A Cellular Resolution Census of the Developing Human Brain

项目概要/摘要 我们的目标是创建一个时空单细胞分辨率地图的发展人类新皮层，以 确定存在多少不同的细胞类型，并解开它们复杂的发育历史。我们将 我们的分析建立在基于转录组特征的细胞类型的多模式分类上， 在可能的情况下辅以生理和表观遗传特征。我们还将研究瞬时细胞 种群只存在于发育阶段，我们将保留所有细胞的位置信息， 数据来创建发育细胞图谱，该图谱根据细胞类型在细胞中的位置绘制细胞类型的多样性。 人类大脑的成长我们已经开发出了大规模平行分析分子的创新策略， 使用基于液滴的捕获技术的原代人皮质细胞的生理特性， 内容显微镜，和成对的生理反应的转录状态。我们建议进行 在大脑皮层的特定区域，以及纹状体， 丘脑，下丘脑和小脑，我们将使用单核测序来解开发育 传统上很难研究的时间点。我们的项目将揭示细胞的起源 通过解决三个具体目标来研究人类大脑皮层的多样性：1）我们将使用单细胞RNA测序， 询问神经发生和胶质细胞生成如何进行，并在发育的大脑中产生关键细胞类型。 我们假设，促进区域化和连通性的关键事件可以在转录水平上发生。 从怀孕的头三个月到出生后的各个阶段，提供了细胞身份是如何确定的见解。 2)我们对人脑细胞图谱的发展方法提供了一个机会， 在边缘区和亚板区发育早期出现的细胞群， 新生儿阶段据推测，这些细胞类型在建立大脑结构中发挥重要作用， 功能，但它们在人类大脑发育中的特征仍然很差。我们假设 这些群体可以在转录上被识别，并且可以解释这些短暂的 人口数量。3)转录状态是细胞类型鉴定的有力工具，但它们不能捕获细胞的转录状态。 分子特征的全部复杂性。我们将分析细胞特异性激动剂反应和染色质状态， 反映了定义的转录类别内的异质性。我们假设 生理状态和表观遗传状态对转录的影响将为细胞类型分类提供额外的细微差别。 我们的研究结果将为发育中的人脑提供一个细胞分类学的框架， 细胞功能区中分子定义的细胞类型的全面细胞分辨率图 人类大脑在发育过程中这一独特的资源将作为人类大脑研究的蓝图 功能，疾病中细胞类型的选择性脆弱性，以及使我们独特的大脑进化特征。

Project Summary/Abstract We aim to create a spatiotemporal single cell resolution map of the developing human neocortex in order to establish how many distinct cell types are present and to unravel their complex developmental history. We will build our analysis on a multimodal classification of cells types based on transcriptomic signatures but complemented where possible by physiological and epigenetic features. We will also examine transient cell populations present only during developmental stages, and we will retain positional information for all our cell data to create a developmental cell atlas that plots the diversity of cell types according to their locations in the growing human brain. We have developed innovative strategies for massively parallel profiling of molecular and physiological properties of primary human cortical cells using droplet based capture technologies, high content microscopy, and paired physiological responses to transcriptional state. We propose to conduct our integrated cellular survey of developing human brain in specific regions of the cortex, as well as in the striatum, thalamus, hypothalamus and cerebellum, and we will use single nuclei sequencing to unlock developmental time points that have been traditionally difficult to study. Our project will shed light on the origins of cellular diversity in the human cortex by addressing three specific aims: 1) We will use single cell RNA-sequencing to interrogate how neurogenesis and gliogenesis proceed and give rise to key cell types in the developed brain. We hypothesize that key events promoting regionalization and connectivity can be transcriptionally distinguished from the first trimester to postnatal stages, providing insights into how cell identity is determined. 2) Our developmental approach to the human brain cell atlas provides an opportunity to characterize transient cell populations that appear early in development in the marginal zone and subplate regions, and disappear at neonatal stages. These cell types are presumed to play important roles in establishing brain architecture and function, but they remain poorly characterized in developing human brain. We hypothesize the heterogeneity of these populations can be identified transcriptionally and can explain a diverse set of roles for these transient populations. 3) Transcriptional states are a powerful tool for cell type identification, but they do not capture the entire complexity of molecular features. We will profile cell-specific agonist responses and chromatin state that reflect heterogeneity within defined transcriptional classes. We hypothesize that the intersection of physiological state and epigenetic state to transcription will provide additional nuance to cell type classification. Our results will provide a framework of cellular taxonomy in the developing human brain and create a comprehensive cellular resolution map of molecularly defined cell types throughout functional regions of the human brain during development. This unique resource will serve as a blueprint for studies of human brain function, selective vulnerability of cell types in disease, and the features of brain evolution that make us unique.