ENVIRONMENTAL LIGHT AND RETINAL MEMBRANE DEVELOPMENT

环境光与视网膜膜发育

• 批准号: 6518282
• 负责人: DANIEL T ORGANISCIAK
• 金额: $ 30.57万
• 依托单位: WRIGHT STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1977
• 资助国家: 美国
• 起止时间: 1977-04-01 至 2004-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6518282
• 关键词: DNA damage; SDS polyacrylamide gel electrophoresis; aging; albino rat; apoptosis; circadian rhythms; cysteine endopeptidases; cytochrome c; electron microscopy; enzyme activity; gene induction /repression; gene mutation; genetic susceptibility; immunocytochemistry; laboratory rat; light adverse effect; light microscopy; membrane biogenesis; northern blottings; oxidative stress; retina degeneration; retinal pigment epithelium; rhodopsin; thiourea; visual photoreceptor; western blottings

DESCRIPTION (Adapted from applicant's abstract): This study seeks to determine the mechanisms of intense light-mediated photoreceptor cell degeneration. The hypotheses to be tested are: (1) that there is a sensitive period within the circadian cycle which results in enhanced susceptibility to retinal light damage and which is mediated by the expression of intrinsic factors in the retina (aim 1) and (2) that susceptibility to light damage is affected by genetic inheritance, which can enhance the rate of photoreceptor cell loss from environmental insult in the form of dim cyclic light exposure or acute intense light treatment (aim 2). This proposal will focus on identifying the circadian expression of intrinsic factors in the retina that enhance or prevent light-induced cell damage leading to apoptotic cell death. The investigator will study the interactions of light-and dark rearing conditions and the timing of intense light treatments in vivo that produce synchronous photoreceptor cell damage in normal albino rats and in transgenic rats with mutations in the rhodopsin primary amino acid sequence. Photoreceptor cell loss will be assayed using rhodopsin and photoreceptor DNA measurements, electrophoretic analysis of DNA fragmentation, and light and electron microscopy of light-damaged retinas. The circadian expression of melatonin and dopamine will be measured and Western and Northern analysis will be used to identify potential modulators of retinal damage. These measurements will be complimented by 2D gel electrophoresis of retinal proteins and ordered differential display of mRNA from retinas at different times of the day and night. The synthetic antioxidant dimethylthiourea will be used as a probe to uncover early events in the mechanism of cell death, including mitochondrial involvement and the potential for oxidative stress to induce apoptosis. In addition, the time course of the appearance of cytoplasmic cytochrome C and the activation of cellular caspases will be determined by antibody-based techniques, immunocytochemistry and enzymatic analysis.

描述(摘自申请者的摘要)：这项研究旨在 强光介导的光感受器细胞机制的确定 退化。要检验的假设是：(1)存在一个 昼夜节律周期内的敏感期，导致 对视网膜光损伤的敏感性，这是由 视网膜内固有因子的表达(目标1)和(2) 对光伤害的敏感性受基因遗传的影响，这种遗传 可以提高环境中感光细胞的损失率 以昏暗的周期性光线暴露或强烈光线的形式侮辱 治疗(目标2)。这项提案将集中于确定昼夜节律 视网膜中增强或抑制内源性因子的表达 光诱导的细胞损伤导致细胞凋亡。这个 研究人员将研究光照和黑暗养育的相互作用 体内强光治疗的条件和时间 对正常白化大鼠产生同步性光感受器细胞损伤 在视紫红质初级氨基酸突变的转基因大鼠中 序列。光感受器细胞损失将使用视紫红质和 光感受器DNA测量，DNA的电泳分析 碎裂，以及光和电子显微镜的光损伤 视网膜。褪黑激素和多巴胺的昼夜表达将是 将使用测量和西部和北部分析来确定 视网膜损伤的潜在调节剂。这些测量结果将是 由视网膜蛋白质双向凝胶电泳仪点播 视网膜在一天中不同时间的差异显示 还有黑夜。合成抗氧化剂二甲基硫脲将用作 一项揭示细胞死亡机制早期事件的探测器， 包括线粒体的参与和氧化的可能性 应激诱导细胞凋亡。此外，在时间进程中 胞质细胞色素C的出现与细胞的活化 半胱氨酸酶将通过基于抗体的技术来确定， 免疫细胞化学和酶分析。