Ion Channel Function in Auditory & Vestibular hair cells

这些研究集中在KCNQ钾通道在听觉和前庭系统的感觉毛细胞中的功能。在这类新发现的电压门控离子通道的五个成员中，有四个成员的突变会导致人类遗传性疾病。这些蛋白中至少有三个在听觉和前庭外周表达：KCNQ1、3和4。其中两个蛋白KCNQ1和4的突变会导致严重的听觉功能障碍。虽然这些遗传性疾病的病因尚不清楚，但这些严重的感觉缺陷意味着KNCQ蛋白在正常的听觉功能中扮演着重要的角色。这个项目有两个主要目标。第一个目标是将KCNQ钾通道的表达与听觉和前庭毛细胞的正常生理联系起来。这将为KCNQ基因家族的突变如何导致病理状态提供新的见解。第二个目标是研究KCNQ通道在前庭外周突触传递中的作用。具体地说，我们将测试这一假设，即I型毛细胞传入突触利用一种新的K依赖神经传递形式。为了解决这些问题，我们制定了一项共同战略。这些钾通道的孔形成区域内的突变以显性方式作用于阻断传导。利用病毒介导的基因转移，我们将在器官培养的细胞中表达突变的KCNQ基因，这些细胞来自小鼠的听觉和前庭器官。突变的KCNQ基因在正常细胞中的表达会抑制野生型KCNQ亚基的活性。为了检测功能中断，我们将表征感染细胞的电生理特性，通过绿色荧光蛋白的共同表达来鉴定，以及邻近的未感染对照细胞。因此，我们将以一种特定和受控的方式将分子同一性与其生理联系联系起来。

The studies proposed here focus on the function of KCNQ potassium channels in the sensory hair cells of the auditory and vestibular systems. Mutations in four of the five members of this newly discovered class of voltage-gated ion channels cause inherited human diseases. At least three of these proteins are expressed in the auditory and vestibular periphery: KCNQ1, 3 and 4. Mutations in two of them, KCNQ1 and 4, cause severe auditory dysfunction. Although the etiologies of these inherited conditions are not well understood, the profound sensory deficits imply an important role for KNCQ proteins in normal auditory function. This project has two main goals. This first goal is to correlate expression of KCNQ potassium channels with the normal physiology of auditory and vestibular hair cells. This will provide new insight into how mutations in the KCNQ gene family lead to pathological states. The second goal is to investigate the role of KCNQ channels in synaptic transmission in the vestibular periphery. Specifically, we will test the hypothesis that the type I hair cell afferent synapse utilizes a novel form of K+-dependent neurotransmission. To address these questions we have devised a common strategy. A mutation within the pore-forming region of these potassium channels acts in a dominant manner to block conduction. Using virus-mediated gene transfer we will express mutant KCNQ genes in cells of organotypic cultures from the mouse auditory and vestibular organs. Expression of mutant KCNQ genes in normal cells will suppress the activity of wildtype KCNQ subunits. To assay for disrupted function we will characterize the electrophysiological properties of infected cells, identified by coexpression of green fluorescent protein, and neighboring uninfected control cells. Thus, in a specific and controlled manner we will link a molecular identity with its physiologic correlate.