Neuronal circuit controlling sleep-promoting ventrolateral preoptic neurons

项目总结/摘要 睡眠是一个活跃的过程，需要促进睡眠的细胞群的参与。 过去二十年的研究表明，腹外侧视前区表达甘丙肽的神经元 （VLPOGal）神经元是正常睡眠和皮质慢波活动所必需的。然而， 在我们理解VLPOGal神经元通过细胞和突触基础上的几个基本空白 是有规律的。一种用于行为睡眠-觉醒控制的非常有影响力的电路模型是“触发器”模型， 睡眠状态转换，提出睡眠-觉醒转换受相互抑制的调节 在促进睡眠的VLPOGal神经元和单胺能和胆碱能唤醒促进节点之间， 前脑和脑干VLPO和VLPO之间存在相互的解剖学联系， 单胺和胆碱能神经元，这些细胞群分别表现出睡眠和觉醒活性， 射击特征为该模型提供了一般支持。然而，VLPO接收来自 许多其他来源，包括直接参与睡眠的非胆碱能和非单胺能神经元， 尾流调节我们最近发现外侧下丘脑GABA能神经元的激活 通过直接抑制VLPOGal神经元有效地促进唤醒。在这里，我们试图通过以下方式扩展这一发现： 识别控制唤醒水平的其他潜在突触驱动，包括长距离和局部驱动 突触驱动为此，虽然一段时间以来已经认识到VLPOGal神经元在局部神经元中处于低水平。 突触控制，这种内VLPO电路的细节仍然在很大程度上未被表征。我们发现 VLPOGal神经元被一组局部GABA能神经元直接抑制。我们建议这个地方 GABA能回路作为共同的入口节点，通过它传入唤醒和睡眠促进通路 控制VLPOGal神经元，具体的假设是，这是通过1）直接和前馈抑制， 产生唤醒和2）解除VLPOGal神经元的抑制以产生睡眠。因此，目前的建议旨在 首先在体外然后在体内鉴定直接或间接调节VLPOGal的长程传入输入 神经元通过局部GABA能网络驱动行为和皮层唤醒。为此，我们将首先 在特定目的1中使用体外电路映射和VLPO中GABA能传递的局灶性缺失 神经元，以确定用于控制VLPOGal神经元的局部GABA能回路的必要性。在 具体目标2我们将通过体外记录来鉴定VLPO中的突触后神经元， 唤醒和睡眠促进输入。最后，在《特定目标3》中，我们将使用体内光遗传学来测试 在体外切片中首次发现的VLPO的传入输入是产生睡眠和觉醒行为所必需的 的变化和纤维光度法检查这些传入VLPO的状态依赖性活动。鉴于 这一建议试图填补一个巨大的知识空白，我们希望通过这项合作工作取得成果， 对支持睡眠和觉醒调节的大脑回路的重要和新颖的见解。

Project Summary/Abstract Sleep is an active process requiring the participation of delimited nodes of sleep-promoting cell populations. Work over the last twenty years has demonstrated that galanin expressing neurons in the ventrolateral preoptic (VLPOGal) neurons are necessary for normal sleep and cortical slow wave activity. There remain however several fundamental gaps in our understanding of the cellular and synaptic basis by which VLPOGal neurons are regulated. One highly influential circuit model for behavioral sleep-wake control is the `flip-flop' model of sleep-state switching, which proposes that sleep-wake transitions are regulated by a reciprocal inhibition between sleep-promoting VLPOGal neurons and monoaminergic and cholinergic wake-promoting nodes of the forebrain and brainstem. The findings that reciprocal anatomical connections exist between the VLPO and monoamine and cholinergic neurons and that these cell groups exhibit, respectively, sleep- and wake-active firing profiles have provided general support for the model. However, the VLPO receives afferent inputs from many other sources including non-cholinergic and non-monoaminergic neurons directly involved in sleep and wake regulation. We have recently found that activation of GABAergic neurons of the lateral hypothalamus potently promotes arousal by directly inhibiting VLPOGal neurons. Here we seek to extend this finding by identifying other potential synaptic drives that control arousal levels, including both long-range and local synaptic drives. To this end, while it has been appreciated for some time that VLPOGal neurons are under local synaptic control, the details of this intra-VLPO circuit remains largely uncharacterized. We have found that VLPOGal neurons are directly inhibited by a group of local GABAergic neurons. We propose that this local GABAergic circuit serves as common entry node through which afferent wake- and sleep-promoting pathways control VLPOGal neurons, with the specific hypotheses that this occurs by 1) direct and feedforward inhibition to produce arousal and 2) disinhibition of VLPOGal neurons to produce sleep. The current proposal thus seeks to identify, first in vitro and then in vivo, the long-range afferent inputs that directly or indirectly regulate VLPOGal neurons to drive behavioral and cortical arousal via the local GABAergic network. To do so, we will first employ in Specific Aims 1 in vitro circuit mapping and focal deletion of GABAergic transmission in VLPO neurons to determine the necessity of the local GABAergic circuit for the control of VLPOGal neurons. In Specific Aim 2 we will identify with in vitro recordings the postsynaptic neurons in VLPO that are targeted by wake- and sleep-promoting inputs. Finally, in Specific Aim 3 we will use in vivo optogenetics to test whether the afferent inputs to the VLPO first identified in vitro slices are necessary to produce sleep and wake behavioral changes and fiber photometry to examine the state-dependent activity of these afferents to VLPO. Given the large knowledge gap this proposal seeks to fill, we expect results from this collaborative work to provide important and novel insights into the brain circuitry supporting sleep and wake regulation.