Roles of CRF-PACAP systems in sleep in mice (Carlezon)

摘要：项目 3 许多精神疾病都会影响睡眠。有些人患有与压力相关的疾病（例如，后 创伤性应激障碍 [PTSD]）睡眠少于正常水平，而其他人睡眠较多，或睡眠分散 反映更频繁的睡眠和觉醒发作的睡眠模式。有大量证据表明 压力会扰乱睡眠，我们已经证明，小鼠的慢性压力会导致睡眠和睡眠的深刻改变。 昼夜节律。相比之下，睡眠失调对介导压力效应的大脑系统的影响是 没有彻底理解。睡眠不足或受限的时期可能会产生分子改变， 进而影响睡眠。在啮齿类动物中的研究表明了 CRF 之间的重要相似点和不同点 （促肾上腺皮质激素释放因子）和 PACAP（垂体腺苷酸环化酶激活多肽）对 行为。尽管这两种肽都会增加听觉惊吓，但临床前和临床试验中经常使用的一种警惕性措施 焦虑和恐惧的临床研究显示，PACAP 效果往往是持久的（持续 >1 周），而 CRF 效果 尽快解决。这些类型的发现，以及 CRF 和 PACAP 系统中已知的遗传改变 在易患压力相关疾病的人群中，提供比较和对比神经生物学的基本原理 这些肽。 CRF 和 PACAP 对睡眠的影响尚未直接比较，特别是超过 较长的一段时间。我们的前提是这些肽系统影响睡眠并受睡眠影响。我们的 假设这些肽与睡眠之间存在相互关系，因此激活 CRF 和/或 PACAP 系统将影响睡眠，相反，睡眠限制将影响 CRF 和/或 PACAP 系统，但这些影响的持续时间会有所不同。测试将在雄性和雌性小鼠中进行 植入无线发射器，可连续收集脑电图、肌电图、体温和 几周的活动数据。分子分析将重点关注与情绪方面有关的大脑区域 压力的影响因素，包括杏仁核 (AMG)、终纹床核 (BNST) 和前额皮质 (PFC)， 以及传统上与睡眠和生物节律有关的区域。我们还将进行电路映射 研究旨在提供有关调节情绪行为的区域与那些区域之间的神经联系的见解 调节睡眠。在目标 1 中，我们将比较改变 CRF 和 PACAP 系统功能对睡眠的影响 和生物节律（昼夜节律）。我们还将检查 PAC1R 或 CRFR1 消融的影响 在 AMG 中，对试验数据的跟踪显示，压力会增加该区域 PAC1R 的表达。在目标 2 中，我们 将使用以下方法直接比较急性和重复睡眠限制对 PACAP 和 CRF 系统的影响 RNAscope 可量化任何变化的幅度并识别发生变化的细胞类型。瞄准 3，我们将使用跨突触病毒标记来绘制与睡眠和情绪有关的回路之间的连接 行为。项目 3 实验是根据现有知识设计的，但正如 Conte 中心所预期的那样， 我们将利用所有项目的新发现，通过管理核心来完善和优化我们的目标。

SUMMARY: PROJECT 3 Sleep is affected in many psychiatric illnesses. Some with people with stress-related disorders (e.g., Post- Traumatic Stress Disorder [PTSD]) sleep less than normal, whereas others sleep more, or have fragmented sleep patterns that reflect more frequent bouts of sleep and wakefulness. There is considerable evidence that stress disrupts sleep, and we have shown that chronic stress in mice causes profound alterations in sleep and diurnal rhythms. In contrast, the effects of sleep dysregulation on brain systems that mediate stress effects are not thoroughly understood. Periods of poor or restricted sleep may produce molecular alterations that subsequently affect sleep. Work in rodents demonstrates important similarities and differences between CRF (corticotropin-releasing factor) and PACAP (pituitary adenylate cyclase-activating polypeptide) effects on behavior. Although both peptides increase acoustic startle, a measure of vigilance often used in preclinical and clinical studies of anxiety and fear, PACAP effects tend to be persistent (lasting >1 week), whereas CRF effects resolve quickly. These types of findings, together with known genetic alterations in CRF and PACAP systems in people vulnerable to stress-related illness, provide a rationale for comparing and contrasting the neurobiology of these peptides. The effects of CRF and PACAP on sleep have not been directly compared, particularly over extended periods of time. Our premise is that these peptide systems affect—and are affected by—sleep. Our hypothesis is that there is a reciprocal relationship between these peptides and sleep, such that activation of CRF and/or PACAP systems will affect sleep and, conversely, sleep restriction will affect CRF and/or PACAP systems, but that the persistence of these effects will differ. Tests will be conducted in male and female mice implanted with wireless transmitters that enable continuous collection of EEG, EMG, body temperature, and activity data for several weeks. Molecular analyses will focus on brain areas implicated in the emotional aspects of stress, including amygdala (AMG), bed nucleus of the stria terminalis (BNST), and prefrontal cortex (PFC), as well as areas more traditionally implicated in sleep and biological rhythms. We will also perform circuit mapping studies to provide insights on neural connections between regions that regulate emotional behavior and those that regulate sleep. In Aim 1, we will compare the effects of altering CRF and PACAP system function on sleep and biological (diurnal and circadian) rhythms. We will also examine the effects of PAC1R or CRFR1 ablation in AMG, following up on pilot data showing that stress increases PAC1R expression in this region. In Aim 2, we will directly compare the effects of acute and repeated sleep restriction on PACAP and CRF systems using RNAscope to quantify the magnitude of any changes and identify the cell-types in which changes occur. In Aim 3, we will use trans-synaptic viral labeling to map connections between circuits implicated in sleep and emotional behavior. Project 3 experiments are designed based on current knowledge, but as expected with Conte Centers, we will use new discoveries from all projects to refine and optimize our objectives via the Administrative Core.