Optogenetics-assisted identification of the rapid stress receptor in zebrafish

与压力相关的疾病不仅包括创伤后应激障碍(PTSD)、精神分裂症、焦虑和抑郁等精神障碍，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)是一种关键的保守应激激素，它是各种应激诱导的生理和行为变化的主要调节器。GCS在快速和长期时间域都有许多目标和功能。关于GC如何调节长期细胞变化，有大量的信息可用。与之形成鲜明对比的是，尽管经过几十年的努力，GCs快速调节动物行为和生理的机制在很大程度上是未知的。这一研究领域的主要瓶颈一直是无法确定负责GCs快速效应的受体，这被认为是一种新的GPCR，不同于介导长期效应的GC受体。目前在应力场中的方法还没有产生对这个问题的解决方案，主要是因为还没有快速和具体地触发GC响应的方法。该项目的目标是利用斑马鱼的光遗传学设计一种新的策略，以克服这一技术限制，并解决这一重要的生物和医学挑战。具体地说，在这项研究中，我们建议利用光遗传学来开发一种高通量的体内快速GC效应的检测方法，并使用候选的反向遗传方法来寻找快速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GPCRs。最后，我们将利用这个合作项目提供的独特机会，生成一个更大的可光激活GPCR库，重点放在大脑功能中识别的受体上，并将它们传播给社区。这个文库将解决目前缺乏光遗传学工具来研究斑马鱼这一重要受体类别的问题。因此，利用光遗传学，这项工作将发现新的应激受体(S)，这将为更好地理解应激障碍开辟新的途径，并为研究参与大脑功能不同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.