CYCLIC NUCLEOTIDES IN CONE METABOLISM AND DISEASE

锥体代谢和疾病中的环状核苷酸

• 批准号: 2158485
• 负责人: DEBORA B FARBER
• 金额: $ 32.05万
• 依托单位: UNIVERSITY OF CALIFORNIA LOS ANGELES
• 依托单位国家: 美国
• 财政年份: 1978
• 资助国家: 美国
• 起止时间: 1978-08-01 至 1996-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2158485
• 关键词: 3'5' cyclic nucleotide phosphodiesterase; Rodentias; adenylate cyclase; affinity chromatography; cone cell; congenital vision disorder; cyclic AMP; cyclic GMP; dogs; enzyme mechanism; gel electrophoresis; genetic library; genetic manipulation; genetic recombination; guanylate cyclase; histochemistry /cytochemistry; human tissue; laboratory mouse; lizards; macular degeneration; nucleic acid probes; nucleic acid sequence; nucleotide metabolism; phosphodiesterase inhibitors; phosphorylation; protein kinase; radioimmunoassay; radiotracer; retinitis pigmentosa; rhodopsin; visual photoreceptor

The objective is to investigate the metabolism of cyclic nucleotides in normal and diseased visual cells with a special emphasis in the less biochemically characterized cones. In addition, we wish to investigate the mechanism by which elevated cyclic nucleotide levels affect the viability of photoreceptor cells in some retinal degenerations. Our work has suggested that in contrast to the function that cGMP plays in rods, cAMP may be more important than cGMP in cone-dominant retinas since cAMP concentration is higher, cAMP levels are decreased by light, and elevated levels of cAMP seem to be more toxic and cause the specific degeneration of cones. However, to establish that cones and rods utilize different cyclic nucleotides as intracellular regulators it is necessary to study isolated cone photoreceptors because the rest of the retina interferes with its own cAMP metabolism. Using isolated cone cells we will study their cyclic nucleotide content, activities, kinetics and regulation of cyclic nucleotide synthetic and degradative enzymes, and investigate if cyclic nucleotide action is modulated by the phosphorylation of specific proteins. We will determine whether light reduces cyclic nucleotide levels and whether activities of the metabolic enzymes are coupled to bleaching of the visual pigment. The same enzyme systems will also be studied in developing cone-like photoreceptors grown in culture, obtained from dissociated retinas of chick embryos. In addition, we will continue to collect baseline data on cyclic nucleotide metabolism in normal human retina to be used for comparison with results from retinas affected with Retinitis Pigmentosa, macular degeneration or other retinal diseases, whenever these retinas become available. To gain insight into how elevated cyclic nucleotide levels participate in visual cell degeneration, we will study first degeneration of rods in the rd mouse, since we have several probes on hand and clues about what components need to be analyzed. In the future, when more is known about cones and probes are developed, we will focus our attention on cone degeneration. We will use antibodies and molecular biology methods to measure concentration and synthesis of cGMP-phosphodiesterase (PDE), the activity of which is abnormal in rd retina, and mRNAs that code for G-protein and rhodopsin, which are needed for PDE activation. We will also clone cDNAs for PDE and rhodopsin from control and rd retinas. Analyses of the cDNA sequences may reveal anomalies associated with the cause of the rd mouse disease and further our understanding of photoreceptor degeneration.

目的是研究环状核苷酸在 正常和患病的视觉细胞，特别强调 生化特征锥。 此外，我们希望调查 升高环核苷酸水平影响生存能力的机制 在某些视网膜变性中的感光细胞。 我们的工作有 建议与CGMP在杆中扮演的功能相反， 自cAMP以来 浓度较高，露营水平通过光降低，并升高 营地水平似乎更具毒性，并导致特定的变性 锥。 但是，要确定锥和杆利用不同的环状 核苷酸作为细胞内调节剂，有必要研究分离的 锥形感受器，因为视网膜的其余部分会干扰其自己 营地代谢。 使用孤立的锥细胞，我们将研究它们的周期性 核苷酸含量，活性，动力学和循环调节 核苷酸合成和降解酶，并研究循环是否循环 核苷酸作用是通过特异性的磷酸化调节的 蛋白质。 我们将确定光是否降低环状核苷酸水平 以及代谢酶的活动是否与漂白 视觉色素。 也将在相同的酶系统中研究 开发从培养物中生长的锥状光感受器，从 分离的雏鸡胚胎视网膜。 此外，我们将继续 收集正常人正常人的环状核苷酸代谢的基线数据 视网膜用于与视网膜影响的结果进行比较 色素性视网膜炎，黄斑变性或其他视网膜疾病， 每当这些视网膜可用时。 洞悉如何提升 环状核苷酸水平参与视觉细胞变性，我们将 研究RD小鼠中杆的首次变性，因为我们有几个 手头的探针以及有关需要分析哪些组件的线索。 在 未来，当开发有关锥体和探针的更多了解时，我们将 将注意力集中在锥变性上。 我们将使用抗体， 测量浓度和合成的分子生物学方法 CGMP磷酸二酯酶（PDE），其活性在RD中异常 视网膜和mRNA为G蛋白和Rhodopsin编码，需要 用于PDE激活。 我们还将克隆cDNA的PDE和Rhodopsin。 控制和RD视网膜。 对cDNA序列的分析可能揭示 与RD小鼠疾病的原因相关的异常，进一步 了解光感受器变性。