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

项目摘要/摘要 候选人目标和使命相关性：申请者的广泛、长期目标是调查- 大脑的(电路/行为)和低水平(亚细胞/分子)组织原则协同驱动 学习。拟议的研究活动将为这一长期目标奠定基础，这样做将 通过整合新的技术和概念方法进行因果联系，促进Brain 2025报告目标 内质网的细胞内钙释放(ICR)对神经活动动力学和行为的影响。 项目描述：树突状钙是神经可塑性机制的核心，使动物能够适应 环境。长期以来，人们一直认为ICR塑造了这些机制。申请人最近进行了第一次 研究活体哺乳动物神经元的ICR以揭示这种亚细胞现象是如何形成的 小鼠锥体神经元树突的经验依赖性特征选择 海马区CA1。这项工作提出了关于何时、何地以及如何参与ICR的重要问题 来支持学习。申请者将在以下目标中回答这些问题： 目的1.研究体内树突中可塑性相关的内质网钙动力学(K99)：实现这一目标 目的：申请者将同时进行双色、双平面活体双光子胞浆和ER- 头固定空间导航新虚拟环境中单个CA1-PNS树突内的钙离子。 目的2.明确细胞内钙释放与体内突触可塑性的突触逻辑(K99/R00)： 申请人将首先创造一种新的分子工具来光遗传诱导ICR(目标2.1；K99)。申请人 然后将这种精确的介入工具与单细胞成像相结合，可诱导地阻断突触前 ICR的释放和光发生抑制以剖析ICR参与可塑性的突触逻辑 对行为正常的小鼠的诱导。(目标2.2；R00)。 目的3.解剖兴奋回路--活体(R00)细胞内钙释放的分子机制： 候选人将通过光基因激活特定的兴奋性投射到不同的树突间隔上 单个CA1PN同时监测行为小鼠内质网钙动态。当地的药理操作将 剖析将突触前兴奋性输入转化为突触后ICR的两条典型通路的作用。 职业发展计划：申请者将延长互补性很强的共同导师安排 Franck Polleux博士和Attila Losonczy博士之间，他们在细胞/分子/遗传学和 分别采用活体/行为方法。申请者将得到博士强有力的咨询支持。 哥伦比亚大学理论神经科学中心的Stefano Fusi和细胞主任Darcy Peterka博士 哥伦比亚大学扎克曼研究所的成像技术。申请人的研究和向独立的过渡将受益 来自这个强大的指导团队、最先进的设施、所有必要的设备和众多专业人员 发展资源由哥伦比亚博士后事务办公室、扎克曼研究所、 和大脑计划。

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.