Ion Conductances in the Retinal Pigment Epithelium

描述(由申请人提供)：光感受器的健康和完整性关键取决于其细胞外微环境的组成和体积。视网膜下间隙的离子组成和体积的调节是通过跨视网膜色素上皮(RPE)的离子和水的运输完成的，RPE是并列在光感受器外节和脉络膜血供之间的多功能单层细胞。RPE转运是一组不同的离子转运蛋白和通道的协同活动的结果，这些离子转运蛋白和通道驻留在其顶端和基底侧膜上。随着视网膜活动的变化，视网膜细胞释放的化学信号扩散到RPE，并启动其运输的调整，以补偿光感受器微环境的变化。这些运输过程的中断或它们的调节可能会导致视网膜下腔的不利变化，从而导致视网膜疾病。这些通道和转运体还负责维持RPE细胞的细胞内组成，如果受到干扰，可能会对其他关键的RPE功能产生不利影响，如吞噬作用、光感受器外段的降解以及维生素A的运输和代谢。我们的总体目标是了解钾(K+)通道参与调节视网膜下间隙和RPE细胞质中液体的体积和离子组成的机制。最近的研究发现，在RPE中存在一种类似于神经元中M型电流的外向整流钾电流。本建议的具体目的是：(1)确定RPE中M型电导背后的KCNQ通道的亚基组成；(2)确定M型电导是定位于根尖膜还是基底外侧膜；(3)确定M型电导是否受到已知的调节特定类型KCNQ通道的药物和信号通路的调节；以及(4)确定M型电导在RPE细胞体积调节中的作用。这些目标将使用分子、生化、免疫组织化学、成像和电生理技术的组合来实现，以研究M型通道的结构、功能和调节。这些研究的结果将有助于更好地理解这些至关重要的运输蛋白如何在RPE中发挥作用，以帮助维持健康的光感受器微环境。与公共健康相关：光感受器的健康和完整性关键取决于其细胞外微环境的组成和体积。视网膜下间隙的离子组成和体积的调节是通过跨视网膜色素上皮(RPE)的离子和水的运输完成的，RPE是并列在光感受器外段和脉络膜血供之间的单层细胞。RPE转运是驻留在其膜上的一组不同的离子转运蛋白和通道协调活动的结果。随着视网膜活动的变化，视网膜细胞释放的化学信号扩散到RPE，并启动其运输的调整，以补偿光感受器微环境的变化。这些运输过程的中断或它们的调节可能会导致视网膜下腔的不利变化，从而导致视网膜疾病。

DESCRIPTION (provided by applicant): The health and integrity of photoreceptors critically depend on the composition and volume of their extracellular microenvironment. Regulation of the ionic composition and volume of the subretinal space is accomplished by the transport of ions and water across the retinal pigment epithelium (RPE), a multifunctional monolayer of cells juxtaposed between the photoreceptor outer segments and the choroidal blood supply. RPE transport is the result of the coordinated activity of a diverse group of ion transport proteins and channels residing in its apical and basolateral membranes. With changes in retinal activity, chemical signals released by retinal cells diffuse to the RPE and initiate adjustments in its transport to compensate for alterations in the photoreceptor microenvironment. Disruption of these transport processes or their regulation may cause adverse changes in the subretinal space, contributing to retinal disease. These channels and transporters are also responsible for maintaining the intracellular composition of the RPE cell, which, if disturbed, could adversely affect other key RPE functions such as phagocytosis, the degradation of photoreceptor outer segments, and vitamin A transport and metabolism. Our overall goal is to understand the mechanisms by which potassium (K+) channels participate in the regulation of the volume and ionic composition of the fluid in both the subretinal space and the RPE cytoplasm. Recent studies have identified in the RPE an outwardly rectifying K+ current that resembles the M-type current in neurons. The specific aims of this proposal are to: (1) determine the subunit composition of KCNQ channels that underlie the M-type conductance in the RPE; (2) determine whether the M-type conductance is localized to the apical or basolateral membrane; (3) determine whether the M-type conductance is modulated by pharmacologic agents and signaling pathways known to modulate specific types of KCNQ channels; and (4) determine the role of the M-type conductance in the regulation of RPE cell volume. These aims will be pursued using a combination of molecular, biochemical, immunohistochemical, imaging, and electrophysiological techniques to investigate M-type channel structure, function, and regulation. The outcome of these studies will result in a better understanding of how these critically important transport proteins operate in the RPE to help maintain a healthy photoreceptor microenvironment. PUBLIC HEALTH RELEVANCE: The health and integrity of photoreceptors critically depend on the composition and volume of their extracellular microenvironment. Regulation of the ionic composition and volume of the subretinal space is accomplished by the transport of ions and water across the retinal pigment epithelium (RPE), a monolayer of cells juxtaposed between the photoreceptor outer segments and the choroidal blood supply. RPE transport is the result of the coordinated activity of a diverse group of ion transport proteins and channels residing in its membranes. With changes in retinal activity, chemical signals released by retinal cells diffuse to the RPE and initiate adjustments in its transport to compensate for alterations in the photoreceptor microenvironment. Disruption of these transport processes or their regulation may cause adverse changes in the subretinal space, contributing to retinal disease.