Optogenetics-assisted identification of the rapid stress receptor in zebrafish

与压力有关的障碍不仅包括精神障碍，如创伤后压力障碍（PTSD）、精神分裂症、焦虑和抑郁，而且还包括心脏、代谢和血管问题，代表着巨大的社会成本，而且正在迅速上升。糖皮质激素（GC）是关键的保守应激激素之一，它是各种应激诱导的生理和行为变化的主要调节剂。GC在快速和长期时域中具有许多目标和功能。关于GC如何介导长期细胞变化，有大量的信息可用。与此形成鲜明对比的是，尽管经过数十年的努力，GC快速调节动物行为和生理的机制在很大程度上仍是未知的。在这一研究领域的主要瓶颈一直是无法确定的受体负责GC的快速效应，这被认为是一种新的GPCR不同于GC受体介导的长期影响。目前在应激领域的方法还没有产生解决这个问题的方案，主要是因为快速和特异性地触发GC反应的方法还没有可用。该项目的目标是设计一种新的策略，利用斑马鱼的光遗传学来克服这一技术限制，并解决这一重要的生物和医学挑战。具体来说，我们建议在这项研究中使用光遗传学开发一种高通量的快速GC效应的体内测定方法，并使用候选反向遗传方法找到快速GC受体。为了验证受体的身份，我们将首次在斑马鱼中开发和实施可光激活的GPCR。最后，我们将利用这个合作项目提供的独特机会，生成一个更大的光激活GPCR库，重点关注大脑功能中识别的受体，并将其传播给社区。该文库将解决目前缺乏光遗传学工具来研究斑马鱼中这一重要受体的问题。因此，利用光遗传学，这项工作将确定新的应激受体，这将为更好地了解应激障碍开辟新的途径，并为研究参与大脑功能各个方面的受体创造重要的光遗传学资源。

Stress-related disorders, which include not only psychiatric disturbances such as post-traumatic stress disorder (PTSD), schizophrenia, anxiety and depression, but also heart, metabolic and vascular problems, represent tremendous societal cost, which is rapidly rising. One of the key conserved stress hormones are Glucocorticoids (GCs), which serve as a master regulator of diverse stress-induced changes in physiology and behavior. GCs have many targets and functions in both rapid and long-term time domain. There is large amount of information available on how GCs mediate long-term cellular changes. In stark contrast, despite decades of efforts, the mechanism by which GCs rapidly regulate animal behavior and physiology is largely unknown. The major bottleneck in this research area has been inability to identify the receptor responsible for fast effects of GCs, which is thought to be a novel GPCR distinct from the GC receptors that mediate long-term effects. Current approaches in the stress field have not yielded a solution to this problem primarily because methods to rapidly and specifically trigger GC responses have not been available. The goal of this project is to devise a new strategy using optogenetics in zebrafish to overcome this technical limitation and address this important biological and medical challenge. Specifically, we propose in this study to use optogenetics to develop a high throughput in vivo assay for rapid GC effects and to find the rapid GC receptor using a candidate reverse genetic approach. To validate the identity of the receptor, we will develop and implement for the first time light-activatable GPCRs in zebrafish. Finally, we will use the unique opportunity offered by this collaborative project to generate a larger library of light-activatable GPCRs with a focus on receptors identified in brain function and disseminate them to the community. This library will address the current lack of optogenetic tools to study this important receptor class in zebrafish. Thus, using optogenetics, this work will identify novel stress receptor(s), which will open new avenues to better understand stress disorders and create an important optogenetic resource for investigating receptors involved in diverse aspects of brain function.