Investigating role of neurexin-1 mutation in autism using human induced neurons

描述（由申请人提供）：本提案的目的是提高我们对自闭症谱系障碍（ASD）病理生理学的有限理解，ASD是一种破坏性的神经发育状况，影响88名美国儿童中的1名。这将通过研究从人类胚胎干细胞（ESC）诱导的人类神经元来实现，这些胚胎干细胞有条件地敲除neurexin-1（Nrxn 1），这是一种在ASD患者中经常突变的基因。Nrxn 1的突触作用及其在人类神经元背景下对自闭症的功能贡献尚未研究，这使得这项研究独一无二。初步数据表明，这种方法的可行性，从而胚胎干细胞的条件突变的Nrxn 1基因座已产生。使用这些细胞，将产生人类兴奋性神经元，从而在神经元发育期间有条件地敲除Nrxn 1基因。基于Nrxn 1是人类神经元正常突触功能所必需的假设，我将比较对照和Nrxn 1突变人类神经元的突触发育和功能（突触传递）。nrxn 1在突触发育中的功能将通过检查突触密度和大小以及突触囊泡循环的形态学研究来监测。使用，电生理分析，我将评估前和突触后的功能Nrxn 1在人类兴奋性突触介导的突触传递。总之，这些实验试图确定突变的Nrxn 1影响人类突触传递的机制，并因此导致自闭症，使用评估突触功能的实验方法的组合。!

DESCRIPTION (provided by applicant): The object of this proposal is to enhance our limited understanding of the pathophysiology of autism spectrum disorders (ASDs), which is a devastating neurodevelopmental condition affecting 1 in 88 US children. This will be accomplished by studying human neurons induced from human embryonic stem cells (ESCs) that are conditionally knock out for neurexin-1 (Nrxn1), a gene that is frequently mutated in patients with ASDs. The synaptic role of Nrxn1 and its functional contribution to autism in human neuronal context has not been studied, which makes this study unique. Preliminary data indicates the feasibility of this approach, whereby ESCs conditionally mutant for Nrxn1 locus has been generated. Using these cells, human excitatory neurons will be generated whereby Nrxn1 gene will be conditionally knocked out during neuronal development. Based on the hypothesis that Nrxn1 is required for proper synaptic function in human neurons, I will compare synapse development and function (synaptic transmission) in control and Nrxn1 mutant human neurons. Nrxn1 function in synapse development will be monitored by morphological studies examining synapse density and size as well as synaptic vesicle cycling. Using, electrophysiolgical assays, I will assess both pre and postsynaptic functions of Nrxn1 in mediating synaptic transmission at human excitatory synapses. In total, these experiments attempt to determine the mechanism by which mutated Nrxn1 influence human synaptic transmission and as a consequence, contribute to autism, using a combination of experimental approaches that assess synaptic function. !