Molecular characterization of mouse melanopsin and second messenger pathway.

描述（由申请人提供）：哺乳动物生理和行为的许多方面表现出每日24小时节律。这些每日的振荡是昼夜节律，并由称为视交叉上核（SCN）的大脑结构控制。哺乳动物的生物钟不断地被环境光的开始重置。时钟的光夹带需要来自视网膜的输入，视网膜通过视网膜神经节细胞（RGC）的一小部分的轴突投射与SCN通信。令人惊讶的是，不需要视杆和视锥光感受器;相反，投射到SCN的RGC似乎起自主昼夜光感受器的作用，因为它们表现出独立于视杆和视锥驱动的突触输入的光响应。这些SCN投射的RGCs还表达黑视蛋白，一种新的脊椎动物视蛋白，这是在这些细胞中启动光反应所必需的。在所有已知的脊椎动物视蛋白中，黑视蛋白是独特的;它显示出比其他脊椎动物视蛋白更大的序列相似性，以无脊椎动物的横纹肌感光器视蛋白。目前对黑视素的生物化学性质的理解是初步的，其体内第二信使系统尚未最终建立和广泛表征。拟议研究的总体目标是表征黑视蛋白的光谱和生化特性，并确定介导SCN投射RGC的内在光响应的生化途径。我们假设，黑视蛋白形成一种具有类似于与横纹肌光感受器相关的视色素的生物化学特征的色素，并激活基于Gq的信号通路。此外，我们假设黑视素经历光依赖性修饰，其有助于黑视素依赖性信号级联的光适应。为了验证这些假设，我们提出了一个跨学科的方法相结合的生物化学，电生理学，分子遗传学和行为研究。这种方法旨在确定黑视蛋白的光化学，阐明黑视蛋白激活的第二信使途径的性质，并确定是否存在黑视蛋白的光依赖性翻译后修饰。公共卫生相关性：哺乳动物生理和行为的许多方面都表现出每天24小时的节律。这些每日的振荡是昼夜节律，由一种称为视交叉上核的大脑结构控制。哺乳动物的生物钟不断地被环境光的开始重置。时钟的光夹带需要来自视网膜的输入，视网膜通过视网膜神经节细胞的一小部分的轴突投射与视交叉上核通信。令人惊讶的是，经典的光感受器、视杆和视锥不是必需的;相反，视交叉上核投射的视网膜神经节细胞起自主光感受器的作用。这些神经节细胞还表达一种新的脊椎动物视觉色素，称为黑视素，这是启动这些细胞中的光反应所必需的。拟议的研究项目旨在表征黑视素。了解黑视素激活的信号级联反应对感觉生物学和昼夜节律领域具有重要意义。此外，昼夜节律系统紊乱通常伴有神经退行性疾病、睡眠障碍和失明，并且更常见于跨子午线旅行和轮班工作。在未来，阐明黑视蛋白为基础的信号级联反应，应该使我们能够开发成功的药物治疗这些疾病。

DESCRIPTION (provided by applicant): Many aspects of mammalian physiology and behavior exhibit a daily 24 hour rhythm. These daily oscillations are circadian rhythms and are controlled by a brain structure known as the suprachiasmatic nucleus (SCN). The mammalian circadian clock is constantly being reset by the onset of environmental light. Light entrainment of the clock requires input from the retina, which communicates with the SCN via the axonal projections of a small subset of retinal ganglion cells (RGCs). Surprisingly, rod and cone photoreceptors are not required; instead, RGCs that project to the SCN appear to function as autonomous circadian photoreceptors as they exhibit light responses independent of rod- and cone-driven synaptic input. These SCN-projecting RGCs also express melanopsin, a novel vertebrate opsin, which is necessary for initiating the light response in these cells. Among all known vertebrate opsins, melanopsin is unique; it shows greater sequence similarity to invertebrate rhabdomeric photoreceptor opsins than to other vertebrate opsins. The current understanding of melanopsin's biochemical properties is rudimentary and its in vivo second messenger system has yet to be conclusively established and extensively characterized. The overall goal of the proposed research is to characterize the spectral and biochemical properties of melanopsin and to determine the biochemical pathway that mediates the intrinsic light responses of SCN-projecting RGCs. We hypothesize that melanopsin forms a photopigment that has biochemical characteristics similar to visual pigments associated with rhabdomeric photoreceptors and activates a Gq-based signaling pathway. Furthermore, we hypothesize that melanopsin undergoes light-dependent modifications that contribute to light adaptation of the melanopsin-dependent signaling cascade. To test these hypotheses, we propose an interdisciplinary approach combining biochemistry, electrophysiology, molecular genetics and behavioral studies. This approach is designed to determine melanopsin's photochemistry, to elucidate the nature of the second messenger pathway activated by melanopsin, and to determine if there are light-dependent post-translational modifications of melanopsin. PUBLIC HEALTH RELEVANCE: Many aspects of mammalian physiology and behavior exhibit a daily 24 hour rhythm. These daily oscillations are circadian rhythms and are controlled by a brain structure known as the suprachiasmatic nucleus. The mammalian circadian clock is constantly being reset by the onset of environmental light. Light entrainment of the clock requires input from the retina, which communicates with the suprachiasmatic nucleus via the axonal projections of a small subset of retinal ganglion cells. Surprisingly, classical photoreceptors, rods and cones are not necessary; instead, suprachiasmatic nucleus - projecting retinal ganglion cells function as autonomous photoreceptors. These ganglion cells also express a novel vertebrate visual pigment, known as melanopsin, which is necessary for initiating the light response in these cells. The proposed research project aims to characterize melanopsin. Understanding the signaling cascade activated by melanopsin is of great interest and significance for the field of sensory biology and circadian rhythms. In addition, disorders of the circadian system often accompany neurodegenerative diseases, sleep disorders, and blindness, and are more commonly experienced as a result of transmeridian travel and shift work. In the future, elucidation of the melanopsin-based signaling cascade should allow us to develop successful pharmacological treatments for these disorders.