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）：这是研究计划的研究计划的竞争续约，该计划研究了对耳蜗细胞的胆碱能抑制的生物物理和分子基础。神经递质乙酰胆碱（ACH）开放由19和110个亚基组成的离子（阳离子 - permeant）受体（ACHR）。通过ACHR钙的流入钙涌入钙敏感的钾通道（由SK2基因编码）以超极化并抑制毛细胞。虽然通过ACHR钙涌入似乎足以激活SK通道，但其他观察结果表明，钙可以从内质存储中释放出来，以扩展和放大该过程。传出突触的结构包括毛细胞中的近膜水箱，可以用作钙储存。毛细胞ACHR和相关SK通道的遗传改变为传出功能提供了一些见解。特别是，单个氨基酸在19的渗透路径中的改变产生了一个“超级受体”，其开放时间延长，对ACH的敏感性更高，脱敏速度放缓。在具有该受体的敲蛋白小鼠中，传出的抑制作用得到了极大的增强，并且可以更好地保护动物免受永久性的声音损害。在该模型和其他转基因模型中，我们将继续探测胆碱能抑制的分子机制。此外，我们将利用禽（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。如果要进行进展，我们将构建一种用鸡19取代的转基因小鼠。我们将对缺少一个或多个effer体突触的一个或多个成分的野生型和转基因小鼠中的突触箱进行详细的超微结构分析。水箱和突触后膜之间的细胞质间隙很小，充满了有组织的电子密度，这表明存在可能包括ACHRS和钙释放通道的细胞质部分的分子“连接器”。这种布置强烈让人联想到肌肉中的连接络合物，并表明除了基于钙的信号传导外，ACHR和钙释放通道之间的“构象耦合”的可能性。内耳的传出神经支配会调节灵敏度并预防声学创伤。我们对这一过程的分子碱基的知识日益开始，已经揭示了潜在的治疗靶标，并得到了毛细胞乙酰胆碱受体的独特药理学的支持。透彻了解胆碱能抑制的分子碱基将推进利用这些机制的治疗策略。