Excitability and Excitotoxicity in Type-I Cochlear Afferents: Synapse Structure and Function

I 型耳蜗传入神经的兴奋性和兴奋性毒性：突触结构和功能

基本信息

批准号：
10444754
负责人：
Mark Allen Rutherford
金额：
$ 69.54万
依托单位：
WASHINGTON UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2016
资助国家：
美国
起止时间：
2016-03-10 至 2027-02-28
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10444754
关键词：
Acids Acoustic Nerve Acute Address Adhesions Affect Anatomy Antioxidants Basic Science Cells Chronic Cochlea Collaborations Combined Modality Therapy Complement Complex Data Discipline Dose Drug Delivery Systems Ear Electrophysiology (science)Excitatory Amino Acid Antagonists FDA approved Family Functional disorder Funding Genetic Glutamate Receptor Glutamates Goals Hearing Hearing problem Heterogeneity Histology International Intervention Investigation Knockout Mice Knowledge Labyrinth Measures Mediating Molecular Morphology Mus Nerve Fibers Nervous system structure Neurotransmitter Receptor Noise Permeability Pharmaceutical Preparations Pharmacology Physiological Physiology Polyamines Prevention Property Proteins Proteomics Recombinants Resistance Role Structure Synapses Synaptic Transmission Testing Time antagonist channel blockers cochlear development cochlear synaptopathy design excitotoxicity in vivo local drug delivery mouse genetics neuroligin 1 neuroligin 3 neurophysiology noise exposure noise trauma patch clamp postsynaptic prevent prevent hearing loss receptor response side effect small molecule sound stargazin tool translational potential transmission process

项目摘要

Project Summary: This project investigates the cochlear AMPA-type glutamate receptors (AMPARs) that are necessary for hearing, overactivation of which leads to excitotoxic synapse loss and hearing disorders. Each cochlear afferent synapse expresses many hundreds to a few thousand of these AMPARs, of both the Ca2+-permeable subtype (CP-AMPARs, lacking subunit GluA2) and the Ca2+-impermeable subtype (CI-AMPARs, containing subunit GluA2). The combination of pore-forming GluA subunits and auxiliary subunits of the AMPAR complex, influenced by transsynaptic adhesion factors, determine its physiological properties and pharmacological sensitivities. The cochlear AMPAR complex has properties that make it unique in the nervous system, for example, the absence of GluA1. However, the precise complement of cochlear AMPAR subunits in not known. This proposal uses mouse genetics, in vivo and ex vivo cochlear electrophysiology, proteomics, and ultrastructural molecular anatomy to investigate the subunit composition, pharmacological sensitivity, and functional significance of the cochlear AMPAR complex. We will determine the influence of auxiliary subunit TARP-2 (Stargazin) on cochlear function, synaptic transmission, and GluA subunit expression. We will test the hypothesis that synaptopathy in GluA3KO mice results from an increase in Ca2+-permeability of the AMPAR complex. We will determine how Neuroligin1 and 3 affect AMPAR subunit expression and auditory nerve fiber physiology. We will determine the influence of GluA3, TARP-2, Nlgn1, and Nlgn3 on the intrasynaptic distribution of AMPAR subunits. With recombinant expression of different combinations of GluA pore-forming and auxiliary subunits in HEK cells (with or without GluA3, with or without TARP-2), we will challenge our understanding of the cochlear AMPAR complex by comparing changes in pharmacological sensitivity with those changes observed for the native cochlear synapses (GluA3WT vs GluA3KO, TARP-2WT vs TARP-2KO). The gain of this basic knowledge will inform design of small molecules to target cochlear AMPARs. With chronic systemic administration of the tool compound (CP-AMPAR blocker IEM-1925), we will measure synaptic adaptation and resistance to noise-induced synaptopathy. With acute systemic dosing, we will ask if noise trauma can be prevented if IEM-1925 is given only during, not before, the noise exposure and if IEM- 1925 + antioxidant combination therapy can protect cochlear function from more intense noise exposures. The long-term goal of this line of investigation is to develop systemic drugs to target CP-AMPARs of the inner ear while allowing hearing function to be maintained through CI-AMPARs, and while avoiding unwanted CNS side effects. The successful completion of this collaborative project will determine the precise subunit composition of the cochlear AMPAR complex and its influence on pharmacological sensitivity.

项目概要：该项目研究了耳蜗 AMPA 型谷氨酸受体 (AMPAR)，该受体是听力，过度激活会导致兴奋性毒性突触丧失和听力障碍。传入突触表达数百到数千个 AMPAR，其中包括 Ca2+ 渗透性突触亚型（CP-AMPAR，缺乏 GluA2 亚基）和 Ca2+ 不可渗透亚型（CI-AMPAR，含有 AMPAR 复合物的成孔 GluA 亚基和辅助亚基的组合，受突触粘附因子的影响，决定其生理特性和药理耳蜗 AMPAR 复合体具有使其在神经系统中独一无二的特性。例如，GluA1 的缺失。然而，耳蜗 AMPAR 亚基的精确补充尚不清楚。该提案使用小鼠遗传学、体内和离体耳蜗电生理学、蛋白质组学和超微结构分子解剖学研究亚基组成、药理学敏感性和耳蜗 AMPAR 复合体的功能意义我们将确定辅助亚基的影响。 TARP-2 (Stargazin) 对耳蜗功能、突触传递和 GluA 亚基表达的影响我们将进行测试。 GluA3KO 小鼠的突触病是由于 AMPAR 的 Ca2+ 通透性增加所致的假设我们将确定 Neuroligin1 和 3 如何影响 AMPAR 亚基表达和听觉神经纤维。我们将确定 GluA3、TARP-α2、Nlgn1 和 Nlgn3 对突触内的影响。 AMPAR 亚基的分布与 GluA 成孔的不同组合的重组表达。和 HEK 细胞中的辅助亚基（有或没有 GluA3，有或没有 TARP-2），我们将挑战我们的通过比较药理敏感性的变化来了解耳蜗 AMPAR 复合体观察到的原生耳蜗突触的变化（GluA3WT 与 GluA3KO、TARP-2WT 与 TARP-2KO）。获得这些基础知识将为针对耳蜗 AMPAR 的小分子设计提供信息。工具化合物（CP-AMPAR阻滞剂IEM-1925）的长期全身给药，我们将测量对于急性全身给药，我们会问突触适应和对噪声引起的突触病的抵抗力。如果仅在噪声暴露期间而不是之前给予 IEM-1925，并且如果 IEM- 1925+抗氧化剂联合疗法可以保护耳蜗功能免受更强烈的噪音暴露。该研究方向的长期目标是开发针对内耳 CP-AMPAR 的全身药物同时允许通过 CI-AMPAR 维持听力功能，同时避免不需要的 CNS 侧该合作项目的成功完成将决定精确的亚基组成。耳蜗 AMPAR 复合体的作用及其对药理敏感性的影响。