Cell Type-specific Anterograde Circuit Mapping and Functional Control by Optimizing YFV-17D Transneuronal Systems

细胞类型特定的顺行电路映射和功能控制 优化YFV-17D跨神经元系统 摘要 为了阐明大脑回路的功能组织，我们需要描绘神经元 连接并控制具有特定连接性的神经元的活动。这两项任务都有 越来越依赖于跨神经元的病毒载体。顺行跨神经元病毒载体，它 从突触前神经元扩散到突触后神经元，可以揭示神经元的投射 并选择性地瞄准突触后神经元。由于缺乏理想的顺行跨神经元 病毒载体，我们在过去的几年里一直在开发一种新的顺行系统，该系统基于 黄热病疫苗-YFV-17D。我们已经成功地构建了两个主要的顺行工具： 用于定位神经元投射的包装缺陷YFV∆CME和复制缺陷 YFV∆NS1(或YFV∆CMENS1)用于跨神经元控制突触后神经元的基因表达， 可用于功能观察或操作。神经元毒性可以避免 在YFV∆NS1和YFV∆CMENS1的情况下，但在YFV∆CME中不是。这些新系统提供了以下好处 跨神经元的功效，多样化的应用，以及工程的简易性，但它们仍然具有 许多实验场景的限制。 在这里，我们建议进一步改进这些病毒系统，使其广泛适用， 强大而安全的工具，用于大脑回路的功能解剖。首先，我们将构建自我 限制YFV-17D以进一步减少病毒复制在神经元中引起的神经元毒性， 这将使这些载体在研究和潜在的临床应用中更加有用。 其次，我们将测试多种策略，以将该系统定向到特定神经元的细胞类型 局部电路和远程投影中的特定跟踪或功能控制。其中一些 特定细胞类型的工具将不依赖于转基因小鼠系的可用性，并且可以 适用于广泛的物种。第三，我们将整合常用的标签、传感器或效应器 在优化的跨神经元病毒载体中具有广泛的应用。我们还将应用 改进的版本，以描绘选定的神经元电路，这是非常感兴趣的 神经学家。因此，该项目将产生一套功能强大的工具，广泛适用于 神经科学研究，并将揭示多种经典神经元类型的投射。

Cell Type-specific Anterograde Circuit Mapping and Functional Control by Optimizing YFV-17D Transneuronal Systems Summary To elucidate the functional organization of brain circuitry we need to delineate neuronal connectivity and control the activity of neurons with specific connectivity. Both tasks have increasingly relied on transneuronal viral vectors. Anterograde transneuronal viral vectors, which spread from presynaptic neurons to the postsynaptic neurons, can reveal the neuronal projections and selectively target postsynaptic neurons. Due to the lack of ideal anterograde transneuronal viral vectors, we have spent the last few years developing a new anterograde system based on yellow fever vaccine—YFV-17D. We have successfully constructed two major anterograde tools: the packaging defective YFV∆CME for mapping neuronal projectomes, and the replication defective YFV∆NS1 (or YFV∆CMENS1) for transneuronal control of gene expression in postsynaptic neurons, which can be used for functional observation or manipulation. Neuronal toxicity can be avoided in the case of YFV∆NS1 and YFV∆CMENS1, but not in YFV∆CME. These new systems provide benefits of transneuronal efficacy, diverse applications, and ease of engineering, but they still have limitations for many experimental scenarios. Here we propose to further improve these viral systems to make them broadly applicable, powerful and safe tools for functional dissection of the brain circuits. Firstly, we will construct self- constrained YFV-17D to further minimize neuronal toxicity incurred by viral replication in neurons, which will make these vectors more useful for both research and potential clinical applications. Secondly, we will test multiple strategies to target this system to specific neurons for cell-type specific tracing or functional control in both local circuits and long-range projections. Some of the cell type specific tools will not rely on the availability of genetically modified mouse lines and can be applied to broad species. Thirdly, we will incorporate commonly used tags, sensors or effectors in the optimized transneuronal viral vectors for versatile applications. We will also apply the improved versions to delineate a selected neuronal circuit that is of great interest to neuroscientists. Therefore, this project will yield a set of highly powerful tools widely applicable to neuroscience research, and will reveal the projectomes of multiple classical neuronal types.