High-throughput mapping of synaptic connectivity between transcriptomically defined cell types

项目摘要 识别构成大脑每个区域的细胞类型和突触连接的模式 是理解神经回路如何产生所有感知，认知， 和行为。光学、分子和计算技术的快速进步使大规模的 大规模项目旨在全面绘制构成哺乳动物大脑的细胞类型。 然而，定义大脑中数千种细胞类型的微连接仍然具有挑战性 由于缺乏可扩展的方法。该提案描述了解决这一问题的技术的发展 方法上的差距使用高灵敏度荧光电压成像和单- 神经元光遗传学光刺激，我们将映射突触连接内和之间的大脑区域。 使用这种方法，可以映射突触连接，吞吐量为两到三个数量级 高于现有技术。重要的是，利用全光学方法来映射连接将 使我们能够将突触连接测量与新兴技术相结合， 荧光原位杂交通过这种方式，我们可以识别大神经元的分子身份， 人口及其连通性。我们将在以下文件中演示本提案中开发的技术： 运动皮层，一个对基因表达模式的了解远远超过我们识别能力的区域， 连通性图案。揭示了局部连接基序和细胞的精确分子身份， 接受来自丘脑的长距离输入--皮层活动的重要驱动力--将提供新的 深入了解支持自主运动的电路机制。

PROJECT SUMMARY Identifying the cell types that make up each region of the brain and the patterns of synaptic connections through which they are linked is key to understanding how neural circuits give rise to all perception, cognition, and behavior. Rapid improvements in optical, molecular, and computational technologies are enabling large- scale projects aiming to comprehensively map the cell types that comprise the mammalian brain. Nevertheless, defining the microconnectivity of the thousands of cell types in the brain remains challenging due to a lack of scalable methods. This proposal describes the development of a technology for addressing this methodological gap. Using a novel combination of high-sensitivity fluorescence voltage imaging and single- neuron optogenetic photostimulation, we will map synaptic connectivity within – and between – brain regions. Using this approach, synaptic connectivity can be mapped with throughput two to three orders of magnitude higher than existing techniques. Importantly, leveraging an all-optical approach to mapping connectivity will allow us integrate synaptic connectivity measurements with emerging techniques for highly multiplexed fluorescence in situ hybridization. In this way, we can identify the molecular identities of large neuronal populations and their connectivity. We will demonstrate the technology developed in this proposal in the motor cortex, a region where knowledge of gene expression patterns has far outpaced our ability to identify connectivity motifs. Revealing both local connectivity motifs and the precise molecular identity of cells that receive long-range input from the thalamus – an important driver of cortical activity – will provide new insights into the circuit mechanisms supporting voluntary movements.