Scalable Cell- and Circuit-Targeted Electrophysiology

可扩展的细胞和电路靶向电生理学

• 批准号: 9893932
• 负责人: Edward S. Boyden
• 金额: $ 56.63万
• 依托单位: MASSACHUSETTS INSTITUTE OF TECHNOLOGY
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-07-01 至 2022-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9893932
• 关键词: 3-Dimensional; Algorithms; Anatomy; Antibodies; Bar Codes; Behavioral; Brain; Brain Diseases; Brain Injuries; Calcium; Cells; Chemicals; Clinical; Code; Complex; DNA; Data Set; Development; Disease; Electrodes; Electronics; Electrophysiology (science); Exhibits; Fluorescence Microscopy; Grant; Hippocampus (Brain); Image; In Situ; Individual; Instruction; Interneurons; Isotropy; Label; Learning; Manuscripts; Methods; Microscopy; Morphology; Mus; Neurons; Organism; Parvalbumins; Performance; Physiology; Polymers; Preparation; Probability; Procedures; Property; Proteins; Protocols documentation; Recording of previous events; Reporter; Research Personnel; Robot; Robotics; Scientist; Slice; Streptavidin; Synapses; System; Techniques; Technology; Time; Tissue Expansion; Tissues; Visualization; base; biocytin; cell type; cranium; design; fluorescence microscope; fluorophore; high throughput analysis; hippocampal pyramidal neuron; image guided; improved; in vivo; miniaturize; nanoscale; neural circuit; neuronal circuitry; neurosurgery; novel; patch clamp; preservation; relating to nervous system; sensory stimulus; synaptic function; tool; two photon microscopy; two-photon

The functional activity and dysregulation of neuronal circuits relies critically on the physiology of neuronal synapses, which are challenging to analyze because they appear in great numbers, and they are difficult to record in vivo, especially in relation to the dynamic neural codes generated by specific neurons. To make things even more complex: synapses are incredibly dynamic in fashions that are dependent on recent history, sensory stimuli, disease state, and other behaviorally relevant contexts. Ideally there would be a technology that would allow for individual investigators to rapidly analyze synapses between neurons exhibiting neural codes in a behavioral context, so that it is possible to understand how information is trans formed at synapses. We here propose to develop a simple, easily deployable toolbox for achieving this, building from several recent discoveries. First, we have found (manuscript in preparation) that it is possible to automatically perform whole cell patch clamp neural recording of cells in the living mouse brain that have been identified via two-photon fluorescence microscopy (e.g., cells of a given type that express a genetically encoded fluorophore). We here propose to invent a multiple-neuron patching version of this “imagepatching” robot, to enable the simultaneous characterization of the neural codes in multiple neurons, as well as the synaptic connections between them (Aim 1). We will also develop miniaturized and optimized hardware capable of performing imagepatching, neurosurgery, and patch clamp electrode reuse for improved yield and throughput of synaptic assessment. (Aim 2). Also, we have discovered that it is possible to physically expand preserved neural circuits, by embedding them in swellable polymers, and then chemically expanding those polymers, a technology we call expansion microscopy (ExM), which enables nanoscale imaging of 3-D tissues and organisms. We propose to optimize ExM for the analyses of synapses (Aim 3). We here propose a fast-paced, 4-year grant, to create a powerful, easy-to-use toolbox that makes the critical task of in vivo synaptic physiology into a routine, automated procedure. We will distribute all tools and datasets as freely as possible, sharing all algorithms, circuit designs, and assembly instructions, and hosting visitors to learn these technologies – for which we have an extensive track record.

神经元电路的功能活性和失调依赖于神经元的生理 突触，这是分析的挑战，因为它们出现了很多，而且很难 记录体内的记录，特别是与特定神经元产生的动态神经元有关。做 事情更加复杂：突触在依赖最近历史的时尚中具有令人难以置信的动态性， 感觉刺激，疾病状态和其他行为相关的环境。理想情况下会有一项技术 这将使单个研究者能够快速分析表现神经元的神经元之间的突触 在行为上下文中代码，以便可以了解如何在突触中翻译信息。 我们在这里建议开发一个简单，易于部署的工具箱来实现这一目标，从最近的几个 发现。首先，我们发现（准备中的手稿）可以自动执行整个 通过两光子鉴定的活小鼠大脑细胞的细胞夹夹神经元记录 荧光显微镜（例如，给定类型的细胞表达泛滥的荧光团）。我们在这里 提出发明此“ image绘制”机器人的多神经修补版本，以启用简单 多个神经元中神经元的表征以及它们之间的突触连接 （目标1）。我们还将开发小型和优化的硬件，能够执行图像绘制， 神经外科手术和斑块夹电极再利用，以提高突触评估的产量和吞吐量。 （目标2）。另外，我们发现，可以通过 将它们嵌入膨胀的聚合物中，然后化学扩展这些聚合物，我们称之为这项技术 扩展显微镜（EXM），可实现3-D组织和生物的纳米级成像。我们建议 优化EXM用于突触分析（AIM 3）。我们在这里提出一项快节奏的4年赠款，以创建一个 功能强大，易于使用的工具箱，将体内突触生理学的关键任务变成例程， 自动化过程。我们将尽可能免费分发所有工具和数据集，共享所有算法， 电路设计和集会说明，以及托管访问者学习这些技术 - 我们为此 广泛的记录。

项目成果

Expansion Microscopy: Protocols for Imaging Proteins and RNA in Cells and Tissues.

• DOI: 10.1002/cpcb.56
• 发表时间: 2018-09-01
• 期刊: Current protocols in cell biology
• 作者: Asano, Shoh M;Gao, Ruixuan;Boyden, Edward S
• 通讯作者: Boyden, Edward S

Light microscopy based approach for mapping connectivity with molecular specificity

• DOI: 10.1038/s41467-020-18422-8
• 发表时间: 2020-02
• 期刊: Nature Communications
• 影响因子: 16.6
• 作者: Fred Y. Shen;Margaret M. Harrington;Logan A. Walker;Hon Pong Jimmy Cheng;E. Boyden;Dawen Cai

Expansion microscopy: principles and uses in biological research

• DOI: 10.1038/s41592-018-0219-4
• 发表时间: 2019-01-01
• 期刊: NATURE METHODS
• 影响因子: 48
• 作者: Wassie, Asmamaw T.;Zhao, Yongxin;Boyden, Edward S.
• 通讯作者: Boyden, Edward S.

Expansion microscopy: enabling single cell analysis in intact biological systems.

• DOI: 10.1111/febs.14597
• 发表时间: 2019-04
• 期刊: The FEBS journal
• 作者: Alon S;Huynh GH;Boyden ES
• 通讯作者: Boyden ES