The role of gephyrin translation in inhibitory synaptic plasticity

gephyrin 翻译在抑制性突触可塑性中的作用

• 批准号: 10629239
• 负责人: Theresa Marie Welle
• 金额: $ 3.72万
• 依托单位: UNIVERSITY OF COLORADO DENVER
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-04-01 至 2025-08-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10629239
• 关键词: Affect; Automobile Driving; Biological Assay; Chemosensitization; Cognition; Data; Dendrites; Electrophysiology (science); Epilepsy; Equilibrium; Excitatory Synapse; Functional disorder; Genes; Glutamates; Goals; Image; In Situ; Inhibitory Synapse; Knowledge; Label; Learning; Ligation; Long-Term Depression; Long-Term Potentiation; Maintenance; Measures; Mediating; Memory; Messenger RNA; MicroRNAs; Molecular; N-Methyl-D-Aspartate Receptors; Neurons; Neurophysiology - biologic function; Pathology; Play; Protein Biosynthesis; Proteins; Proteome; Puromycin; Regulation; Research; Role; Scaffolding Protein; Schizophrenia; Shapes; Site; Synapses; Synaptic Transmission; Synaptic plasticity; Testing; Transcript; Translating; Translational Regulation; Translations; Untranslated RNA; Visualization; autism spectrum disorder; confocal imaging; experimental study; gephyrin; immunocytochemistry; insight; mRNA Translation; neural; neural circuit; neuronal cell body; neuronal excitability; overexpression; postsynaptic; receptor; scaffold; single molecule; synaptic function; synaptic inhibition; translational impact

PROJECT SUMMARY Synaptic plasticity, the activity-dependent alteration in the strength of neuronal connections, forms the molecular basis of learning, memory, and cognition. Both excitatory and inhibitory connections undergo bidirectional synaptic plasticity to tune neuronal excitability, sculpt neural circuits, and coordinate the balance of excitation and inhibition (E/I balance). GABAergic synaptic plasticity is crucial for maintaining E/I balance, and its dysfunction is implicated in pathologies such as epilepsy, schizophrenia, and autism spectrum disorders. However, inhibitory synaptic plasticity and its molecular underpinnings are critically understudied. A crucial regulator of inhibitory synaptic plasticity is gephyrin, a synaptic scaffolding protein essential for GABAA receptor clustering and inhibitory synaptic transmission. In one form of inhibitory synaptic plasticity, inhibitory long-term potentiation (iLTP), gephyrin clustering increases rapidly at dendritic inhibitory synapses to strengthen GABAergic synapses exclusively in dendrites by 20 min post-stimulation. Early mechanisms of iLTP are independent of translation, but much less is known about iLTP persistence and how translation of key proteins maintains iLTP. Answering these questions will provide insights into mechanisms of synaptic plasticity and how changes in GABAergic synaptic strength shape neural circuits long-term. In this proposal, I will address this knowledge gap by focusing on the role of gephyrin translation for maintaining iLTP and more specifically where it occurs and how it is regulated to enact precise and persistent potentiation of inhibitory synaptic connections. I hypothesize that local gephyrin translation supports iLTP exclusively in the dendrites and that translational regulation of gephyrin influences gephyrin synaptic clustering and inhibitory synaptic strength. I will utilize fluorescent in situ labeling of mRNA and visualization of actively translating proteins to determine localization and translation of gephyrin transcripts in dendrites following iLTP (Aim 1). miR153 is a short, non-coding transcript identified as a key regulator of gephyrin translation and implicated in learning, memory, and cognition. By modulating its expression levels in neurons and employing a combination of immunocytochemistry and electrophysiology, I will determine the impact of miR153 function on gephyrin clustering and inhibitory synaptic strength during iLTP (Aim 2). Together, this approach will elucidate how the localization and regulation of gephyrin translation impact specific and lasting changes to inhibitory synaptic strength during iLTP and ultimately reveal the mechanisms driving inhibitory synaptic plasticity to maintain E/I balance for proper neural function.

项目摘要 突触可塑性是神经元连接强度的活性依赖性改变，形成了分子 学习，记忆和认知的基础。兴奋性和抑制性连接都会发生双向 突触可塑性调整神经元兴奋性，雕刻神经回路并协调兴奋平衡 和抑制（E/I平衡）。 GABA能突触可塑性对于维持E/I平衡至关重要，它 功能障碍与癫痫，精神分裂症和自闭症谱系障碍等病理有关。 然而，抑制性突触可塑性及其分子基础被严格研究。关键 抑制性突触可塑性的调节剂是Gephyrin，Gephyrin是GABAA受体必不可少的突触脚手蛋白 聚类和抑制性突触传播。以一种抑制性突触可塑性形式的长期抑制作用 增强（ILTP），gephyrin聚类在树突抑制突触下迅速增加 GABA能突触专门在树突中刺激20分钟。 ILTP的早期机制是 独立于翻译，但对ILTP的持久性以及关键蛋白的翻译如何了解不多 维护ILTP。回答这些问题将提供有关突触可塑性机制的见解以及如何 GABA能突触强度的变化长期神经回路。在此提案中，我将解决这个问题 通过关注Gephyrin翻译在维持ILTP的作用，更具体地说的知识差距 它发生的位置以及如何调节抑制性突触的精确和持续增强 连接。我假设局部gephyrin翻译在树突中仅支持ILTP，并且 Gephyrin的翻译调节会影响Gephyrin突触聚类和抑制性突触 力量。我将利用mRNA的荧光原位标记，并可视化主动将蛋白质转化为 确定ILTP之后的树突中Gephyrin转录本的定位和翻译（AIM 1）。 mir153是 简短的非编码成绩单被确定为gephyrin翻译的关键调节剂，并与学习有关， 记忆和认知。通过调节其在神经元中的表达水平并采用 免疫细胞化学和电生理学，我将确定miR153功能对gephyrin的影响 ILTP期间的聚类和抑制性突触强度（AIM 2）。这种方法一起将阐明 Gephyrin Translation的定位和调节会影响特异性和持久变化对抑制性突触 ILTP期间的强度，并最终揭示了驱动抑制性突触可塑性以维持E/I的机制 平衡适当的神经功能。