Cholinergic Response of Cochlear Hair Cells

耳蜗毛细胞的胆碱能反应

• 批准号: 8304285
• 负责人: Paul A Fuchs
• 金额: $ 52.07万
• 依托单位: JOHNS HOPKINS UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1992
• 资助国家: 美国
• 起止时间: 1992-09-30 至 2014-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8304285
• 关键词: 3-Dimensional; Acetylcholine; Acoustic Trauma; Amino Acids; Animals; Birds; Buffers; Calcium; Calcium Channel; Cations; Cell membrane; Chickens; Cholinergic Receptors; Cochlea; Comparative Study; Complex; Coupling; Dependence; Dimensions; Electron Microscopy; Electrons; Genes; Genetic; Hair Cells; Image; Knock-out; Knockout Mice; Knowledge; Labyrinth; Long-Term Effects; Mammals; Membrane; Microscopic; Molecular; Molecular Probes; Mus; Muscle; Mutation; Neurotransmitters; Outcome; Outer Hair Cells; Permeability; Pharmacology; Point Mutation; Postsynaptic Membrane; Potassium Channel; Process; Property; Protocols documentation; Rattus; Research; Resolution; Role; Signal Transduction; Structure; Synapses; Synaptic Membranes; Therapeutic; Time; Transgenic Mice; Transgenic Model; Xenopus oocyte; base; cholinergic; comparative; desensitization; driving force; electron density; insight; mouse model; nerve supply; programs; receptor; release of sequestered calcium ion into cytoplasm; research study; response; sound; therapeutic target; tomography; voltage

DESCRIPTION (provided by applicant): This is the competing renewal of a research program that examines the biophysical and molecular basis of cholinergic inhibition of cochlear hair cells. The neurotransmitter acetylcholine (ACh) opens ionotropic (cation-permeant) receptors (AChRs) composed of 19 and 110 subunits. Calcium influx through the AChR activates calcium-sensitive potassium channels (encoded by the SK2 gene) to hyperpolarize and inhibit the hair cell. While calcium influx through the AChR seems adequate to activate the SK channels, other observations suggest that calcium may be released from endoplasmic stores to extend and amplify that process. The structure of the efferent synapse includes a near-membrane cistern in the hair cell that may serve as a calcium store. Genetic alteration of the hair cell's AChR and associated SK channels has provided some insight into efferent function. In particular, alteration of a single amino acid in the permeation path of 19 produced a 'super receptor' with prolonged open times, greater sensitivity to ACh and slowed desensitization. In the knockin mouse possessing this receptor, efferent inhibition was greatly enhanced, and the animal was better protected from permanent loud sound damage. We will continue to probe the molecular mechanisms of cholinergic inhibition in this and other transgenic models. In addition, we will exploit functional and genetic differences between avian (chicken) and mammalian AChRs to explore the role of calcium flux in detail. Should progress warrant it, we will construct a transgenic mouse substituted with chicken 19. We will perform detailed ultrastructural analyses of synaptic cisterns in wildtype and transgenic mice lacking one or more components of the efferent synapse. The small cytoplasmic gap between cistern and postsynaptic membrane is replete with organized electron-densities, suggesting the presence of molecular 'connectors' that may include the cytoplasmic portions of AChRs and calcium release channels. This arrangement is strongly reminiscent of junctional complexes in muscle and suggests the possibility of 'conformational coupling' between AChRs and calcium release channels, in addition to calcium-based signaling. Efferent innervation of the inner ear modulates sensitivity and protects against acoustic trauma. Our growing knowledge of the molecular bases for this process has begun to reveal potential therapeutic targets, supported by the unique pharmacology of hair cell acetylcholine receptors. Thorough knowledge of the molecular bases of cholinergic inhibition will advance therapeutic strategies exploiting those mechanisms.

描述（由申请人提供）：这是一项研究计划的竞争更新，该计划检查耳蜗毛细胞胆碱能抑制的生物物理和分子基础。神经递质乙酰胆碱 (ACh) 可打开由 19 和 110 个亚基组成的离子型（阳离子渗透）受体 (AChR)。通过 AChR 的钙流入会激活钙敏感钾通道（由 SK2 基因编码），从而超极化并抑制毛细胞。虽然通过 AChR 的钙流入似乎足以激活 SK 通道，但其他观察结果表明，钙可能会从内质储存中释放，以延长和放大该过程。传出突触的结构包括毛细胞中的近膜池，可充当钙储存库。毛细胞 AChR 和相关 SK 通道的遗传改变为了解传出功能提供了一些见解。特别是，19 渗透路径中单个氨基酸的改变产生了“超级受体”，其开放时间延长，对乙酰胆碱更加敏感，并减慢了脱敏速度。在拥有这种受体的敲入小鼠中，传出抑制大大增强，并且更好地保护动物免受永久性大声损伤。我们将继续探讨该转基因模型和其他转基因模型中胆碱能抑制的分子机制。此外，我们将利用鸟类（鸡）和哺乳动物 AChR 之间的功能和遗传差异来详细探讨钙通量的作用。如果进展有保证，我们将构建一种用鸡 19 替代的转基因小鼠。我们将对缺乏传出突触的一种或多种成分的野生型和转基因小鼠的突触池进行详细的超微结构分析。池和突触后膜之间的小细胞质间隙充满了有组织的电子密度，表明分子“连接器”的存在，其中可能包括 AChR 的细胞质部分和钙释放通道。这种排列强烈地让人想起肌肉中的连接复合物，并表明除了基于钙的信号传导之外，AChR 和钙释放通道之间还存在“构象耦合”的可能性。内耳的传出神经支配调节敏感性并防止声损伤。我们对该过程的分子基础的了解不断加深，已开始揭示潜在的治疗靶点，并得到毛细胞乙酰胆碱受体独特药理学的支持。对胆碱能抑制的分子基础的透彻了解将推进利用这些机制的治疗策略。