Intracellular Calcium in Hair Cells

描述（由申请人提供）：长期目标是了解听觉毛细胞的机械传导机制和耳蜗张力异位组织的潜在因素。实验将集中在细胞内ca2 +在调节换能器通道适应和毛束力学中的作用。毛细胞反应将在分离的爬行动物和哺乳动物耳蜗中测量，并将与细胞内ca2 +成像相结合。具体目的是：(1)记录啮齿动物毛细胞在听力开始前后的传感器电流，记录其特性随成熟和耳蜗位置的变化。(2)表征单个换能器通道，并研究ca2 +调制及其随频率的变化。比较海龟和啮齿动物的结果可以说明换能器通道性质的演变；(3)测量毛细胞立体纤毛束的力学特性，寻找哺乳动物外毛细胞自发和主动的束运动。活动束运动和外毛细胞收缩之间的相互作用将用于评估这两个过程在哺乳动物耳蜗放大和调谐中的作用。(4)通过单通道记录和特定通道异构体的免疫标记来测量龟耳蜗中K+通道切片变异的分布。结果可能提供深入了解耳蜗张力异位组织的起源。(5)测量和改变毛束中的ca2 +浓度，并将其与传感器通道激活和主动毛束运动的控制联系起来。将研究ca2 +缓冲和吸收到细胞内室，特别是线粒体，对限制ca2 +瞬态的贡献。由于毛细胞承受较大的ca2 +负荷，ca2 +稳态的紊乱可能是细胞死亡的主要原因。ca2 +对换能器通道的调制可能是所有毛细胞共同的，并且可能是在噪声暴露、耳毒性物质中毒或衰老期间造成不可逆损伤的通道。老化或过度刺激导致的听力丧失通常仅限于高频，并与耳蜗底部毛细胞的退化有关。这项工作将通过映射传感器通道与耳蜗位置的数量和性质来解决灵敏度差异的原因。

DESCRIPTION (provided by applicant): The long-term objectives are to understand the cellular mechanisms of mechanotransduction in auditory hair cells and the factors underlying the cochlea's tonotopic organization. Experiments will focus on the role of intracellular Ca 2+ in regulating transducer channel adaptation and hair bundle mechanics. Hair cell responses will be measured in the isolated cochleas of both reptiles and mammals and will be combined with intracellular Ca 2+ imaging. Specific aims are: (1) to record transducer currents in rodent hair cells before and after the onset of hearing, documenting the changes in properties with maturation and with cochlear location. (2) to characterize single transducer channels and investigate their modulation by Ca 2+ and variation with frequency. Comparison of turtle and rodent results may illustrate the evolution of transducer channel properties; (3) to measure the mechanical properties of hair cell stereociliary bundles and search for spontaneous and active bundle motion in mammalian outer hair cells. The interaction between active bundle motion and outer hair cell contractility will be used to assess the roles of the two processes in amplification and tuning in the mammalian cochlea. (4) to measure the distribution of K+ channel slice variants in the turtle cochlea from both single channel recording and immunolabeling for specific channel isoforms. The results may provide insight into the origin of the cochlea's tonotopic organization. (5) to measure and alter Ca 2+ concentration in hair bundles and relate it to control of transducer channel activation and active hair bundle motion. The contributions of Ca 2+ buffering and uptake into intracellular compartments, especially the mitochondria, to limit Ca 2+ transients will be studied. Since hair cells experience large Ca 2+ loads, disturbance of Ca 2+ homeostasis may be a leading cause of cell death. Ca 2+ modulation of transducer channels is probably common to all hair cells and may be the conduit of irreversible damage during noise exposure, poisoning with ototoxic agents or aging. Loss of hearing with aging or over-stimulation is often restricted to high frequencies and linked to degeneration of hair cells at the base of the cochlea. The work will address reasons for the differential sensitivity by mapping number and properties of transducer channels with cochlear location.