Synaptic and Functional Changes in Cochlear and Vestibular Hair Cells of Tmc Mutant Mice

项目摘要/摘要 跨膜通道样蛋白1和2(TMC1和TMC2)是感觉转导所必需的。 听觉和前庭毛细胞。靶向缺失Tmc1(Tmc1Δ/Δ)的小鼠表现为耳聋 两者缺失者(Δ/Δ、TMC2、Δ/Δ)均表现为耳聋、前庭功能障碍和感觉传导缺失。头发 仅表达TMC1的细胞具有较小的单通道电导和较低的钙通透性 而不是只表达TMC2的基因。而毛束形态、机械转导和毛细胞存活 已经在TMC突变小鼠中进行了评估，感觉转导缺失或受损对 带状突触的维持还没有被研究过。耳蜗毛细胞和前庭毛细胞利用这些 突触介导充满谷氨酸的囊泡的同步释放，以实现时间上的精确传递 听觉上的暗示。突触带缺失或数量改变可导致突触异常 传播、噪声中语音辨别能力受损，以及耳鸣等知觉异常。这项研究 一项提案试图描述感觉受损/缺失的突触和功能后果 耳蜗器和前庭器官的转导。目标1的重点是描述内毛细胞的特征 TMC突变小鼠和野生型小鼠不同发育时间点的突触。初步结果 Tmc1Δ/Δ和Tmc1Δ/ΔTmc2Δ/Δ小鼠在以后的时间点显示突触数量减少，但在 Tmc2Δ/Δ小鼠，提示Tmc1的缺失是观察到的内毛细胞带变化的原因 突触。TMC1和TMC2不仅是听力所必需的，也是正常前庭功能所必需的。 Tmc1Δ/ΔTmc2Δ/Δ小鼠表现出异常行为，包括摇头，仰卧不能直立 摆好姿势，开始转圈。虽然TMC突变小鼠的行为已经被表征，但前庭的状态 外围国家还没有被详细研究过。目标2的重点是分析前庭毛细胞突触 TMC突变小鼠的球囊和球囊及前庭感觉直接评估前庭功能 诱发电位(VsEP)。近年来的研究证实了病毒载体的安全性和有效性 并利用这些病毒基因疗法恢复缺乏听功能的小鼠的听觉功能 TMC1.然而，通过基因治疗恢复前庭功能和突触的研究尚未得到证实。 目标3的重点是评估TMC突变小鼠在基因治疗后的突触和功能恢复 将注射后获得的数据与目标1和目标2收集的数据进行比较。 该提案将为功能区突触维持的机制提供新的见解，增强我们的 对前庭外周的了解，并为继续开发修复的基因疗法提供信息 听力和平衡感。

Project Summary/Abstract Transmembrane channel-like proteins 1 and 2 (TMC1 and TMC2) are required for sensory transduction in auditory and vestibular hair cells. Mice with targeted deletion of Tmc1 (Tmc1Δ/Δ) exhibit deafness and those lacking both (Tmc1Δ/ΔTmc2Δ/Δ) display deafness, vestibular dysfunction, and absent sensory transduction. Hair cells exclusively expressing Tmc1 have smaller single-channel conductance and lower calcium permeability than those expressing only Tmc2. While hair bundle morphology, mechanotransduction, and hair cell survival have been assessed in Tmc mutant mice, consequences of absent or impaired sensory transduction on the maintenance of ribbon synapses have not been examined. Cochlear and vestibular hair cells utilize these synapses to mediate synchronous release of glutamate-filled vesicles for temporally precise transmission of auditory cues. Absent or altered numbers of synaptic ribbons can contribute to abnormal synaptic transmission, impaired speech-in-noise discrimination, and perceptual anomalies like tinnitus. This research proposal seeks to characterize the synaptic and functional consequences of impaired/absent sensory transduction in the cochlea and vestibular organs. The focus of Aim 1 is to characterize inner hair cell synapses at various developmental timepoints in Tmc mutant mice and wildtype mice. Preliminary results demonstrate decreased synapse numbers in Tmc1Δ/Δ and Tmc1Δ/ΔTmc2Δ/Δ mice at later time points but not in Tmc2Δ/Δ mice, suggesting the absence of Tmc1 accounts for the observed changes in inner hair cell ribbon synapses. Tmc1 and Tmc2 are not only required for hearing but also necessary for normal vestibular function. Tmc1Δ/ΔTmc2Δ/Δ mice demonstrate abnormal behaviors including head-bobbing, inability to right from supine positions, and circling. While behavior has been characterized in Tmc mutant mice, the status of the vestibular periphery has not been examined in detail. The focus of Aim 2 is to analyze vestibular hair cell synapses in utricles and saccules of Tmc mutant mice and to directly assess vestibular function using vestibular sensory evoked potentials (VsEPs). Studies in recent years have verified the safety and efficacy of viral vectors for delivery into mouse cochlea and utilized these viral gene therapies to restore auditory function in mice lacking Tmc1. However, recovery of vestibular function and synapses via gene therapy has not yet been examined. The focus of Aim 3 is to assess synaptic and functional recovery in Tmc mutant mice following gene therapy by comparing data acquired after injections to those collected from Aims 1 and 2. Data generated from this proposal will provide novel insight into the mechanisms underlying ribbon synapse maintenance, enhance our understanding of the vestibular periphery, and inform continued development of gene therapies for restoration of hearing and balance.