Development of a scalable strategy for reconstructing cell-type determined connectome of the mammalian brain

摘要 为了充分理解大脑功能的结构基础， 相同神经元的多种属性。重要的属性包括但不限于形态， 连接特性和分子模式，如功能基因的表达和 突触密度的分布。基于光学显微镜的神经元追踪有助于我们 对神经元形态异质性的基本认识。排列形态学 将来自多个大脑样本的极其稀疏标记的神经元重建到一个共同的 坐标系统允许系统比较的感受性树突领域和投影 不同神经元亚型的轴突乔木。然而，神经元的连接伙伴不能被 由于稀疏采样的性质， 有限的分辨率。另一方面，基于电子显微镜的重建标记了所有细胞， 无差别，但允许识别超细结构，包括突触接触，这在理论上 可以映射所有神经元之间的完整连接体。但是，从技术上讲， 经济上具有挑战性的规模无损电子显微镜的大样本，如小鼠 个脑袋此外，在样品处理过程中，分子信息经常丢失。在这个项目中，我们建议 开发一种可扩展的策略，结合几种新技术， 通过光学显微镜观察哺乳动物大脑的分子连接体。能够同时分析 整个小鼠脑的神经元形态、连接性和分子特性将 阐明我们对脑细胞异质性的理解，并推进我们对脑细胞异质性的认识。 哺乳动物大脑的整体结构，具有前所未有的分辨率和规模。

Abstract In order to fully understand the structural substrates underlying the brain function, it is central to curate multiple attributes of the same neurons. The important attributes include, but limited to morphology, connection properties and molecular patterns, such as the expression of functional genes and the distribution of synaptic densities. Light microscopy-based neuronal tracing has contributed our fundamental understanding of the heterogeneity of neuronal morphology. Aligning morphology reconstructions of extremely sparsely labeled neurons from multiple brain samples onto a common coordinate system permitted systematic comparison of the receptive dendritic fields and the projective axonal arbors of distinct neuronal subtypes. However, connection partners of neurons cannot be identified from the light microscopy-based reconstructions due to the nature of sparse sampling and limited resolution. On the other hand, electron microscopy-based reconstructions label all the cell nondifferential but permit recognition of ultra-fine structures, including synaptic contacts, which in theory permits mapping the complete connectome between all neurons. However, it is technically and economically challenging to scale lossless electron microscopy to a large sample, such as the mouse brain. In addition, molecular information is often lost in sample processing. In this project, we propose to develop a scalable strategy that combines several novel technologies to permit reconstructing a molecular connectome of the mammalian brain by light microscopy. Being able to simultaneous profile neuronal morphology, connectivity and molecular properties throughout the whole mouse brain will clarify our understanding of the heterogeneity of brain cells and advance our knowledge about the overall architecture of the mammalian brain with unprecedented resolution and scale.