Dendritome mapping of genetically-defined and sparsely-labeled cortical and striatal projection neurons

遗传定义和稀疏标记的皮质和纹状体投射神经元的树突状图谱

• 批准号: 9768581
• 负责人: Hong-Wei Dong
• 金额: $ 84.16万
• 依托单位: UNIVERSITY OF CALIFORNIA LOS ANGELES
• 依托单位国家: 美国
• 财政年份: 2018
• 资助国家: 美国
• 起止时间: 2018-09-01 至 2023-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9768581
• 关键词: 3-Dimensional; Adult; Age; Anatomy; Atlases; Axon; BRAIN initiative; Brain; Cells; Censuses; Classification; Communities; Computer Hardware; Computer software; Corpus striatum structure; Custom; Data; Data Set; Databases; Dendrites; Dendritic Spines; Development; Dyes; Frequencies; Generations; Genetic; Genotype; Golgi Apparatus; Grant; Image; Image Analysis; Imagery; Imaging Device; Initiator Codon; Label; Maps; Membrane; Methods; Mitotic; Molecular; Molecular Profiling; Monsters; Morphology; Mosaicism; Mus; National Institute of Mental Health; Neuroglia; Neurons; Neurosciences Research; Pathway interactions; Population; Presynaptic Terminals; Property; Proteins; RBP4 gene; Reference Standards; Reporter; Resolution; Translations; Virus; Visualization software; Work; base; bioimaging; body position; brain cell; brain research; cell type; computerized tools; cost; data access; data integration; digital; hippocampal pyramidal neuron; image processing; informatics tool; innovation; logarithm; molecular marker; next generation; novel; novel strategies; postnatal; programs; reconstruction; single-cell RNA sequencing; tool

PROJECT SUMMARY Integrating molecular, morphological, and connectomic properties is critical for unbiased classification of neuronal cell types in the mammalian brain. Here we propose a novel approach to classify neuronal cell types by brainwide comprehensive profiling of the dendritic morphology of genetically-defined neurons in the mouse brain. We have developed an innovative mouse genetic tool, called Mosaicism with Repeat Frameshift (or MORF), which enables sparsely and stochastically labeling of genetically-defined neurons in mice. MORF reporter mice can label in exquisite detail single neurons from dendrite and spines to axons and axonal terminals at a labeling frequency of 1-5% of a given neuronal population. We propose to cross our new MORF lines with Cre mouse lines for striatal medium spiny neurons (MSNs) of direct- and indirect pathways, and for cortical pyramidal neurons of distinct cortical layers (i.e. L2/3/4, L5 and L6). Each MORF/Cre mouse will allow us to image the detailed dendritic morphology for thousands of genetically-defined striatal and cortical neurons (i.e. dendritome). We have also developed and streamlined imaging and computational tools to acquire and register brainwide single neuron morphological data onto a standard reference mouse brain atlas. We will digitally reconstruct hundreds of thousands of MORF-labeled neurons using our novel program called G-Cut. Reconstructed neurons will subsequently used for morphology based clustering to define new morphological subtypes, which in turn can be analyzed for the expression of novel molecular markers neuronal cell types (e.g. from single cell RNA-sequencing). Finally, we will disseminate the data to the Brain Cell Data Center (BCDC) for data integration with those from other BRAIN Initiative Cell Census Network (BICCN) and for data access by the broader neuroscience research community. In addition to dendritome data generation and analyses, we will further advance our MORF method by generating new MORF reporter mouse lines with logarithmic fold decrease in the Cre- dependent labeling frequencies, which will permit imaging of the complete, brainwide morphology of genetically-defined single neurons that include both dendritic and axonal arborization. Such tool should greatly facilitate the neuronal morphology based cell type classification. Finally, we will develop integrated computer hardware and software for domain-specific computing for automated image processing and neuronal reconstruction, a major bottleneck in analyzing large bioimage datasets. Altogether we will provide rich dendritome information to enable unbiased, morphology-based neuronal cell type classification, and novel mouse genetic tools and computer software and hardware to advance the field of large-scale neuronal morphological imaging and analyses for the comprehensive study of the mammalian brain.

项目总结 整合分子、形态和连接性属性对于无偏倚至关重要 哺乳动物脑内神经细胞类型的分类。在这里，我们提出了一种新的方法 通过脑内树突状细胞形态的全面描述对神经细胞类型进行分类 小鼠大脑中由基因定义的神经元。我们开发了一种创新的老鼠遗传工具， 称为带有重复移码的马赛克(或Morf)，它可以实现稀疏和随机的标记 小鼠的基因定义的神经元。Morf报告鼠可以精细地标记来自 树突和棘突至轴突和轴突终末的标记频率为给定神经元的1-5% 人口。我们建议将我们的新的Morf系与CRE小鼠系杂交，以寻找纹状体中棘神经元 (MSN)直接和间接通路，以及不同皮质层的皮质锥体神经元(即 L2/3/4、L5和L6)。每只Morf/Cre小鼠将允许我们成像详细的树突形态 数以千计的基因定义的纹状体和皮质神经元(即树突)。我们还开发了和 用于获取和记录全脑单个神经元的简化成像和计算工具 将形态数据复制到标准参考小鼠脑图谱。我们将以数字方式重建数百个 数以千计的Morf标记神经元使用我们名为G-Cut的新程序。重建的神经元将 随后用于基于形态的聚类以定义新的形态子类型，该新形态子类型进而 可以分析新的分子标记神经细胞类型的表达(例如来自单细胞 RNA测序)。最后，我们将把数据发布到脑细胞数据中心(BCDC)进行数据 与其他大脑倡议细胞普查网络(BICCN)的数据集成，并通过以下方式访问数据 更广泛的神经科学研究社区。 除了树状基因组数据的生成和分析之外，我们还将进一步推进我们的Morf 方法通过产生Cre-Cre对数倍递减的新的Morf报告小鼠品系 依赖的标记频率，这将允许成像完整的，全脑的形态 基因定义的单个神经元，包括树突和轴突分枝。这类工具应 极大地方便了基于神经元形态的细胞类型分类。最后，我们将发展 用于自动图像的领域特定计算的集成计算机硬件和软件 处理和神经元重建，这是分析大型生物图像数据集的主要瓶颈。 总之，我们将提供丰富的树突组信息，以实现无偏见的、基于形态的神经元 细胞类型分类，新型小鼠遗传工具和计算机软硬件的进步 大尺度神经元形态成像与分析领域的综合研究 哺乳动物的大脑。