Fast, large-scale neuronal imaging with multi-z confocal microscopy

使用多 Z 共聚焦显微镜进行快速、大规模神经元成像

基本信息

批准号：
10524735
负责人：
Jerome Mertz
金额：
$ 44.14万
依托单位：
BOSTON UNIVERSITY (CHARLES RIVER CAMPUS)
依托单位国家：
美国
项目类别：
财政年份：
2020
资助国家：
美国
起止时间：
2020-02-01 至 2024-11-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10524735
关键词：
3-Dimensional Address Animals Behavior Biology Brain imaging Calcium Cannulas Collection Communication Communities Computer software Confocal Microscopy Data Detection Development Devices Effectiveness Engineering Exhibits Fluorescence Generations Genetic Goals Head Image Lasers Light Lighting Microscope Microscopy Modality Modification Monitor Mus Neurobiology Neurons Optics Penetration Performance Photobleaching Phototoxicity Population Preparation Protocols documentation Reaction Time Research Resolution Sampling Scanning Signal Transduction Speed Spottings Techniques Technology Testing Time Viral design effectiveness evaluation experience fluorescence microscope improved in vivo in vivo calcium imaging instrument interest lens millimeter millisecond multi-photon neuroregulation neurotransmission novel optogenetics prototype recruit reflectance confocal microscopy sample fixation spatiotemporal technology development user-friendly voltage

项目摘要

ABSTRACT Neuronal signals can vary on millisecond timescales, with communicating neurons often separated by hundreds of microns. Imaging such fast dynamics over extended volumes presents a challenge for standard fluorescence microscopes. For example, a new generation of genetically encoded voltage indicators are becoming available whose response times are on the order of milliseconds. To address this challenge, we propose to develop a new type of microscope that can perform near- 1kHz-rate high resolution volumetric imaging over 1mm x 1mm x 0.2mm scales. Our proposed solution, called Multi-Z confocal microscopy, is based on two key ideas. First, it combines high-NA detection with low-NA illumination. The former leads to high signal collection efficiency; the latter leads to axially extended illumination over an extended range of Z depths. Second, it detects multiple signals from this extended depth range using multiple confocal pinholes that are axially distributed. The pinholes are reflecting, so that signal rejected by one pinhole is sent to the next pinhole, and so forth. In this manner, no signal is lost, and signal collection efficiency remains high. Two versions of our microscope will be developed, based on line-scan and sheet-scan illumination. The former provides better optical sectioning and will be designed for calcium imaging. The latter provides much higher speed (near kHz-rate) and will be designed for voltage imaging. In contrast to conventional line-scan or light-sheet microscopes, our lines and sheets are oriented parallel to the optical axis rather than perpendicular to the axis. The versatility of both versions of our microscope will be augmented with the addition of optogenetic stimulation and combined confocal reflectance contrast. We have enlisted the help of Drs. Alberto Cruz-Martin (BU, Biology) and Xue Han (BU, BME), who both specialize in in-vivo mouse imaging and have expertise in the genetic or viral delivery of novel probes, animal preparation, head fixation, behavior protocols, etc.. For voltage imaging, we will test a state-of-the-art indicator called SomArchon1 (provided by the Dr. Ed Boyden lab). The ability to image large sample volumes at 1kHz rates is of general applicability and can be broadly impactful. Our goal will be to demonstrate the effectiveness of our Multi-Z microscopy technique by performing calcium and voltage imaging over entire populations of neurons in behaving mice.

抽象的神经元信号可以在毫秒时间尺度上变化，通信神经元通常被数百微米。在大体积范围内对如此快速的动态进行成像对标准荧光显微镜。例如，新一代基因编码电压响应时间为毫秒级的指标正在变得可用。为了应对这一挑战，我们建议开发一种新型显微镜，可以进行近乎在 1mm x 1mm x 0.2mm 尺度上进行 1kHz 速率高分辨率体积成像。我们提出的解决方案，称为多 Z 共焦显微镜，基于两个关键思想。首先，它将高数值孔径检测与低数值孔径照明。前者信号采集效率高；后者导致轴向在更大的 Z 深度范围内提供更大的照明。其次，它检测到来自此的多个信号使用轴向分布的多个共焦针孔扩展深度范围。针孔是反射，使得被一个针孔拒绝的信号被发送到下一个针孔，依此类推。以这种方式，无信号丢失，信号采集效率保持较高。我们将开发基于线扫描和片扫描照明的两种版本的显微镜。前者提供更好的光学切片，专为钙成像而设计。后者提供更高的速度（接近 kHz 速率），并将设计用于电压成像。相比之下传统的线扫描或光片显微镜，我们的线和片平行于光学方向轴而不是垂直于轴。我们两个版本显微镜的多功能性将是通过添加光遗传学刺激和组合共焦反射对比度来增强。我们已经获得了博士的帮助。 Alberto Cruz-Martin（生物系）和Xue Han（生物系）两者都专注于小鼠体内成像，并且在新型药物的遗传或病毒传递方面拥有专业知识探针、动物准备、头部固定、行为协议等。对于电压成像，我们将测试最先进的指标，称为 SomArchon1（由 Ed Boyden 博士实验室提供）。形象化能力 1kHz 速率下的大样本量具有普遍适用性，并且可以产生广泛的影响。我们的目标将通过执行钙和对行为小鼠的整个神经元群进行电压成像。

项目成果

期刊论文数量（7）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（0）

Correcting sampling bias in speckle contrast imaging.

纠正散斑对比度成像中的采样偏差。

DOI：
发表时间：
2022-12-15
期刊：
影响因子：
3.6
作者：
Zheng, Shuqi;Mertz, Jerome
通讯作者：
Mertz, Jerome

High-contrast multifocus microscopy with a single camera and z-splitter prism.

配备单相机和 z 分光棱镜的高对比度多焦点显微镜。

DOI：
发表时间：
2020-11-20
期刊：
影响因子：
10.4
作者：
Xiao, Sheng;Gritton, Howard;Tseng, Hua;Zemel, Dana;Han, Xue;Mertz, Jerome
通讯作者：
Mertz, Jerome

Fast, multiplane line-scan confocal microscopy using axially distributed slits.

使用轴向分布狭缝的快速多平面线扫描共焦显微镜。

DOI：
发表时间：
2021-03-01
期刊：
影响因子：
3.4
作者：
Tsang, Jean;Gritton, Howard J;Das, Shoshana L;Weber, Timothy D;Chen, Christopher S;Han, Xue;Mertz, Jerome
通讯作者：
Mertz, Jerome

Direct characterization of tissue dynamics with laser speckle contrast imaging.

利用激光散斑对比成像直接表征组织动力学。