Optogenetic Control of Vigilance State Transition

描述（由申请人提供）：下丘脑泌素，也称为食欲素，是两种神经肽递质，仅由下丘脑中的几千个神经元产生。在小鼠、犬和人类中的研究表明，下丘脑泌素系统的功能丧失导致发作性睡病/癫痫。我们最近发现，使用光激活阳离子通道，视紫红质2（ChR 2）的遗传定义的Hcrt神经元的光刺激增加了睡眠到觉醒过渡的概率。然而，Hcrt神经元在清醒动物中的作用仍然知之甚少。在这里，我们将使用Hcrt神经元的光遗传学操作来测试这些神经元在清醒期间的阶段性活动是否足以稳定唤醒。首先，我们将使用慢病毒在Hcrt神经元中引入ChR 2，并且我们将使用不同的光刺激模式来操纵Hcrt神经元在清醒期间的活动，以测试持续的活动是否会延长清醒。我们将通过微透析监测光刺激诱导的Hcrt释放的变化。我们还将监测运动活动、焦虑、进食、行为和睡眠/觉醒参数。为了确定Hcrt诱导觉醒的机制，我们将光遗传学刺激Hcrt系统的三个主要突触后靶点：蓝斑中的去甲肾上腺素能神经元、腹侧被盖区中的多巴胺能神经元和基底前脑中的胆碱能神经元。在第三个具体目标中，我们将寻找证据证明在适当条件下阻断几分钟内的阶段性活动足以诱导野生型小鼠的cataerosis。与药理学研究不同，光遗传学提供了前所未有的毫秒级时间分辨率，允许对睡眠不稳定的物种（如啮齿动物）的神经递质功能进行警觉状态特异性分析。 我们的数据将扩大最先进的光遗传学方法在睡眠研究中的应用，并将提供关于下丘脑泌素能系统如何稳定觉醒的机制模型。这些实验也可能导致新的和增强的治疗嗜睡症，失眠和其他睡眠障碍。 公共卫生相关性：我们的数据将扩大最先进的光遗传学方法在睡眠研究中的应用，并将提供关于下丘脑泌素能系统如何稳定觉醒的机制模型。这些实验也可能导致新的和增强的治疗嗜睡症，失眠和其他睡眠障碍。

DESCRIPTION (provided by applicant): The hypocretins, also known as orexins, are two neuropeptide transmitters produced exclusively by a few thousand neurons in the hypothalamus. Studies in mice, dogs and humans have shown that loss-of- function of the hypocretin system results in narcolepsy/cataplexy. We have recently shown that optical stimulation of genetically defined Hcrt neurons using the light activated cation channel, Channelrhodopsin 2 (ChR2) increases the probability of sleep-to-wake transitions. However, the role of Hcrt neurons in awake animals is still poorly understood. Here, we will use optogenetic manipulation of Hcrt neurons to test whether phasic activity of these neurons during wakefulness is sufficient to stabilize arousal. First, we will introduce ChR2 in Hcrt neurons using a lentivirus and we will use different photostimulation patterns to manipulate Hcrt neuronal activity during wakefulness to test whether sustained activity extends waking. We will monitor photostimulation-induced changes in Hcrt release by microdialysis. We will also monitor locomotor activity, anxiety, feeding, behavior and sleep/wake parameters. To determine the mechanism by which Hcrt induces wakefulness, we will optogenetically stimulate three of the main postsynaptic targets of the Hcrt system: noradrenergic neurons in the locus coeruleus, dopaminergic neurons in the ventral tegmental area, and cholinergic neurons in the basal forebrain. In a third specific aim, we will seek evidence demonstrating that blocking phasic activity during a few minutes under the appropriate conditions is sufficient to induce cataplexy in wild-type mice. Unlike pharmacological studies, optogenetics offers unprecedented millisecond scale temporal resolution, which allows vigilance state-specific analysis of neurotransmitter function in species with unconsolidated sleep such as rodents. Our data will expand the use of state-of-the-art optogenetic methods in sleep studies and will provide mechanistic models on how the hypocretinergic system stabilizes arousal. These experiments may also lead to new and enhanced treatments for narcolepsy, insomnia and other sleep disorders. PUBLIC HEALTH RELEVANCE: Our data will expand the use of state-of-the-art optogenetic methods in sleep studies and will provide mechanistic models on how the hypocretinergic system stabilizes arousal. These experiments may also lead to new and enhanced treatments for narcolepsy, insomnia and other sleep disorders.