Population Imaging of Action Potentials by Novel Two-Photon Microscopes and Genetically Encoded Voltage Indicators

通过新型双光子显微镜和基因编码电压指示器对动作电位进行群体成像

• 批准号: 9588470
• 负责人: Jerry L Chen
• 金额: $ 268.09万
• 依托单位: BOSTON UNIVERSITY (CHARLES RIVER CAMPUS)
• 依托单位国家: 美国
• 财政年份: 2018
• 资助国家: 美国
• 起止时间: 2018-09-30 至 2022-10-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9588470
• 关键词: Action Potentials; Address; Anatomy; Animals; Behavior; Biological; Brain; Brain Diseases; Calcium; Cells; Code; Communication; Communities; Development; Electrophysiology (science); Elements; Engineering; Fiber; Frequencies; Future; Genetic Engineering; Goals; Image; Individual; Interneurons; Investigation; Kinetics; Label; Lasers; Measurement; Measures; Membrane; Methods; Microscope; Microscopy; Molecular; Monitor; Mus; Neocortex; Neurons; Neurosciences; Noise; Opsin; Optical Methods; Optics; Phase; Photons; Physiologic pulse; Population; Process; Property; Protein Engineering; Reporting; Research Personnel; Resolution; Scanning; Signal Transduction; Source; Speed; System; Techniques; Technology; Testing; Thulium; Time; Training; Vibrissae; Work; awake; base; design; detector; experimental study; genetic information; imaging system; in vivo; in vivo evaluation; instrumentation; millisecond; neuronal circuitry; neuronal patterning; new technology; non-invasive imaging; novel; operation; optical imaging; relating to nervous system; sapphire laser; scale up; screening; sensor; somatosensory; tool; two-photon; voltage

PROJECT SUMMARY Understanding how information is processed in the mammalian neocortex has been a longstanding question in neuroscience. While the action potential is the fundamental bit of information, how these spikes encode representations and drive behavior remains unclear. In order to adequately address this problem, it has become apparent that experiments are needed in which activity from large numbers of neurons can be measured in a detailed and comprehensive manner across multiple timescales. Direct measurements of action potentials have primarily been achieved by electrophysiology. However, such measurements cannot easily be combined with other methods to assess the connectivity and molecular properties of neurons. Integrating functional, anatomical, and genetic information is critical for understanding how neuronal circuits are organized and computed. There have been long-standing efforts in developing optical methods for measuring neuronal activity due to its compatibility to simultaneously measure connectivity and molecular identity using fluorescent labeling techniques. Newly engineered genetically-encoded voltage-sensitive indicators have now opened the door for optical imaging of action potentials. Two-photon microscopy has been a proven method for deep non-invasive imaging into the brain. However, the fast millisecond transience of action potentials and the membrane localization of genetically-encoded voltage-sensitive indicators both contribute to conditions of limited photon flux. This creates fundamental challenges in the application of two- photon microscopy for voltage imaging that requires scanning at kilohertz frame rates with high signal to noise. To achieve this requires a concerted effort between optical engineers and protein engineers to develop new instrumentation and sensors to arrive at an optimal solution. This multi-investigator effort proposes to advance two-photon microscopy and genetically-encoded voltage-sensitive indicators to enable non-invasive population-level measurements of action potentials with single-cell spatial resolution and single-spike temporal precision deep into the mammalian brain of awake behaving animals.

项目概要 了解哺乳动物新皮质如何处理信息一直是一个长期存在的问题 神经科学。虽然动作电位是信息的基本位，但这些尖峰如何编码 陈述和驾驶行为仍不清楚。为了充分解决这个问题， 很明显，需要进行实验来研究大量神经元的活动 跨多个时间尺度进行详细而全面的测量。直接测量 动作电位主要是通过电生理学来实现的。然而，此类测量不能 很容易与其他方法结合来​​评估神经元的连接性和分子特性。 整合功能、解剖和遗传信息对于理解神经元回路如何进行至关重要 被组织和计算。长期以来，人们一直致力于开发光学方法 测量神经元活动，因为它具有同时测量连接性和分子的兼容性 使用荧光标记技术进行身份识别。新设计的基因编码电压敏感 指标现在为动作电位的光学成像打开了大门。双光子显微镜有 是一种经过验证的对大脑进行深度非侵入性成像的方法。然而，快速的毫秒瞬态 动作电位和基因编码电压敏感指标的膜定位 有助于限制光子通量的条件。这给二元化的应用带来了根本性的挑战。 用于电压成像的光子显微镜，需要以千赫兹帧速率进行扫描，并具有高信噪比。 为了实现这一目标，需要光学工程师和蛋白质工程师共同努力开发新的 仪器和传感器以得出最佳解决方案。这项多位研究者的努力旨在推进 双光子显微镜和基因编码电压敏感指示器，可实现非侵入性 具有单细胞空间分辨率和单尖峰时间的群体水平动作电位测量 精确深入到清醒行为动物的哺乳动物大脑中。