SPECTRAL AND METABOLIC BASIS OF VISUAL RESPONSES

描述(改编自申请者摘要)：本项目将重点关注 调节脊椎动物视网膜杆和视网膜杆敏感性的机制 圆锥体。膜电流的电生理记录， 视觉色素的显微分光光度测定及荧光分析 细胞内钙离子([Ca~(2+)]_i)的测定将从单细胞 视黄醇及其类似物结合的视杆和视锥 黑暗和随后的亮光(漂白)来确定胞浆有多大 信使物质在暗适应过程中受到调节。三总 问题将被解决：(1)视黄醇的性质是什么 相互作用是在黑暗和漂白期间：最低限度的适应 视蛋白和视网膜的结构相互作用是恢复视神经功能所必需的 将测定暗适应过程中的敏感性；PDE和鸟苷酸 循环酶的速度将被估计在已经漂白和 用11-顺式视网膜及其类似物治疗；13-去甲基视网膜和 10-去甲基视网膜将被加入漂白的视杆和视锥中以 测试扭转在视网膜C-11：C-12键中的重要性 暗适应；(2)[Ca2+]i在光适应中的作用 与色素再生：胞浆钙和膜电流的变化 将在孤立的暗适应和漂白适应的杆状和圆锥体中测量 在存在和不存在视网膜及其类似物的情况下 暗适应中敏感度的恢复与暗适应的关系 漂白后视网膜中的[Ca~(2+)]i；9-去甲基视网膜 以确定适应的早期阶段是否 由于视紫红质关闭，因为9-去甲基视紫红质被认为表现出 异常延长的激活寿命；无论这是否延长 寿命是通过改变[Ca~(2+)]i来确定的；和(3) 是否需要视蛋白结合部位的全视网膜占位 以适应视素。后一个问题将通过测量 漂白棒和漂白棒中PDE和环化酶速度的敏感性和估测 在已知浸出剂存在和不存在的情况下的圆锥体 全反式视网膜/视黄醇来自完整的光感受器。

DESCRIPTION (Adapted from applicant's abstract): This project will focus on mechanisms which regulate the sensitivity of vertebrate retinal rods and cones. Electrophysiological recordings of membrane current, microspectrophotometric measurements of visual pigment and fluorometric determinations of cytosolic calcium ([Ca2+]i) will be obtained from solitary rods and cones in which retinal and its analogs have been incorporated in darkness and following bright light (bleaching) to determine how cytosolic messenger substances are regulated during dark-adaptation. Three general questions will be addressed: (1) what the nature of the opsin-retinoid interaction is during dark- and bleaching-adaptation: the minimum structural interactions of opsin and retinal required for recovery of sensitivity during dark-adaptation will be determined; PDE and guanylyl cyclase velocities will be estimated in cells which have been bleached and treated with 11-cis retinal and analogs; and 13-demethyl retinal and 10-demethyl retinal will be incorporated into bleached rods and cones to test the importance of torsion in the C-11:C-12 bond in retinal during dark-adaptation; (2) what the involvement of [Ca2+]i is in light adaptation and pigment regeneration: changes in cytosolic calcium and membrane current will be measured in solitary dark-adapted and bleach-adapted rods and cones in the presence and absence of retinal and its analogs in order to determine the relationship between recovery of sensitivity during dark-adaptation and levels of [Ca2+]i; 9-demethyl retinal will be incorporated into bleached photoreceptors in order to determine whether an early phase of adaptation is due to rhodopsin turn-off since 9-demethyl rhodopsin is thought to exhibit an abnormally prolonged activated lifetime; whether or not this prolonged lifetime is modified by changes in [Ca2+]i will be determined; and (3) whether all-trans retinal occupancy of the opsin binding site is required for opsin adaptation. This latter question will be addressed by measuring sensitivity and estimating PDE and cyclase velocities in bleached rods and cones in the presence and absence of agents which are known to leach all-trans retinal/retinol from the intact photoreceptors.