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High-throughput measurement of neuronal projections and synapses using Synapse-seq

使用 Synapse-seq 对神经元投射和突触进行高通量测量

基本信息

批准号：
10640361
负责人：
Evan Z Macosko
金额：
$ 175.03万
依托单位：
BROAD INSTITUTE, INC.
依托单位国家：
美国
项目类别：
财政年份：
2023
资助国家：
美国
起止时间：
2023-04-01 至 2026-03-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10640361
关键词：
Address Alzheimer&apos s Disease Axon Bar Codes Biological Assay Brain Cells Consumption Data Dendritic Spines Development Distal Genetic Recombination Genomics Hippocampus Human In Situ In Vitro Individual Label Maps Measurement Measures Messenger RNA Methods Modernization Molecular Nervous System Physiology Neurons Neurosciences Pathway Analysis Pattern Preparation Presynaptic Terminals RNA Reporter Resolution Sensory Deprivation Series Slide Somatosensory Cortex Spatial Distribution Stereotyping Synapses Synaptosomes System Technology Thalamic structure Time Tissues Vertebral column Vibrissae Viral Virion Work arm body position cell type density digital entorhinal cortex in situ sequencing in vivo innovation mouse model neural neural circuit new technology novel postsynaptic presynaptic sequencing platform single cell sequencing technology development technology validation tool trafficking transcriptomics viral DNA

项目摘要

SUMMARY The brain is composed of thousands of highly specialized cell types that form very specific synaptic connections with each other. Together, these connections form neural circuits that are the structural basis of brain function. Despite their importance, synaptic connections amongst cell types are largely unascertained, because of the dearth of existing tools to do so. In this project, we will develop a suite of genomics-based tools--collectively termed Synapse-seq--that allow us to quantify neuronal projections, postsynaptic densities, and synaptic connections in individual cells in vivo. First, we will use AAV to introduce a molecular tool that trafficks a barcoded mRNA reporter to the presynaptic compartments of infected cells, combined with single-cell sequencing and spatial transcriptomics (Slide-seq) to jointly match cell body positions to projection targets. Next, we will use a postsynaptic targeting system, also delivered by AAV, combined with synaptosome preparations and in situ sequencing to quantify spine densities across thousands of single cells. Finally, we will combine our pre- and postsynaptic labeling systems with novel in situ sequencing readouts to measure synaptic connectivity in vivo. Together, the successful development of these technologies will provide neuroscientists with a suite of powerful tools to routinely measure fundamental aspects of neural connectivity in vivo.

总结大脑是由数千种高度特化的细胞组成的，彼此之间的突触连接这些连接一起形成了神经回路，大脑功能的结构基础尽管它们很重要，但突触之间的联系细胞类型在很大程度上是不确定的，因为缺乏现有的工具来这样做。在这项目，我们将开发一套基于基因组学的工具-统称为Synapse-seq-，使我们能够量化神经元投射，突触后密度和突触连接，体内的单个细胞。首先，我们将使用AAV引入一种分子工具，将条形码mRNA报告基因与感染细胞的突触前区室结合，单细胞测序和空间转录组学（Slide-seq）联合匹配细胞体位置投射目标。接下来，我们将使用同样由AAV递送的突触后靶向系统，结合突触体制备和原位测序来量化棘密度在数千个单细胞中。最后，我们将联合收割机结合我们的突触前和突触后标记新的原位测序读出系统，以测量体内突触连接。总之，这些技术的成功开发将为神经科学家提供一个一套强大的工具，用于常规测量体内神经连接的基本方面。