VISUAL PIGMENT AND PHOTORECEPTOR ADAPTATION

描述(由申请人提供)：视杆感受器暴露在明亮的(视紫红质漂白)光下会触发视黄醇视觉周期的操作，并显著降低视杆闪光反应的敏感度。随后对视杆的暗适应，除了兴奋衰减(关闭激活的转导中间产物和cGMP补充)外，还需要从“沉默的”脱敏过程中恢复，即使在循环电流接近完全恢复后，这种脱敏过程也会抑制转导信号。体外研究表明，沉默脱敏的幅度远远超过漂白剂诱导的量子捕获量的减少，并已确定了许多可能的贡献反应。然而，目前缺乏关于活体眼睛静默脱敏操作的信息，这是最终了解体内参与反应的机制和相关作用的主要障碍。该项目的一个主要焦点是使用配对闪光视网膜电波(ERG)记录来确定无声脱敏的大小和时间在小鼠和人视杆中的体内漂白依赖性，这是一种新开发的技术，可以无创地确定视杆弱闪光反应的全时间过程。在正常功能的小鼠和人类受试者的视杆中，以及由于缺乏全反式视网膜漂白产物的ABCA4转运体而表现出迟缓兴奋衰退的abcr-/-小鼠，我们将检验兴奋衰减率限制静默脱敏持续时间的特定假设。成对闪光法在定量测定杆状闪光反应中的准确性--具体地说，ERG b波侵袭偏离杆状反应幅度推导的可能性--将在nob小鼠身上进行实验，这是一种b波缺陷模型。该项目的另一个重点是测试最近提出的关于视网膜色素上皮(RPE)中视黄素代谢的假说，这一过程直接与RPE在支持黑暗适应棒中的视紫红质再生方面的作用有关。这一假设--RPE中的11-顺式维甲酸调节RPE基底膜的全反式视黄醇的外流--将在表现出正常或视觉周期操作受损的小鼠身上进行放射性标记实验。该项目的研究结果将促进对视杆暗适应的基础知识，并为进一步研究受试者和实验动物的感光细胞疾病奠定基础。

DESCRIPTION (provided by applicant): Exposure of rod photoreceptors to bright (rhodopsin bleaching) light triggers operation of the retinoid visual cycle and markedly desensitizes the rod flash response. Subsequent dark adaptation of the rods requires, in addition to excitation decay (shut-off of activated transduction intermediates and cGMP replenishment), recovery from "silent" desensitization processes that depress transduction signaling even after the near-complete recovery of circulating current. In vitro studies show that the magnitude of silent desensitization far exceeds that attributable to bleach-induced reduction in quantum catch, and have identified numerous likely contributing reactions. However, the lack of information on the operation of silent desensitization in the living eye is a major current obstacle to ultimately understanding in vivo mechanisms and relative roles of the contributing reactions. A main focus of the project is to determine the in vivo bleaching-dependence of the silent desensitizations's magnitude and timing in mouse and in human rods using paired-flash electroretinographic (ERG) recording, a recently developed technique that permits noninvasive determination of the full time course of the rod weak-flash response. In normally functioning rods of mice and of human subjects, and in abcr-/- mice that exhibit sluggish excitation decay due to lack of the ABCA4 transporter of all-trans retinal bleaching product, we will test the specific hypothesis that excitation decay rate-limits the duration of silent desensitization. The accuracy of the paired-flash method in quantitatively determining the rod flash response -- specifically, the possibility that ERG b-wave intrusion skews derivation of the rod response amplitude -- will be tested in experiments on the nob mouse, a b-wave deficient model. A further focus of the project is to test a recently raised hypothesis concerning retinoid metabolism in the retinal pigment epithelium (RPE), a process directly linked with the RPE's role in supporting rhodopsin regeneration in dark adapting rods. The hypothesis -- that 11-cis retinoid in the RPE regulates the efflux of all-trans retinol at the RPE basolateral membrane -- will be tested in radiolabeling experiments on mice that exhibit normal or impaired visual cycle operation. Results of the project will advance fundamental knowledge of rod dark adaptation and provide foundation for further studies of photoreceptor disease in human subjects and experimental animals.