Properties and Functions of Ca2+ Channels in Hair Cells

毛细胞 Ca2 通道的特性和功能

• 批准号: 6929994
• 负责人: EBENEZER N YAMOAH
• 金额: $ 33.18万
• 依托单位: UNIVERSITY OF CALIFORNIA AT DAVIS
• 依托单位国家: 美国
• 财政年份: 2000
• 资助国家: 美国
• 起止时间: 2000-01-01 至 2010-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6929994
• 关键词: Rana; calcium channel; calcium channel blockers; calcium flux; chickens; cochlea; ear hair cell; ear pharmacology; electrophysiology; hearing; immunoelectron microscopy; in situ hybridization; laboratory mouse; magnesium ion; molecular cloning; neurotransmitter transport; polymerase chain reaction; potassium channel; protein isoforms; voltage /patch clamp; voltage gated channel

Ca regulates multiple functions in hair cells. Our objectives are to understand how multiple voltage-gated Ca channels confer these Ca-dependent functions. The overall thrust of this proposal is to deploy innovative molecular biological, electrophysiological, and imaging techniques- many inspired from previous Ca channel studies- for the discovery of fundamental and newly accessible arenas of Ca channel physiology in hair cells. This thrust drives three aims that address various aspects of hair cell Ca channel physiology, each with fundamental and therapeutic implications. The overall hypothesis is that hair cells express Cav3.x (T-type) Ca channels that may confer hair cell-specific functions (e.g. characteristic bursting activity at certain stages in development) and contribute towards maturation of hair cells. The Aims are: (1) To clarify unresolved aspects of the subtypes of Ca channels in hair cells and their functions. The identity of non L-type Ca channels remains uncertain, despite large potential physiological ramifications. Using biophysical and molecular biological techniques, we will identify the non L-type channels in hair cells as T-type channels and show that alternative splicing of mRNA produces different variants of T-type channels that are specific to hair cells. (2) To determine the role of Ca-channel subtypes in functional development of hair cells. We predict that there is a developmental switch in the expression of VGCCs in hair cells. We expect that different Ca channel subtypes are involved in early developmental processes of hair cells. (3) To define the role of Ca entry through L- and T- type Ca channels in the control of local and global Ca domains in hair cells. For Ca ions to be effective as signals in different intracellular processes, their local concentration has to be precisely controlled to prevent crosstalk between different pathways. We hypothesize that heterogeneous expression of Ca channel subtypes from single to different-sized clusters confers varying Ca domains and multiple Ca dependent functions. Here, we will determine the elementary properties of Ca channels in hair cells by performing whole-cell and single-channel recordings and simultaneous recording of Ca domains using high resolution imaging techniques. The spatial and temporal characteristics of Ca handling in hair cells will be identified and a model of local and global Ca handling will be formulated. Overall, this proposal will answer fundamental unknowns of Ca channel physiology in hair cells.

Ca 调节毛细胞的多种功能。我们的目标是了解多个电压门控 Ca 通道如何赋予这些 Ca 依赖性功能。该提案的总体主旨是部署创新的分子生物学、电生理学和成像技术（许多技术受到以前的 Ca 通道研究的启发），以发现毛细胞中 Ca 通道生理学的基本和新近领域。这一推动力推动了解决毛细胞 Ca 通道生理学各个方面的三个目标，每个目标都具有基础和治疗意义。总体假设是，毛细胞表达 Cav3.x（T 型）Ca 通道，可能赋予毛细胞特异性功能（例如，在发育的某些阶段的特征性爆发活性）并有助于毛细胞的成熟。目的是： (1) 阐明毛细胞中 Ca 通道亚型及其功能的未解决问题。尽管有很大的潜在生理影响，但非 L 型 Ca 通道的身份仍然不确定。利用生物物理和分子生物学技术，我们将毛细胞中的非 L 型通道识别为 T 型通道，并证明 mRNA 的选择性剪接会产生毛细胞特有的 T 型通道的不同变体。 (2)确定Ca通道亚型在毛细胞功能发育中的作用。我们预测毛细胞中 VGCC 的表达存在发育转变。我们预计不同的 Ca 通道亚型参与毛细胞的早期发育过程。 (3) 定义通过 L 型和 T 型 Ca 通道的 Ca 进入在控制毛细胞局部和整体 Ca 结构域中的作用。为了使 Ca 离子在不同的细胞内过程中发挥有效的信号作用，必须精确控制它们的局部浓度，以防止不同途径之间的串扰。我们假设 Ca 通道亚型从单一到不同大小的簇的异质表达赋予不同的 Ca 结构域和多种 Ca 依赖性功能。在这里，我们将通过使用高分辨率成像技术进行全细胞和单通道记录以及同时记录 Ca 域，来确定毛细胞中 Ca 通道的基本特性。将确定毛细胞中 Ca 处理的空间和时间特征，并制定局部和全局 Ca 处理模型。总体而言，该提议将回答毛细胞中 Ca 通道生理学的基本未知数。