Genetic access to cortical cell types with epigenetic assays and high-throughput, low-cost enhancer screening

项目摘要 神经系统的功能依赖于神经元网络之间的复杂相互作用 由多种神经元类型组成。了解这些网络如何在健康和疾病中发挥作用 依赖于了解特定神经元类型与其功能之间的精确连接 完整大脑中的相互作用。因此，很明显，为了充分了解 神经系统，有必要有详细的神经元连接的描述与相同水平的 这些系统运行的精确度，并选择性地操纵和测量特定细胞的活动 在正常运行的网络环境中键入。这样的研究将极大地便利于能够 在AAV载体的背景下，针对特定细胞类型的靶基因表达。这里提出的研究是 旨在开发和使用一种新的高通量策略和聚合酶链式反应激活的细胞分选 识别细胞类型特定的增强子元件。所使用的方法将确定可以推动 转基因在特定类型的皮质神经元中的表达。进一步的研究将使这些增强剂在 实际使用环境，以便于使用遗传编码的钙指示剂进行成像并促进 光遗传失活。这些试剂将允许未来的研究测试特定的功能贡献 抑制神经元类型对知觉、认知和行为的影响。

Project Summary The function of the nervous system is dependent on complex interactions between networks of neurons composed of multiple neuron types. Understanding how these networks function both in health and disease is dependent on understanding the precise connectivity between specific neuron types and their functional interactions in the intact brain. It is therefore apparent that, in order to have an adequate understanding of the nervous system, it is necessary to have detailed descriptions of neuronal connectivity with the same level of precision at which these systems operate and to selectively manipulate and measure the activity of specific cell types in the context of the normal functioning network. Such studies would be greatly facilitated by the ability to target gene expression to specific cell types in the context of AAV vectors. The research proposed here is aimed at developing and using a novel high throughput strategy and PCR-activated cell sorting for identification of cell type specific enhancer elements. The approach used will identify enhancers that can drive transgene expression in specific types of cortical neurons. Further studies will valdiate these enhancers in a real-use context for ability to facilitate imaging with genetically-encoded calcium indicators and to facilitate optogenetic inactivation. These reagents will allow future studies testing the functional contributions of specific inhibitory neuron types to perception,cognition and behavior.