Defining the circuit, synaptic, and molecular mechanisms linking intracellular Ca2+ release to learning using subcellularly-targeted manipulations and imaging techniques in dendrites in vivo

使用体内树突的亚细胞靶向操作和成像技术定义连接细胞内 Ca2 释放与学习的电路、突触和分子机制

• 批准号: 10502363
• 负责人: Justin O'Hare
• 金额: $ 13.62万
• 依托单位: COLUMBIA UNIVERSITY HEALTH SCIENCES
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-07-15 至 2024-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10502363
• 关键词: Action Potentials; Address; Alzheimer' s Disease; Animals; Apical; Area; Automobile Driving; BRAIN initiative; Behavior; Behavioral; Biophysics; Brain; Calcium; Cells; Color; Communities; Consultations; Dendrites; Development Plans; Electroporation; Endoplasmic Reticulum; Environment; Equipment; Fire - disasters; Foundations; Glutamates; Goals; Head; Hippocampus (Brain); Image; Imaging Techniques; In Vitro; Institutes; Intervention; Investigation; Knowledge; Laboratories; Learning; Light; Link; Literature; Logic; Memory; Mentorship; Mission; Molecular; Molecular Genetics; Monitor; Morphology; Mus; Neurodegenerative Disorders; Neuronal Plasticity; Neurons; Neurosciences; Output; Parkinson Disease; Pathway interactions; Pattern; Pharmacology; Play; Prevalence; Reporter; Reporting; Research; Research Activity; Resource Development; Role; Shapes; Supervision; Synapses; Synaptic plasticity; Testing; Work; autism spectrum disorder; awake; base; career development; cellular imaging; experience; experimental study; gain of function; hippocampal pyramidal neuron; imaging approach; improved; in vivo; innovation; insight; nervous system disorder; neuronal cell body; novel; optogenetics; place fields; post-doctoral training; postsynaptic; presynaptic; receptive field; relating to nervous system; spatiotemporal; subcellular targeting; tool; two-photon; virtual environment; virtual reality; way finding

Project Summary/Abstract Candidate Goals and Mission Relevance: The applicant’s broad, long-term objective is to investigate how high- (circuit/behavioral) and low- (subcellular/molecular) level organizational principles of the brain cooperate to drive learning. The proposed research activities will build a foundation for this long-term goal and, in so doing, will promote BRAIN 2025 Report goals by integrating new technological and conceptual approaches to causally link intracellular Ca2+ release (ICR) from endoplasmic reticulum (ER) to neural activity dynamics and behavior. Project description: Dendritic Ca2+ is central to neural plasticity mechanisms allowing animals to adapt to the environment. ICR has long been thought to shape these mechanisms. The applicant recently carried out the first investigation of ICR in mammalian neurons in vivo to uncover how this subcellular phenomenon shapes experience-dependent feature selectivity across the dendritic arbor of pyramidal neurons (PNs) in mouse hippocampal area CA1. This work raises important questions regarding when, where, and how ICR is engaged to support learning. The applicant will address these questions in the following Aims: Aim 1. Characterize plasticity-associated ER Ca2+ dynamics in dendrites in vivo (K99): To achieve this Aim, the applicant will perform simultaneous dual-color, dual-plane in vivo 2-photon imaging of cytosolic and ER- resident Ca2+ in dendrites of single CA1 PNs during head-fixed spatial navigation of novel virtual environments. Aim 2. Define the synaptic logic tying intracellular Ca2+ release to in vivo synaptic plasticity (K99/R00): The applicant will first create a novel molecular tool to optogenetically induce ICR (Aim 2.1; K99). The applicant will then combine this precise interventional tool with single-cell imaging, inducible blockade of presynaptic release, and optogenetic dampening of ICR to dissect the synaptic logic by which ICR participates in plasticity induction in behaving mice. (Aim 2.2; R00). Aim 3. Dissect excitatory circuit-molecular mechanisms driving intracellular Ca2+ release in vivo (R00): The candidate will optogenetically activate specific excitatory projections onto distinct dendritic compartments of single CA1PNs while monitoring ER Ca2+ dynamics in behaving mice. Local pharmacological manipulations will dissect contributions of the two canonical pathways that convert presynaptic excitatory input to postsynaptic ICR. Career development plan: The applicant will extend a highly complementary Co-Mentorship arrangement between Drs. Franck Polleux and Attila Losonczy who possess deep expertise in cellular/molecular/genetic and in vivo/behavioral approaches, respectively. The applicant will receive robust consultative support from Dr. Stefano Fusi of Columbia’s Center for Theoretical Neuroscience and Dr. Darcy Peterka, Director of Cellular Imaging at Columbia’s Zuckerman Institute. The applicant’s research and transition to independence will benefit from this strong mentorship team, state-of-the-art facilities, all necessary equipment, and numerous Professional Development resources offered through the Columbia Office of Postdoctoral Affairs, the Zuckerman Institute, and the BRAIN Initiative.

