Molecular Logic Sculpting Cell-Specific Contributions of Neurexin-1 at the Tripartite Synapse

摘要 大量证据表明，编码神经毒素-1(NRXN1)的基因存在拷贝数变异(CNV)， 突触细胞黏附分子在自闭症谱系障碍、精神分裂症和抽动症发病机制中的作用 综合症。然而，关于NRXN1在发育和成体中的功能和调控，人们知之甚少 突触，对它的了解可能会提高我们对大脑疾病的了解，并揭示新的治疗方法 目标。我的初步数据显示，NRXN1不仅在神经元中表达，而且在星形胶质细胞中也有表达， 挑战NRXN1的经典观点，认为它完全是突触前的。在这里，我建议星形细胞NRXN1 调节星形胶质细胞和突触之间正常所需的一种重要形式的通讯 兴奋性突触的发育和功能。我的初步发现显示星形细胞和神经细胞的NRXN1 在主要异构体表达、选择性剪接和硫酸乙酰肝素(HS)修饰方面存在根本差异。 星形胶质细胞NRXN1的选择性丢失导致AMPA受体(AMPAR)的显著和选择性减少。 在不影响大脑重要区域海马区突触数量的情况下介导突触传递 用于将信息从短期记忆巩固到长期记忆和空间记忆。何以 突触后膜，含有大量的Neuresin配体(如LRRTM、神经连接素等)， 区分突触前NRXN1和星形细胞NRXN1，以允许三方内的隔室特定信号 Synapse？为了更好地了解星形细胞NRXN1利用分子机制来指导 突触发育I将在NRXN1星形胶质细胞条件性基因敲除(ACKO)小鼠中进行救援实验 使用AAV递送NRXN1变体，这些变体在主要异构体同源性、选择性剪接、HS修饰和 细胞内信号。自发和诱发的兴奋性突触反应的录音将是 对CA1区锥体神经元进行抢救实验，观察抢救效果。接下来，我将扩展我们对 通过研究基础和/或是否需要星形胶质细胞NRXN1缺失来研究NRXN1缺失的功能后果 活动诱导的树突棘形态和密度的变化以及树突棘的三维密度 正常工作的AMPAR。最后，在星形细胞NRXN1缺失后观察到的突触缺陷可能是 星形胶质细胞功能受损的下游后果。因此，我将测量星形胶质细胞的几个方面 星形胶质细胞中NRXN1缺失后的功能，包括生理功能(即膜特性和通道 电流)、形态(即区域、缝隙连接耦合和与突触的关联)以及电路相关 (即钙动力学)特性。预计拟议的研究将为以下方面提供关键的见解 NRXN1 CNV如何引起神经发育障碍并将脱落的分子和细胞基础 一种支持神经元和星形胶质细胞间通讯的新分子程序 三方突触。

ABSTRACT Substantial evidence implicates copy number variations (CNVs) of the gene encoding Neurexin-1 (Nrxn1), a synaptic cell adhesion molecule, in the pathoetiology of Autism spectrum disorder, schizophrenia, and Tourette syndrome. However, very little is known regarding the function and regulation of Nrxn1 at developing and adult synapses, an understanding of which may improve our knowledge of brain disorders and reveal novel therapeutic targets. My preliminary data show that Nrxn1 is not only expressed by neurons, but also by astrocytes, challenging the classical view of Nrxn1 as being exclusively presynaptic. Here, I propose that astrocytic Nrxn1 mediates an important form of communication between astrocytes and synapses that is required for normal excitatory synapse development and function. My preliminary findings show that astrocytic and neuronal Nrxn1 fundamentally differ in major isoform expression, alternative splicing, and heparan sulfate (HS) modification. Selective loss of astrocytic Nrxn1 leads to a significant and selective reduction in AMPA receptor (AMPAR)- mediated synaptic transmission without affecting synapse number in the hippocampus, a brain region important for consolidation of information from short-term memory to long-term memory and spatial memory. How does the postsynaptic membrane, which contains numerous neurexin ligands (e.g. LRRTMs, neuroligins, etc.), distinguish between presynaptic and astrocytic Nrxn1 to allow compartment-specific signaling within the tripartite synapse? In order to better understand the molecular mechanisms utilized by astrocytic Nrxn1 to instruct synapse development I will perform rescue experiments in Nrxn1 astrocyte conditional knockout (acKO) mice using AAVs to deliver Nrxn1 variants differing in major isoform identity, alternative splicing, HS modification, and intracellular signaling. Recordings of both spontaneous and evoked excitatory synaptic responses will be performed on CA1 pyramidal neurons to measure rescue efficacy. Next, I will extend our understanding of the functional consequences of Nrxn1 deletion through studying whether astrocytic Nrxn1 is required for basal and/or activity-induced changes in dendritic spine morphology and density, as well as the three-dimensional density of functional AMPARs. Finally, the synaptic deficits observed following deletion of astrocytic Nrxn1 may be a downstream consequence of impaired astrocyte function. Thus, I will measure several aspects of astrocyte function following Nrxn1 deletion in astrocytes, including physiological (i.e. membrane properties and channel currents), morphological (i.e. territory, gap-junction coupling, and association with synapses), and circuit-related (i.e. calcium dynamics) properties. It is anticipated that the proposed research will provide critical insights into the molecular and cellular basis of how Nrxn1 CNVs give rise to neurodevelopmental disorders and will shed light on a novel molecular program underpinning communication between neuronal and astrocytic compartments of the tripartite synapse.