BRAIN EAGER: Panoramic, dynamic, multi-region two-photon microscopy for systems neuroscience

该奖项由生物基础设施部（DBI； BIO理事会）的生物研究仪器开发计划（IDBR）授予。在哺乳动物的进化过程中，作为大脑外层的大脑皮层在表面上急剧膨胀。这一皮层区域被划分为独立的功能区，包括视觉皮质、运动皮质和语言区。这些大脑区域协同行动来支持行为。尽管我们已经了解了如何将功能归因于特定的大脑区域，但我们对进行这种协调的细胞机制知之甚少。模型系统，包括老鼠，在它们的大脑中有独立的功能区域。然而，目前神经科学家用于研究大脑活动的工具要么局限于分布在大面积上的稀疏神经元样本，要么局限于在500-700微米宽的单个区域内的大密度神经元。因此，在理解皮层区域如何协同行动以支持行为方面取得进展是非常困难的。拟议中的研究项目将开发一种新型显微镜，它将能够在几毫米的视野范围内检测单个神经元的峰值。这个区域可以包括老鼠的五个或更多的皮质区域。此外，该显微镜将包含高速聚光灯，用于同时成像多个皮质区域的神经元活动。这种时间分辨率对于理解神经元编码的信息、它们在行为过程中的动态以及它们的连通性至关重要。将在美国和全球范围内培养一个科学家社区，以传播这项研究，帮助其实施，并加速神经科学领域的合作进展。还将举办讲习班，培训先进光学和神经科学方面的科学家。最终，该项目将提供对BRAIN计划至关重要的新技术，并将促进更广泛的科学界在双光子成像领域取得进一步进展。研究小组将开发一种双光子（2p）成像系统，具有宽视场（FOV）（~ 3毫米）和跨全视场的细胞分辨率。为了确保记录活动的高时间分辨率，他们还将开发多路波束，在视场内高速成像大脑区域。这些多路复用光束可以动态重新配置，以瞄准全视场内的不同区域，如聚光灯。该方法是对整个系统建模，并创建优化的光学组件，包括自定义物镜。该团队将进行计算工程权衡，以保持蜂窝分辨率，同时仍然实现宽视场。高速扫描将采用共振扫描仪和光子计数电子学。该系统具有可扩展性，光束复用可以模块化，多个模块可以堆叠以增加光束数量，只要荧光寿命短于激光脉冲间隔。因此，Trepan2p（双区域，全景2p）将能够直接测量扩展大脑网络中的相互关联和实时动态。这项技术将使以前不可能的实验成为可能，在包括小鼠和灵长类动物在内的一系列模型系统中，以单细胞分辨率在扩展的神经回路中成像神经元活动。

This award is made by the Instrument Development for Biological Research program (IDBR)in the Division of Biological Infrastructure (DBI; BIO Directorate).The cerebral cortex, the outer layer of brain, has greatly expanded in surface are during mammalian evolution. This cortical region is parcellized into discrete functional areas including visual cortex, motor cortex, and language areas. These brain areas act in concert to support behavior. Although we have learned much about how to ascribe function to particular brain areas, we know little about the cellular mechanisms by which this concert is conducted. Model systems, including mice have discrete functional areas in their brains. However, the current tools that neuroscientists have for investigating activity in brains are limited to either a sparse sampling of neurons distributed over large areas, or a large density of neurons in a single area just 500-700 microns across. Thus, it is tremendously difficult to make progress in understanding how cortical areas act in concert to support behavior. The proposed research project will develop a new type of microscope which will be able to detect single neuron spiking across a field of view of several millimeters. This area can encompass five or more cortical areas in a mouse. In addition, this microscope will contain high speed spotlights for simultaneously imaging neuronal activity in multiple cortical areas. This time resolution is crucial for understanding the information neurons encode, their dynamics during behavior, and their connectivity. A community of scientists across the US and the globe will be cultivated to disseminate the research, aid in its implementation, and accelerate collaborative progress in neuroscience. Workshops will also be held to train scientists in advanced optics and neuroscience. Ultimately, this project will provide new technology that is crucial for the BRAIN Initiative, and will foster a broader scientific community for further progress in the field of two-photon imaging. The research team will develop a two photon (2p) imaging system with a wide field-of-view (FOV) (~ 3 mm) and cellular resolution across the full FOV. To ensure high temporal resolution of recorded activity, they will also develop multiplexed beams that image brain regions within the FOV at high speed. These multiplexed beams can be dynamically reconfigured to target different areas within the full FOV, like spotlights. The approach is to model the full system and create optimized optical subassemblies, including a custom objective. The team will make calculated engineering tradeoffs to preserve cellular resolution while still achieving a wide FOV. High speed scanning will be developed using resonant scanners and photon counting electronics. This system is scalable, as the beam multiplexing can be modularized, and multiple modules can be stacked to increase the number of beams, so long as the fluorescence lifetime is shorter than the interval between laser pulses. Thus, the Trepan2p (Twin-Region, Panoramic 2p), will enable direct measurements of cross-correlations and moment-to-moment, dynamics in extended brain networks. This technology will enable previously impossible experiments, imaging neuronal activity with single cell resolution across extended neuronal circuitry in an array of model systems including mice and primates.