Molecular Analysis of Functional Neural Circuits

功能神经回路的分子分析

• 批准号: 7847362
• 负责人: Leon Reijmers
• 金额: $ 247.5万
• 依托单位: TUFTS UNIVERSITY BOSTON
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-09-30 至 2014-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7847362
• 关键词: Address; Affinity Chromatography; Brain; Brain Diseases; Circadian Rhythms; Development; Epilepsy; Event; Fright; Lead; Maintenance; Memory; Memory impairment; Messenger RNA; Methods; Modification; Molecular; Molecular Analysis; Mus; Neurons; Pain; Protein Biosynthesis; Proteins; Research; Ribosomes; Transgenic Mice; Translating; Work; abstracting; addiction; conditioned fear; experience; interest; neural circuit; neuron loss; public health relevance; relating to nervous system; spinal cord regeneration; therapy development; tool

DESCRIPTION (Provided by the applicant) Abstract: A functional neural circuit consists of a group of connected neurons that collaborate to execute a specific function of the brain. The understanding of the molecular mechanisms that underlie the development, maintenance and experience-dependent modification of functional neural circuits is incomplete due to limitations of existing methods. I propose to generate a transgenic mouse that addresses these limitations by exploiting two recent methodological advances. The first advance is the TetTag mouse, which is a transgenic mouse that can be used to genetically tag a single functional neural circuit. This tag enables the selective molecular analysis of neurons that have a shared function. This method is more sensitive to changes within a single functional neural circuit than other available methods, which have to rely on spatial criteria and thereby include neurons that do not participate in the circuit of interest. The second advance is the Translating Ribosome Affinity Purification (TRAP) method, which enables purification of actively translated messenger RNA from a genetically defined group of neurons. TRAP analysis reflects changes at both the transcriptional and translational level, while other available methods only detect transcriptional changes. I will combine TetTag and TRAP within a single transgenic mouse to generate the first tool that enables the comprehensive analysis of all translational events within a single functional neural circuit. Neurons tagged with the TetTag mouse during fear conditioning provide a stable neural correlate of the fear memory. I will use the TetTag/TRAP mouse to purify actively translated messenger RNA from these tagged neurons in order to detect the protein synthesis events that underlie the storage of a memory. The TetTag/TRAP mouse can be used for the molecular analysis of various functional neural circuits, including those involved in memory, addiction, epilepsy, circadian rhythms, spinal cord regeneration, pain, brain development, and neuronal cell death. Public Health Relevance: The proposed research will lead to a better understanding of how the synthesis of new proteins helps neurons in the brain to work together and execute the functions of the brain. This increased understanding can be used to develop therapies for various brain diseases, including memory impairment, addiction and epilepsy.

描述(由申请人提供) 摘要：功能神经回路由一组相互连接的神经元组成，这些神经元相互协作执行大脑的特定功能。由于现有方法的局限性，对功能神经电路的发展、维护和经验依赖的修饰的分子机制的理解是不完整的。我建议通过利用最近的两个方法学进展来产生一种解决这些限制的转基因小鼠。第一个进展是TetTag小鼠，这是一种转基因小鼠，可以用来对单个功能神经回路进行基因标记。这个标签可以选择性地对具有共同功能的神经元进行分子分析。与其他现有方法相比，该方法对单个功能神经回路内的变化更敏感，其他方法必须依赖于空间标准，从而包括不参与感兴趣回路的神经元。第二个进展是翻译核糖体亲和纯化(TRAP)方法，它能够从遗传定义的一组神经元中纯化主动翻译的信使RNA。TRAP分析反映了转录和翻译水平的变化，而其他可用的方法只检测转录变化。我将在一只转基因小鼠中结合TetTag和Trap，以产生第一个工具，能够对单个功能神经回路中的所有翻译事件进行全面分析。在恐惧条件作用过程中，用TetTag小鼠标记的神经元提供了与恐惧记忆稳定的神经关联。我将使用TetTag/Trap小鼠从这些标记的神经元中提纯主动翻译的信使RNA，以检测记忆储存背后的蛋白质合成事件。TetTag/Trap小鼠可用于各种功能神经回路的分子分析，包括那些涉及记忆、成瘾、癫痫、昼夜节律、脊髓再生、疼痛、大脑发育和神经细胞死亡的回路。 与公共健康的相关性：拟议的研究将使人们更好地理解新蛋白质的合成如何帮助大脑中的神经元协同工作并执行大脑的功能。这种增加的认识可以用来开发各种大脑疾病的治疗方法，包括记忆障碍、成瘾和癫痫。