定和脊柱回缩条件下，哪些蛋白质被一氧化氮亚硝化，并用质谱仪分析蛋白质组成的变化。这将形成一种新的分子途径的基础，该途径是突触可塑性的基础。此外，这项研究计划将进一步加深我们对神经结构和脑细胞交流方式的理解。