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

分子逻辑塑造 Neurexin-1 对三联突触的细胞特异性贡献

基本信息

批准号：
10594568
负责人：
Justin Howard Trotter
金额：
$ 15.17万
依托单位：
STANFORD UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2021
资助国家：
美国
起止时间：
2021-04-01 至 2024-03-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10594568
关键词：
3-Dimensional AMPA Receptors Address Adult Affect Alternative Splicing Astrocytes Behavioral Binding Proteins Biological Assay Brain Brain Diseases Brain region Calcium Cell Adhesion Molecules Cells Communication Communications Media Copy Number Polymorphism Coupled Coupling Data Dendritic Spines Development Disease Dyes Electrophysiology (science)Ensure Etiology Excitatory Synapse Fluorescence Resonance Energy Transfer Gap Junctions Genes Genetic Gilles de la Tourette syndrome Glutamate Transporter Glutamates Goals Grant Heparitin Sulfate Hippocampus Impairment Knockout Mice Knowledge Ligand Binding Ligands Link Logic Measures Mediating Membrane Mentors Messenger RNA Modification Molecular Morphology Mus Neurodevelopmental Disorder Neuroglia Neurons Physiological Postsynaptic Membrane Potassium Channel Potassium Glutamate Prevalence Probability Property Protein Isoforms Proteins Regulation Research Role Schizophrenia Short-Term Memory Signal Transduction Synapses Synaptic Transmission Testing Training Variant academic preparation autism spectrum disorder biocytin career conditional knockout density design differential expression experimental study hippocampal pyramidal neuron improved insight long term memory mutant new therapeutic target novel novel therapeutics patch clamp postsynaptic postsynaptic neurons preference presynaptic programs receptor function response spatial memory synaptic function transmission process two photon microscopy

项目摘要

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.

抽象的大量证据表明编码 Neurexin-1 (Nrxn1) 的基因存在拷贝数变异 (CNV)，Neurexin-1 是一种突触细胞粘附分子，在自闭症谱系障碍、精神分裂症和抽动秽语症的病理学中综合症。然而，关于 Nrxn1 在发育和成人中的功能和调节知之甚少。突触，对突触的理解可能会提高我们对大脑疾病的认识并揭示新的治疗方法目标。我的初步数据表明，Nrxn1不仅由神经元表达，还由星形胶质细胞表达，挑战 Nrxn1 完全是突触前的经典观点。在这里，我建议星形胶质细胞Nrxn1 介导星形胶质细胞和突触之间的正常通信所需的重要形式兴奋性突触的发育和功能。我的初步发现表明星形胶质细胞和神经元 Nrxn1 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 的选择性丢失会导致 AMPA 受体 (AMPAR) 显着且选择性减少 - 介导突触传递而不影响海马体的突触数量，海马体是大脑的重要区域用于巩固从短期记忆到长期记忆和空间记忆的信息。怎么样突触后膜，含有大量神经毒素配体（例如 LRRTM、神经毒素等），区分突触前 Nrxn1 和星形胶质细胞 Nrxn1，以允许三方内的隔室特异性信号传导突触？为了更好地理解星形胶质细胞Nrxn1利用的分子机制来指导突触发育我将在 Nrxn1 星形胶质细胞条件敲除 (acKO) 小鼠中进行救援实验使用 AAV 传递在主要亚型身份、选择性剪接、HS 修饰等方面不同的 Nrxn1 变体细胞内信号传导。自发和诱发的兴奋性突触反应的记录将对 CA1 锥体神经元进行测试以测量救援效果。接下来我就来扩展一下我们的理解通过研究星形胶质细胞 Nrxn1 是否需要基础和/或 Nrxn1 缺失的功能后果活动引起的树突棘形态和密度以及三维密度的变化功能性 AMPAR。最后，删除星形胶质细胞 Nrxn1 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，包括生理学（即膜特性和通道电流）、形态学（即领土、间隙连接耦合以及与突触的关联）和电路相关（即钙动力学）特性。预计拟议的研究将提供重要的见解 Nrxn1 CNV 如何引起神经发育障碍并脱落的分子和细胞基础揭示支持神经元和星形细胞区室之间通讯的新型分子程序三方突触。