AMPAR Function in Synaptic and Extrasynaptic Membranes

AMPAR 在突触和突触外膜中的功能

基本信息

批准号：
10018120
负责人：
Jacques Wadiche
金额：
$ 42.31万
依托单位：
UNIVERSITY OF ALABAMA AT BIRMINGHAM
依托单位国家：
美国
项目类别：
财政年份：
2019
资助国家：
美国
起止时间：
2019-09-15 至 2024-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10018120
关键词：
AMPA Receptors Address Adult Affect Affinity Agonist Behavior Binding Biological Models Biophysics Brain Brain Diseases Calcium Cell membrane Cells Cerebellum Characteristics Data Dependence Development Disease Exhibits Fiber Frequencies Fright Genetic Transcription Glutamate Receptor Glutamates Goals Image Interneurons Knowledge Ligand Binding Literature Location Mediating Membrane Modeling Neuraxis Neurons Pathology Pattern Permeability Pharmacology Phenotype Physiology Polyamines Property Proteins Proxy Regulation Resolution Role Signal Transduction Site Slice Source Spermine Stress Structure Structure of molecular layer of cerebellar cortex Synapses Synaptic Transmission System Testing Toxin base biophysical properties channel blockers experience genetic manipulation indexing insight novel postsynaptic presynaptic prevent receptor receptor binding receptor function simulation trafficking two photon microscopy two-photon voltage

项目摘要

AMPA receptors (AMPARs) mediate the majority of excitatory glutamatergic synaptic transmission in the central nervous system. Most AMPARs, once bound to glutamate, allow Na+ and K+ flux across the cell membrane, causing neurons to depolarize. However, AMPARs that lack the GluR2 subunit are also permeable to Ca2+. These Ca2-permeable (CP) AMPARs are highly expressed during development when they are essential for activity-dependent plasticity, and this function persists at some synapses throughout adulthood. A biophysical characteristic known as rectification is commonly used to differentiate CP-AMPARs from Ca2+-impermeable (CI) AMPARs. Whereas CP-AMPARs exhibit strong inward rectification, CI-AMPA receptors display linear current-voltage relationships. Inward rectification of CP-AMPARs results from intracellular polyamines that act as open channel blockers to prevent outward current flux. Thus, inward rectification and sensitivity to antagonists that bind at the polyamine site provide biophysical signatures of AMPAR subunit composition and hence Ca2+ permeability, and these characteristics have been widely used to establish rules of AMPAR subunit plasticity. Molecular layer interneurons of the cerebellum provide a well-established model system for understanding AMPAR localization and trafficking because repetitive synaptic stimulation or a single experience of fear triggers a form of plasticity called subunit-switching wherein CP-AMPARs at synapses are replaced by CI-AMPARs from a pool of extrasynaptic AMPARs. Although rectification index and sensitivity to polyamine site toxins are widely used to distinguish between GluR2-containing and -lacking AMPARs, there are many examples from the literature that show these biophysical properties do not exclusively reflect subunit composition. A separate literature has converged on gating models of AMPARs that include multiple conductance states, but the functional implications are unclear. Now, our preliminary data show that CP-AMPAR rectification and pharmacology are sensitive to factors that regulate AMPAR conductance states, potentially complicating the interpretation of results using these biophysical properties as sole proxies of subunit composition. We propose to understand how the multiple sub-conductance states of AMPARs contribute to the hallmark biophysical properties CP-AMPARs. We will use high resolution Ca2+ imaging, heterologous expression systems and genetic manipulation to understand regulation of CP-AMPAR biophysical properties and use that understanding to critically evaluate CP-AMPAR localization and plasticity in cerebellar molecular layer interneurons. AMPAR subunit composition has important functional consequences, ranging from regulating the ability of postsynaptic cells to precisely follow high-frequency synaptic activity and mediating Ca2+ influx that can trigger plasticity or pathology. Successful completion of the proposed studies will reveal novel properties of AMPARs that are essential for understanding their function within synapses and intact circuits in the normal and diseased brain.

AMPA 受体 (AMPAR) 介导大脑中大部分兴奋性谷氨酸能突触传递中枢神经系统。大多数 AMPAR 一旦与谷氨酸结合，就会允许 Na+ 和 K+ 穿过细胞膜，导致神经元去极化。然而，缺乏 GluR2 亚基的 AMPAR 也是可渗透的至 Ca2+。这些 Ca2 渗透性 (CP) AMPAR 在发育过程中高度表达，当它们对于活动依赖性可塑性至关重要，并且这种功能在整个成年期的某些突触中持续存在。一个称为整流的生物物理特征通常用于区分 CP-AMPAR 与 Ca2+ 不可渗透 (CI) AMPAR。 CP-AMPAR 表现出强烈的内向整流作用，而 CI-AMPA 受体则表现出强烈的内向整流作用。显示线性电流-电压关系。 CP-AMPAR 的内向整流来自细胞内多胺作为开放通道阻断剂以防止向外的电流通量。因此，向内整顿和对结合在多胺位点的拮抗剂的敏感性提供了 AMPAR 亚基的生物物理特征组成以及 Ca2+ 渗透性，这些特性已被广泛用于建立规则 AMPAR 亚基可塑性。小脑的分子层中间神经元提供了一个完善的模型用于理解 AMPAR 定位和运输的系统，因为重复的突触刺激或单个恐惧体验会触发一种称为亚基转换的可塑性，其中突触处的 CP-AMPAR 被突触外 AMPAR 池中的 CI-AMPAR 取代。尽管整改指数和敏感度多胺位点毒素广泛用于区分含有 GluR2 和缺乏 GluR2 的 AMPAR，文献中的许多例子表明这些生物物理特性并不完全反映亚基作品。另一篇文献集中讨论了 AMPAR 的门控模型，其中包括多个电导状态，但功能含义尚不清楚。现在，我们的初步数据表明 CP-AMPAR 整流和药理学对调节 AMPAR 电导状态的因素敏感，使用这些生物物理特性作为唯一指标可能会使结果的解释变得复杂亚基组成。我们建议了解 AMPAR 的多个亚电导状态如何有助于 CP-AMPAR 的标志性生物物理特性。我们将使用高分辨率 Ca2+ 成像，异源表达系统和基因操作以了解 CP-AMPAR 的调控生物物理特性，并利用这种理解来批判性地评估 CP-AMPAR 的定位和可塑性小脑分子层中间神经元。 AMPAR 亚基组成具有重要的功能后果，范围从调节突触后细胞的能力以精确跟踪高频突触活动，介导可引发可塑性或病理学的 Ca2+ 流入。成功完成拟议的研究将揭示 AMPAR 的新特性，这对于理解它们在突触内的功能至关重要正常和患病大脑中的完整电路。