Experimental study of the cochlear amplifier

描述(申请人提供)：哺乳动物耳蜗通过一种名为“耳蜗放大器”的有源放大机制实现其非凡的灵敏度、精致的频率选择性和巨大的动态范围，该机制利用新陈代谢能量促进基底膜(BM)振动。虽然外毛细胞可以通过体细胞和发束运动产生作用力，但其机械机制仍不清楚。这项应用是通过使用我们最新研制的扫描低相干外差干涉仪在正常和转基因耳蜗中进行一系列新颖的实验来研究体细胞和发束运动在耳蜗声放大中的作用。具体目的一是测量敏感小鼠和沙土鼠耳蜗骨质膜、网状板(RL)和覆盖膜(TM)的振动。假说是，在敏感的小鼠耳蜗中，基于OHC的耳蜗放大器在RL产生的振动比在BM产生的振动更大、更非线性。小鼠的BM和RL振动高度敏感，表现出尖锐的调谐和非线性，就像其他常用的耳蜗力学研究动物，如沙土鼠。这项实验的数据将是研究小鼠和沙土鼠耳蜗微力学的必要数据。具体目标二将通过测量没有躯体运动的小鼠耳蜗体运动的BM、RL和TM振动来确定OHC躯体运动的作用。假设是，在Prestin 499敲门小鼠中，缺乏躯体运动导致BM、RL和TM振动的敏感度降低、尖锐调谐和非线性丢失。由于Prestin 499敲击小鼠的力电传导、僵硬和OHC的形态是正常的，预期的变化将归因于缺乏躯体运动。具体目标三将通过测量毛束运动无效的小鼠耳蜗的BM、RL和TM振动来确定OHC毛束运动的作用。假说是，带有独立发束的缩短和分离的TM损害了TectaC1509G/C1509G小鼠的毛束运动、机械电转导和耳蜗放大，降低了声音诱发的BM、RL和TM振动的敏感度、敏感度和非线性。然而，电刺激同时激活躯体运动和发束运动，并导致BM、RL和TM振动和电诱发耳声发射。在用水杨酸盐抑制体细胞运动后，可以在野生型小鼠中测量到毛束运动介导的反应，但在TectaC1509G/C1509G小鼠中则不能。通过测量和比较TectaC1509G/C1509G和野生型小鼠应用水杨酸盐后的电诱发反应，确定发束运动对耳蜗声放大的贡献。这项研究的新数据将提供有关OHC躯体和发束运动在耳蜗放大中的重要作用以及理解人类听力障碍机制的关键信息。

DESCRIPTION (provided by applicant): The mammalian cochlea achieves its remarkable sensitivity, exquisite frequency selectivity, and enormous dynamic range through an active amplification mechanism called the "cochlear amplifier", which uses metabolic energy to boost basilar membrane (BM) vibration. Although it is well established that outer hair cells (OHCs) can generate forces through somatic and hair-bundle motility, the mechanical mechanism of cochlear amplification remains unclear. This application is to study the role of somatic and hair-bundle motility in cochlear amplification by conducting a set of novel experiments in normal and genetically modified cochleae using our recently developed scanning low-coherence heterodyne interferometer. Specific aim one will measure the BM, reticular lamina (RL), and tectorial membrane (TM) vibration in sensitive mouse and gerbil cochleae. The hypothesis is that, in sensitive mouse cochleae, the OHC-based cochlear amplifier produces a larger and more nonlinear vibration at the RL than that at the BM. BM and RL vibrations in mice are highly sensitive, and show sharp tuning and nonlinearity as in other commonly used animals for cochlear mechanics study, such as gerbils. The data from this experiment will be essential for studying cochlear micromechanics in mice and gerbils. Specific aim two will determine the role of the OHC somatic motility by measuring the BM, RL, and TM vibration in mouse cochleae without somatic motility. The hypothesis is that, in prestin 499 knockin mice, the lack of somatic motility results in decreased sensitivity and loss of sharp tuning and nonlinearity of the BM, RL, and TM vibration. Because the mechanoelectrical transduction, stiffness, and morphology of OHCs in prestin 499 knockin mice are normal, the expected changes will be attributed to the absence of somatic motility. Specific aim three will determine the role of the OHC hair-bundle motility by measuring the BM, RL, and TM vibration in mouse cochleae with ineffective hair-bundle motility. The hypothesis is that shortened and detached TM with free-standing hair bundles compromises hair-bundle motility, mechanoelectrical transduction, and cochlear amplification in TectaC1509G/C1509G mice, decreasing sensitivity, sharp tuning, and nonlinearity of acoustically evoked BM, RL, and TM vibration. However, electrical stimulation activates both somatic and hair-bundle motility and results in BM, RL, and TM vibration and electrically evoked otoacoustic emissions. After somatic motility is suppressed by salicylate, hair-bundle motility-mediated responses can be measured in wild-type but not in TectaC1509G /C1509G mice. The contribution of hair-bundle motility to cochlear amplification will be determined by measuring and comparing electrically evoked responses in TectaC1509G /C1509G and wild-type mice after salicylate application. The new data from this study will provide critical information on the essential role of OHC somatic and hair-bundle motility in cochlear amplification and for understanding mechanisms of auditory disorders in humans.