Experimental study of the cochlear amplifier

描述（由申请人提供）：哺乳动物耳蜗通过一种被称为“耳蜗放大器”的主动放大机制，利用代谢能促进基底膜（BM）振动，实现了其卓越的灵敏度、精细的频率选择性和巨大的动态范围。虽然外毛细胞（OHCs）可以通过体细胞和毛束运动产生力，但耳蜗放大的机械机制尚不清楚。本应用程序是通过使用我们最近开发的扫描低相干外差干涉仪在正常和转基因耳蜗中进行一组新颖的实验，研究体细胞和毛束运动在耳蜗放大中的作用。具体目的一是测量敏感小鼠和沙鼠耳蜗的基底膜、网状层（RL）和毡膜（TM）振动。假设是，在敏感的小鼠耳蜗中，基于ohc的耳蜗放大器在RL产生的非线性振动比在BM产生的振动更大。小鼠耳蜗的BM和RL振动高度敏感，并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一样，如沙鼠。该实验的数据对研究小鼠和沙鼠的耳蜗微力学具有重要意义。具体目的二将通过测量小鼠耳蜗无体动力时耳蜗的BM、RL和TM振动来确定OHC体动力的作用。假设是，在prestin 499敲入小鼠中，缺乏体细胞运动性导致BM、RL和TM振动的灵敏度降低、锐调谐丧失和非线性。由于prestin 499敲入小鼠OHCs的机电转导、刚度和形态正常，因此预期的变化将归因于缺乏体细胞运动性。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。假设缩短和分离的TM具有独立的毛束，影响了TectaC1509G/C1509G小鼠的毛束运动性、机电转导和耳蜗放大，降低了声诱发BM、RL和TM振动的灵敏度、尖锐调谐和非线性。然而，电刺激激活体细胞和毛束的运动，导致BM、RL和TM振动和电诱发的耳声发射。在水杨酸抑制体细胞运动后，在野生型小鼠中可以测量毛束运动介导的反应，而在TectaC1509G /C1509G小鼠中则没有。通过测量和比较水杨酸应用后TectaC1509G /C1509G和野生型小鼠的电诱发反应来确定毛束运动对耳蜗放大的贡献。这项研究的新数据将为OHC体细胞和毛束运动在耳蜗放大中的重要作用以及理解人类听觉障碍的机制提供重要信息。

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.