ENVIRONMENTAL LIGHT AND RETINAL MEMBRANE DEVELOPMENT

本提案的长期目标是更好地了解 可见光诱导的光感受器细胞损伤机制和 光暴露后发生的修复和恢复过程。 我们将研究年龄和光照饲养环境对 视网膜光损伤的大鼠维持长达一年， 弱周期性光照或黑暗的受控条件。我们的重点将是 光引发的核内事件和视觉细胞反应， 作为氧化的标记物或标记感光细胞 死亡 为了检测强光照射对小鼠脑组织的氧化作用， 视网膜，将在以下大鼠中进行光损伤易感性的比较： 各种年龄，有或没有抗氧化剂预处理。标准 将使用连续曝光技术，以及曝光 加速光损伤过程的范例。我们将测试 老年大鼠适应饲养条件突变的能力 和抵抗光损伤相比，年轻的动物， 从弱循环光到黑暗，反之亦然。时间 光致视网膜损伤与修复的关系 将研究光暴露后的恢复情况，以了解事件 导致视觉细胞死亡或存活。 生化指标 视细胞损失，包括视紫红质分析和视细胞DNA 内容，将通过协作形态评估予以补充 视觉细胞和视网膜色素上皮细胞的损伤。我们 将研究DNA损伤的时间过程和相互关系， 修复、应激蛋白反应、mRNA诱导和酶活性 在光照期间和在光照后的暗恢复期间。 这将通过限制性内切酶的Southern分析来完成。 来自在光感受器中表达或不表达的基因的片段， 溴化乙锭法检测视网膜总DNA链断裂的Poisson分析 染色琼脂糖凝胶电泳，并通过HPLC测定氧化 鸟苷核苷。西方和北方分析 电泳分离的视觉传导和热休克蛋白 mRNA将与酶的动力学分析一起沿着使用， 对视觉细胞功能很重要。本提案的具体目标 主要是：（1）确定年龄和抗氧化剂的影响 （二甲基硫脲）处理特定分子在损伤激烈 在弱循环光下饲养长达12个月的大鼠的光暴露，或 在黑暗中(2)为了确定年龄和光线变化的影响， 饲养条件对影响感光细胞的特定分子 细胞功能和光损伤敏感性。(3)确定时间 视细胞损伤后的修复和恢复过程 在有或没有RPE损伤的大鼠中对强可见光的敏感性。的 光诱导视紫红质介导的事件的表征 年轻和年老大鼠的光感受器可能提供有用的见解， 光对动物的伤害机制， 长期光照史和年龄相关的视觉细胞丧失之间的关系， 人类

The long term objectives of this proposal are a better understanding of the mechanisms of visible light induced photoreceptor cell damage and of the processes of repair and recovery that occur following light exposure. We will study the influences of age and light rearing environment on retinal light damage in rats maintained for up to one year under carefully controlled conditions of weak cyclic light or darkness. Our focus will be on light initiated events in the nucleus and on visual cell responses that serve as markers of oxidation or that mark the photoreceptor cell for death. To examine the oxidative effects of intense light exposure in the retina, comparisons of light damage susceptibility will be made in rats of various ages, with or without antioxidant pretreatment. Standard techniques of continuous light exposure will be used, as well as exposure paradigms that accelerate the light damage process. We will test the ability of older rats to adapt to an abrupt change in rearing conditions and to resist light damage in comparison to young animals, following a change from weak cyclic light to darkness and vice versa. The temporal relationship between light initiated retinal damage and repair and recovery following light exposure will be studied to understand the events which lead to visual cell death or survival. Biochemical measures of visual cell loss, including rhodopsin analysis and visual cell DNA content, will be complemented by collaborative morphological evaluations of damage in the visual cells and in the retinal pigment epithelium. We will study the time course and interrelationships of DNA damage and repair, stress protein responses, mRNA induction and enzyme activities during light and during the dark recovery period following light exposure. This will be accomplished by Southern analysis of restriction enzyme fragments from genes expressed or not expressed in photoreceptors, by Poisson analysis of total retinal DNA strand breaks using ethidium bromide stained agarose gel electrophoresis, and by HPLC determination of oxidized guanosine nucleosides. Western, and Northern analysis of electrophoretically separated visual transduction and heat shock proteins and mRNAs will be used along with kinetic analysis of enzymes thought to be important for visual cell function. The specific aims of this proposal are: (1) To determine the effects of age and antioxidant (dimethylthiourea) treatment on specific molecules during damaging intense light exposure of rats reared for up to 12 months in weak cyclic light or in darkness. (2) To determine the effects of age and a change in light rearing conditions on specific molecules which can impact photoreceptor cell function and light damage susceptibility. (3) To determine the time course of visual cell repair and recovery from damage following exposure to intense visible light in rats with or without RPE damage. The characterization of light induced-rhodopsin mediated events in the photoreceptors of young and older rats may provide useful insights into the mechanisms of light damage in animals and into the relationship between long term light history and age related visual cell loss in humans.