Molecular mechanisms of cochlear hair bundle mechanics

项目摘要 耳蜗放大是我们的听觉系统放大和调谐对 传入的声音，赋予我们出色的音量灵敏度、大的动态范围和精细的 频率歧视。据推测，听觉感觉细胞有两个过程对耳蜗起作用。 放大：发生在细胞体中的体细胞运动和发生在细胞胞体中的活跃毛束机制。 顶端立体纤毛发束。探讨主动发束力学对耳蜗术前放大的作用 需要进一步了解与之相关的流程。毛细胞机械转导(MET)，过程 将声音刺激转化为发束中的电信号，是活跃的发束力学的驱动因素。 MET适应是一种关键的机制，被认为有助于积极的毛束机制。 以前在非哺乳动物模型中的工作表明，适应分为快过程和慢过程，两者都是 其中依赖于钙的流入来驱动这一过程。哺乳动物耳蜗中的数据表明，适应 也包括快和慢的成分，但我们的工作表明，驱动快和慢的潜在生物 耳蜗中缓慢的过程与以前报道的非耳聋的过程有根本的不同。 哺乳动物的毛细胞。因此，需要新的研究来了解相关的分子机制。 快适应和慢适应，以及它们对哺乳动物听觉处理的贡献。 从有关耳蜗肌的特性的最新数据中，我们假设张力对适应是必不可少的。 机械装置。在这项研究的目标1中，我们将调查肌球蛋白马达对适应和毛发的贡献。 捆绑机械师。我们使用新的、更快的刺激和高速成像来监测机械 发束的变化与毛细胞电生理学和药物操作有关。使用 许多已知的肌球蛋白马达对听觉功能很重要，在目标2中我们将探索其贡献。 利用现有的小鼠模型，将特定的肌球蛋白马达应用于适应和毛束机制。对于目标3，我们 利用CRISPR/Cas9技术开发了一种新的小鼠模型，可以迅速灭活肌球蛋白VIIa运动功能， 我们将评估肌球蛋白VIIa在紧张产生中的作用。 这一方案中的实验将进一步加深我们对哺乳动物分子机制的理解 利用耳蜗机能和发束机制建立新的哺乳动物听觉MET模型 进程。我们处于独特的地位，可以通过我们拥有并将继续使用的新技术来实现这一目标 发展。听觉MET的基本机制知识将引导我们进行实验，在那里我们可以询问 系统在体内，以确定特定的分子贡献的耳蜗扩增。了解人工耳蜗术 放大可以更好地预防和/或恢复听力。

Project Summary Cochlear amplification is the process by which our auditory system amplifies and tunes responses to incoming sounds, bestowing us with our excellent sound level sensitivity, large dynamic range, and fine frequency discrimination. Auditory sensory cells have two processes hypothesized to contribute to cochlear amplification: somatic motility that occurs in the cell soma and active hair bundle mechanics that occurs in the apical stereocilia hair bundle. To assay the contribution of active hair bundle mechanics to cochlear amplification requires further understanding of the processes related to it. Hair cell mechanotransduction (MET), the process of converting sound stimuli into electrical signals in the hair bundle, is the driver of active hair bundle mechanics. MET adaptation is one key mechanism that is hypothesized to contribute to active hair bundle mechanics. Previous work in non-mammalian models show that adaptation is separated into fast and slow processes, both of which rely on the influx of calcium to drive the process. Data in the mammalian cochlea indicate that adaptation also consists of fast and slow components, but our work shows that the underlying biology driving the fast and slow processes in the cochlea is fundamentally different from what has been previously reported in non- mammalian hair cells. Thus, new investigations are needed to understand the molecular machinery responsible for both fast and slow adaptation, and their contributions to mammalian auditory processing. From new data about properties of cochlear MET, we hypothesize that tension is essential for adaptation mechanisms. In Aim 1 of this study, we will investigate the contribution of myosin motors to adaptation and hair bundle mechanics. We assay this using new, faster stimulation and high-speed imaging to monitor mechanical changes in the hair bundle coupled with hair cell electrophysiology and pharmacological manipulation. With numerous myosin motors known to be important for auditory function, in Aim 2 we will explore the contributions of specific myosin motors to adaptation and hair bundle mechanics using existing mouse models. For Aim 3, we developed a new mouse model using CRISPR/Cas9 technology to acutely inactivate myosin VIIa motor function, and we will assess the role of myosin VIIa in tension generation. The experiments in this proposal will further our understanding of the molecular mechanisms of mammalian cochlear adaptation and hair bundle mechanics to develop a new model of the mammalian auditory MET process. We are uniquely positioned to accomplish this with the new technologies that we have and continue to develop. Basic mechanistic knowledge of auditory MET will lead to experiments where we can interrogate the system in vivo to determine specific molecular contributions to cochlear amplification. Understanding cochlear amplification can lead to better prevention and/or restoration of hearing.