Physiology of Photoreceptors

说明： 钙是脊椎动物光感受器外节的重要信使。在黑暗中，它通过依赖光的通道快速进入，但同样迅速地被移走，因此其浓度被调节到一个狭窄的范围内。暴露在光中会导致钙的一过性下降，钙是光适应过程中的一个重要信号。钙在视网膜退化中也可能起着至关重要的作用。越来越多的证据表明，外段钙的上升或下降，如果足够大和持续时间长，可以触发细胞程序性死亡。这项应用的目的是了解钙是如何在光感受器中调节的，并在视网膜营养不良的动物模型中确定钙浓度与光感受器退行性变之间的关系。我们将研究钙在杆状外节段圆盘和细胞质之间的运动，使用两种荧光染料独立地标记每个隔室。这些测量应该使我们能够估计钙交换的速度，以显示从视盘中释放或摄取的钙是否足以影响细胞质的浓度。此外，我们还将在动物的视杆上使用荧光染料和吸入电极记录，并对外节蛋白进行有针对性的修饰或删除，以解决以下问题：在长期缺乏维生素A的情况下，视蛋白转导的持续激活和伴随的钙离子减少是退化的原因吗？为什么外节蛋白的一些突变会导致良性静止性夜盲，而另一些则会导致光感受器死亡和失明？需要多大的钙变化才能引发感光细胞死亡？操控钙水平能挽救视杆细胞和视锥细胞的死亡吗？外段钙的增加是否通过扩散到内段，特别是线粒体来杀死光感受器？我们希望我们的实验能够更全面地了解钙在光感受器动态平衡和疾病中的作用。

DESCRIPTION: Calcium is an important messenger in the outer segment of vertebrate photoreceptors. In darkness, it enters through light-dependent channels at a rapid rate, but it is equally rapidly removed, so that its concentration is regulated to within a narrow range. Exposure to light causes a transient decrease in calcium that serves as an important signal during light adaptation. Calcium may also be critically important in retinal degeneration. Increasing evidence indicates that a rise or fall of outer segment calcium, if sufficiently large and prolonged, can trigger programmed cell death. The goal of this application is to understand how calcium is regulated in the photoreceptor, and to determine the relationship between calcium concentration and the rate of photoreceptor degeneration in animal models of retinal dystrophy. We shall study movements of calcium between rod outer segment disks and cytoplasm, using two fluorescent dyes to label each compartment independently. These measurements should enable us to estimate the rate of calcium exchange, to show whether release or uptake of calcium from disks is sufficient to affect the cytoplasmic concentration. We shall in addition use fluorescent dyes and suction-electrode recording on rods in animals with targeted modifications or deletions of outer segment proteins to address the following questions: Is the continuous activation of transduction by opsin and a concomitant decrease in calcium the cause of degeneration during prolonged vitamin A deprivation? Why do some mutations of outer segment proteins cause benign stationary night blindness, but others the death of photoreceptors and blindness? How large a change in calcium is required to trigger photoreceptor death? Can the death of the rods and cones be rescued by manipulating the calcium level? Does an increase in outer segment calcium kill the photoreceptor by diffusing into the inner segment, and in particular into the mitochondria? It is our hope that our experiments will provide a more complete understanding of the role of calcium in photoreceptor homeostasis and disease.