Developing cell type-specific enhancers and connectivity mapping pipelines for marmosets

项目总结 尽管基因工具极大地提高了我们对大脑功能的理解，但它们 在很大程度上仅限于老鼠。虽然小鼠是神经科学许多领域的基本模型，但也有 高级大脑功能的许多方面不能在啮齿类动物身上充分建模。同样，许多大脑 紊乱会影响高级认知功能，而这些功能在啮齿类动物中没有明显的相似之处。此外，最近的大型- Scale单细胞转录分析揭示了许多神经元类型、连接和基因表达 灵长类动物特有的图案。因此，迫切需要有大脑的新的遗传模型。 结构和功能更接近人类。非人灵长类动物(NHP)与人类的亲缘关系要密切得多 这一点反映在它们的大脑发育、结构和生理上。因此，它是 越来越多的人认识到，它们为研究高级大脑功能和大脑疾病提供了一个有吸引力的模型。 一种很有前途的NHP模型是普通的绒猴，一种小型的新世界灵长类动物，有许多 神经科学和基因研究的优势。然而，缺乏具有细胞类型特异性的工具一直是一种 推进NHP结构和功能研究的主要障碍。与组合的单细胞rna-seq 和单细胞ATAC-seq，现在可以提名短细胞类型特异性增强子序列。如果 经过验证，这些增强功能将提供一种有效的工具来映射连通性和使用 病毒介导的表达。困难在于从数百个或更多的功能增强剂中确定 NHP中数千个被提名的潜在增强子序列。在这里，我们建议(1)使用一种新的高 体内吞吐量方法，以确定功能增强子，以及(2)建立全脑回路图 用于使用纹状体电路的管道来验证我们的方法用于细胞类型特定的连接映射 绒猴。这些研究完成后，将提供急需的基本工具、方法和 用于绒猴大脑细胞类型特定映射和功能询问的计算管道 健康和疾病模型。

PROJECT SUMMARY Although genetic tools have dramatically advanced our understanding of brain function, they have largely been confined to mice. While mice are essential models for many areas of neuroscience, there are also many aspects of higher brain function that cannot be adequately modeled in rodents. Similarly, many brain disorders affect higher cognitive functions that have no clear parallels in rodents. Furthermore, recent large- scale single cell transcriptomic analyses have revealed many neuron types, connections and gene expression patterns that are unique to primates. Thus, there is an urgent need for new genetic models that have brain structure and function closer to humans. Non-human primates (NHP) are much more closely related to humans than are rodents, and this is reflected in their brain development, structure and physiology. Hence, it is increasingly recognized that they provide an attractive model to study higher brain function and brain disorders. A promising emerging NHP model is the common marmoset, a small new world primate that has many advantages for neuroscience and genetic research. However, lack of tools with cell type specificity has been a major obstacle in advancing structural and functional studies in NHP. With the combined single cell RNA-seq and single cell ATAC-seq, it is now possible to nominate short cell type-specific enhancer sequences. If validated, these enhancers will provide an effective tool to map connectivity and interrogate function using virus mediated expression. The difficulty lies in the identification of functional enhancers from the hundreds or thousands of nominated potential enhancer sequences in NHP. Here we propose (1) to use a novel high throughput in vivo approach to identify functional enhancers, and (2) to establish a whole-brain circuit mapping pipeline for use striatal circuitry to validate our approach for cell type-specific connectivity mapping in marmosets. When completed, these studies will provide much needed essential tools, methods and computational pipelines for cell type-specific mapping and functional interrogation of the marmoset brain in healthy and disease models.