Links among synaptic activity, translation and structure

描述（由申请人提供）： 突触功能特征的变化被认为是 大脑发育和学习的基本过程。总目标 的建议项目是确定谷氨酸之间的相互关系 受体激活、蛋白质合成和树突棘形状， 参与稳定这种突触变化。拟议的研究 有三个具体目标：1）确定空间对应度 突触之间的活动水平与功能 通过涉及帽依赖性的机制的蛋白质合成的变化和位点 和内部核糖体进入序列（IRES）翻译机制; 2） 定义帽依赖性和内部引发的相对贡献 树突中活性诱导蛋白质合成的翻译机制; 以及，3）确定翻译和脊柱之间的一般相互依赖关系 结构，并测试蛋白质合成在形成中的具体作用。 在脆性X综合征模型中观察到的异常长的脊柱， 代谢型谷氨酸受体的刺激。新型荧光试剂 已经开发了进行这些研究，包括标记，基于 荧光共振能量转移（FRET），用于标记突触经历 可塑性变化，以及几种帽依赖性和IRES的荧光报告基因 介导的树枝状翻译。研究将在分离的 海马的器官型切片培养，并将使用假共聚焦 一种由软件包控制的荧光显微镜系统，所述软件包包括 用于定量荧光报告基因表达的图像分析工具。的 建议的实验应该为我们理解 树突状蛋白质合成如何与突触活动、结构和 效能的形式改变了人们对大脑全球过程的看法 发展、学习和记忆。

DESCRIPTION (provided by applicant): Changes in the functional characteristics of glutamatergic synapses are thought to underlie basic processes of brain development and learning. The overall goal of the proposed project is to define interrelationships between glutamate receptor activation, protein synthesis, and dendritic spine shape that may be involved in the stabilization of such synaptic changes. The proposed studies have three specific aims: 1) to determine the degree of spatial correspondence between synapses undergoing activity at a level associated with functional change and sites of protein synthesis via mechanisms that involve cap-dependent and internal ribosomal entry sequence (IRES) mechanisms of translation; 2) to define the relative contributions of cap-dependent and internal initiation mechanisms of translation to activity-induced protein synthesis in dendrites; and, 3) to determine the general interdependence between translation and spine structure, and to test the specific role of protein synthesis in the formation of abnormally long spines seen in a fragile X syndrome model and with stimulation of metabotropic glutamate receptors. Novel fluorescent reagents have been developed to conduct these studies, including a marker, based on fluorescence resonance energy transfer (FRET), for marking synapses undergoing plastic change, and several fluorescent reporters of cap-dependent and IRES mediated dendritic translation. The studies will be conducted in dissociated and organotypic slice cultures of hippocampus, and will use a pseudoconfocal fluorescence microscope system controlled by a software package that includes image analysis tools for quantification of fluorescent reporter expression. The proposed experiments should provide valuable information for our understanding of how dendritic protein synthesis relates to synaptic activity, structure, and forms of efficacy change thought to underlie global processes of brain development, learning, and memory.