Investigating the role of Tsc1 in neocortical circuit assembly

描述(申请人提供)：哺乳动物的新皮质是一个六层结构，通过整合感觉信息来创建世界的表征，并通过产生适当的运动输出来控制行为。每一层都以不同的方式处理信息，因此了解特定于层的神经元网络是如何组织的，将为理解不同层在电路中的功能提供基础。内侧前额叶皮质(MPFC)整合了许多脑区的信息，参与复杂的行为，包括社会互动和决策。重要的是，在自闭症等神经发育障碍中，mPFC回路经常被破坏，但人们对mPFC的精细组织或mPFC层如何整合不同类型的信息知之甚少。罗实验室最近开发了一种改进的基于狂犬病的单跨突触追踪技术，除了可以进行全脑远程成像外，还可以对局部电路进行详细分析。这种新的狂犬病技术可以与层特定的Cre驱动鼠系结合起来，生成内侧前额叶皮质(MPFC)的层特定的电路的详细地图。依赖Cre的狂犬病追踪还将与细胞类型特定基因敲除相结合，因此这些图谱将以前所未有的精度和范围为研究发育和疾病中新皮质连接的基因调控提供基础。为了开始剖析建立特定mPFC连接所涉及的分子通路，该项目将重点放在自闭症相关基因TSC1的作用上。最近的研究表明，TSC1的缺失增加了兴奋性突触的连接，改变了兴奋和抑制的平衡，导致海马神经元的高兴奋性。这种表型可能与TSC1在结节性硬化症中的作用有关，结节性硬化症是一种患者患有良性肿瘤、癫痫和自闭症的疾病。在有条件的Tsc1fl/fl小鼠中进行依赖于Cre的狂犬病追踪，将有助于研究TSC1在mPFC连接和层特异性组织发展中的细胞自主作用。由于TSC1是哺乳动物雷帕霉素复合体靶点mTORC1的负调控因子，因此还将研究TSC1在体内发挥作用的分子机制，以阐明这些信号通路是如何调节大脑发育的。这项工作将提供对TSC1突变患者引起自闭症和癫痫的分子和电路水平机制的更深层次的理解。

DESCRIPTION (provided by applicant): The mammalian neocortex is a six-layered structure that creates a representation of the world by integrating sensory information and controls behavior by generating the appropriate motor output. Each layer processes information differently, so understanding how layer-specific neuronal networks are organized will provide a foundation for understanding how different layers function within circuits. The medial prefrontal cortex (mPFC) integrates information from many brain regions and is involved in complex behaviors including social interaction and decision-making. Importantly, mPFC circuits are often disrupted in neurodevelopmental disorders such as autism, but little is known about the fine scale organization of mPFC or how mPFC layers integrate different kinds of information. The Luo lab recently developed a modified rabies-based mono-trans- synaptic tracing technique that allows for detailed analysis of local circuitry in addition to whole brain long- distance mapping. This new rabies technique can be combined with layer-specific Cre driver mouse lines to generate detailed maps of layer-specific circuits in medial prefrontal cortex (mPFC). Cre-dependent rabies tracing will also be combined with cell-type specific gene knockout, so these maps will provide a basis for studying genetic regulation of neocortical connectivity in development and disease with unprecedented precision and scope. To begin to dissect the molecular pathways involved in establishing specific mPFC connectivity, this project will focus on the role of the autism-related gene Tsc1. Recent studies showed that Tsc1 deletion increases excitatory synaptic connectivity and alters the balance of excitation and inhibition, causing hyperexcitability in hippocampal neurons. This phenotype may be related to the role of Tsc1 in Tuberous Sclerosis Complex, a disease in which patients suffer from benign tumors, epilepsy, and autism. Performing Cre-dependent rabies tracing in conditional Tsc1fl/fl mice crossed with layer-specific Cre-drivers will facilitate investigation of the cell-autonomous role o Tsc1 in the development of mPFC connectivity and layer- specific organization. As Tsc1 is a negative regulator of the mammalian target of rapamycin complex, mTORC1, the molecular mechanisms underlying the in vivo function of Tsc1 will also be investigated to elucidate how these signaling pathways regulate brain development. This work will provide a deeper understanding of the molecular and the circuit-level mechanisms that give rise to autism and epilepsy in patients with Tsc1 mutations.