项目概要/摘要 候选人目标和使命相关性：申请人的广泛、长期目标是调查 大脑的（电路/行为）和低（亚细胞/分子）水平组织原则合作驱动 拟议的研究活动将为这一长期目标奠定基础，并以此为基础。 通过整合新技术和概念方法来建立因果关系，促进 BRAIN 2025 报告目标 细胞内 Ca2+ 释放（ICR）从内质网（ER）到神经活动动力学和行为。 项目描述：树突状 Ca2+ 是神经可塑性机制的核心，使动物能够适应 长期以来，ICR 一直被认为可以塑造这些机制。 对哺乳动物神经元体内 ICR 的研究，揭示这种亚细胞现象是如何形成的 小鼠锥体神经元（PN）树突轴上的经验依赖性特征选择性 这项工作提出了有关 ICR 何时、何地以及如何参与的重要问题。 为了支持学习，申请人将在以下目标中解决这些问题： 目标 1. 表征体内树突中与可塑性相关的 ER Ca2+ 动力学 (K99)：为了实现这一目标 目标是，申请人将对细胞质和 ER- 进行同步双色、双平面体内 2 光子成像。 在新型虚拟环境的头部固定空间导航过程中，单个 CA1 PN 的树突中存在驻留的 Ca2+。 目标 2. 定义将细胞内 Ca2+ 释放与体内突触可塑性联系起来的突触逻辑 (K99/R00)： 申请人将首先创建一种新颖的分子工具来光遗传学诱导ICR（目标2.1；K99）。 然后将这种精确的介入工具与单细胞成像、突触前诱导阻断相结合 ICR 的释放和光遗传学抑制，以剖析 ICR 参与可塑性的突触逻辑 对行为小鼠进行诱导（目标 2.2；R00）。 目标 3. 剖析体内驱动细胞内 Ca2+ 释放的兴奋性回路分子机制 (R00)： 候选人将通过光遗传学激活特定的兴奋性投射到不同的树突区室上 单个 CA1PNs，同时监测行为小鼠的 ER Ca2+ 动态 局部药理操作将。 剖析将突触前兴奋性输入转化为突触后 ICR 的两种典型途径的贡献。 职业发展计划：申请人将提供高度互补的共同导师安排 Franck Polleux 博士和 Attila Losonczy 博士在细胞/分子/遗传和 申请人将分别获得体内/行为方法的强有力的咨询支持。 哥伦比亚理论神经科学中心的 Stefano Fusi 和细胞主任 Darcy Peterka 博士 哥伦比亚大学祖克曼研究所的成像将使申请人的研究和向独立的过渡受益。 来自这个强大的指导团队、最先进的设施、所有必要的设备以及众多的专业人士 通过哥伦比亚博士后事务办公室、祖克曼研究所提供的发展资源， 和大脑计划。