Spectral and Metabolic Basis of Visual Responses

视觉色素再生是哺乳动物维持视觉敏感度的绝对要求 视网膜。然而，色素再生与以下敏感度恢复之间的关系 暴露在强光下(暗适应)目前还知之甚少。对这些基本原理的说明 机制是非常重要的，因为色素再生的损害和由此导致的减慢 感光细胞的暗适应是包括Oguchi在内的许多致盲眼病的特征 疾病，Stargardt‘s病，Leber’s先天性黑色素，眼底白斑，以及与年龄相关的 黄斑病变。总体目标是确定分离的杆状感光细胞的细胞机制(S)。 来自视网膜色素上皮，调节色素再生和暗适应的速度 哺乳动物的视杆，这一过程比哺乳动物的视锥细胞慢得多。恢复敏感度 将电生理学测量的全反式视黄醇还原为全反式视黄醇后的色素 漂白将用微量荧光法测量，色素再生率将测量 显微分光光度法。这种多学科的方法将应用于野生型(WT)和转基因 小鼠视杆色素漂白前后及外源11-顺式视网膜处理后 再生视觉色素。提出了两个目标：在目标I中，我们将确定 调节从视网膜色素中分离的杆状感光细胞的视色素再生速度 暴露在强光下的皮肤上皮细胞。我们将检验这样的假说：生命的衰变 漂白的光产物(Meta III)限制了色素再生的速度，在再生过程中11-cis 视网膜从细胞外间隙的脂质结合伙伴漂白的视盘中分裂成细胞内盘 视蛋白通过被动扩散包围视杆，视蛋白在视杆细胞内盘中的滞留速度减慢 与视蛋白位于质膜中的视锥相比，色素再生速度更快。在AIM II中，我们 将确定和表征再生的视觉色素导致 与我们的合作者Jeannie Chen博士一起，我们将恢复杆状感光细胞的视觉敏感性 验证漂白和再生的小鼠视杆的敏感性恢复在暂时性发生的假说 和与C-末端残基的去磷酸化有关的机械上不同的阶段 再生视紫红质和/或转导蛋白易位。我们将获得的知识是 了解正常和疾病相关条件下的暗适应，这是发育所必需的 治疗失明眼疾的新疗法。

Visual pigment regeneration is an absolute requirement for maintenance of visual sensitivity in the mammalian retina. However, the relationship between pigment regeneration and the recovery of sensitivity following exposure to bright light (dark adaptation) is presently poorly understood. The elucidation of these fundamental mechanisms is of great importance, as the impairment of pigment regeneration and the resulting slowing of dark-adaptation in photoreceptor cells are hallmarks of a number of blinding eye diseases, including Oguchi's disease, Stargardt's disease, Leber's congential amaurosis, fundus albipunctatis, and age-related maculopathy. The overall aims are to determine the cellular mechanism(s) within rod photoreceptors isolated from the retinal pigment epithelium that regulate the rate of pigment regeneration and dark adaptation in mammalian rods, processes that are substantially slower than in mammalian cones. Recovery of sensitivity will be measured electrophysiologically, the reduction of all-trans retinal to all-trans retinol following pigment bleaching will be measured microfluorometrically, the rate of pigment regeneration will be measured microspectrophotometrically. This multidisciplinary approach will be applied to wild-type (WT) and transgenic mouse rods before and following pigment bleaching, and after exogenous treatment with 11-cis retinal to regenerate visual pigment. Two Aims are proposed: In Aim I we will determine the cellular mechanisms that regulate the rate of visual pigment regeneration in rod photoreceptors isolated from the retinal pigment epithelium following their exposure to bright light. We will test the hypotheses that the decay of long-lived photoproducts of bleaching (Meta III) limits the rate of pigment regeneration, that during regeneration 11-cis retinal partitions into intracellular disks of bleached rods from lipid binding partners in the extracellular space surrounding rods by a passive diffusion, and that the residence of opsin in intracellular disks in rods slows pigment regeneration rate compared to cones in which opsin is located in the plasma membrane. In Aim II we will determine and characterize the time course of mechanisms by which regenerated visual pigment leads to recovery of visual sensitivity in rod photoreceptors Together with our collaborator, Dr. Jeannie Chen,we will test the hypotheses that recovery of sensitivity in bleached and regenerated mouse rods occurs in temporally and mechanistically distinct stages that are associated with dephosphorylation of C-terminal residues on regenerated rhodopsin and/or transducin translocation. The knowledge we will gain is prerequisite for understanding dark adaptation under normal and disease-related conditions, and is required for development of novel therapies to treat blinding eye conditions.