Optogenetic Control of Hypocretin Neurons and Stress

描述(申请人提供)：下丘脑神经元光遗传控制和应激对急性应激反应的神经基础涉及产生促肾上腺皮质激素释放因子(CRF)的神经元的激活和多个稳态回路之间的相互作用。然而，人们对这些互动的细节知之甚少。在小鼠、狗和人类受试者中，下丘脑外侧核(LHCRT)中产生下丘脑(Hcrt)的神经元对维持唤醒稳定性非常重要，因为Hcrt功能的丧失与发作性睡病有关。下丘脑神经元受急性应激激活，接受CRF终末的神经支配，并被CRF去极化。相反，输注hcrt-1可以激活下丘脑-垂体-肾上腺(HPA)轴，而下丘脑-垂体-肾上腺(HPA)受体拮抗剂可以阻断急性应激诱导的ACTH的释放。在这里，我们建议使用一种新开发的光遗传学方法来测试下丘脑下丘脑神经元的活动是否是激活HPA轴所必需的和充分的。在第一个目标中，我们将通过监测Hcrt缺陷小鼠的急性应激反应来确定Hcrt神经元是否是激活HPA轴所必需的。在特定的目标2中，我们将使用光遗传学方法来确定Hcrt神经元的活动是否足以诱导应激样反应。我们还将使用转导慢病毒的小鼠来测试这种激活是必要的，慢病毒在下丘脑细胞中表达可光激活的氯离子通道。这项技术将使我们能够破译与应激反应相关的下丘脑肌素网络的神经密码。在第三个目标中，我们将测试HCRT的功能连接性。我们将测试光刺激对HPA轴的影响是由下丘脑室旁核的CRF信号直接介导的，还是通过其他脑结构间接产生的。这项修订提案中收集的数据将以前所未有的时间分辨率加强我们对应激反应的神经基础的理解，并可能导致针对应激障碍和相关疾病的新疗法，以及确定针对HCRT系统治疗其他疾病的药物的潜在副作用。 公共卫生相关性：这项研究计划中收集的数据将加强我们对压力反应的理解，并可能为普通公众的压力障碍和相关疾病带来新的治疗方法。

DESCRIPTION (provided by applicant): Optogenetic control of hypocretin neurons and stress The neural underpinnings of the response to acute stress involve activation of neurons producing corticotrophin releasing factor (CRF) and interactions between multiple homeostatic circuits. However, the details of these interactions are poorly understood. Hypocretin (Hcrt, also known as Orexin)-producing neurons in the lateral hypothalamus (LH) are important for maintaining arousal stability since loss of Hcrt function has been linked to narcolepsy in mouse, dog and human subjects. Hypocretin neurons are activated by acute stress, receive innervation from CRF terminals and are depolarized by CRF. Conversely, infusion of Hcrt-1 activates the hypothalamo-pituitary-adrenal (HPA) axis, and hypocretin receptor antagonists can block the release of ACTH induced by acute stress. Here we propose to use a newly developed optogenetic method to test whether the activity of hypocretin neurons is necessary and sufficient to activate the HPA axis. In the first aim, we will determine whether Hcrt neurons are necessary to activate the HPA axis by monitoring the acute stress response in Hcrt-deficient mice. In specific aim 2, we will determine whether the activity of Hcrt neurons is sufficient to induce a stress-like response by using an optogenetic approach. We will also test when is this activation required by using mice transduced with a lentivirus expressing a photoactivatable chloride channel in hypocretin cells. This technology will allow us to decipher the neural code of the hypocretin network that is associated with the stress response. In the third aim, we will test the functional connectivity of Hcrt. We will test whether the effects of photostimulation on the HPA axis are mediated directly by CRF signaling in the paraventricular hypothalamic nucleus, or whether the effect is indirect through other brain structures. The data collected in this revised proposal will enhance our understanding of the neural basis of the stress response with unprecedented temporal resolution and may lead to novel therapeutics for stress disorders and related diseases, as well as identify potential side effects for drugs that target the Hcrt system for other disorders. PUBLIC HEALTH RELEVANCE: The data collected in this research proposal will enhance our understanding of stress response and may lead to novel therapeutics for stress disorders and related diseases in the general public.