Optogenetic dissection of basal forebrain neurons involved in sleep homeostasis

描述（由申请人提供）：鉴于睡眠剥夺（SD）与人类健康、福祉和认知表现的相关性，稳态睡眠调节的生物学基础是睡眠研究的一个重要方面。越来越多的人要么是出于职业需要，要么是出于自愿，长时间保持清醒，要么是在生理上不理想的时间保持清醒。此外，在抑郁症、创伤后应激障碍、阿尔茨海默病和帕金森病等病症中，经常观察到与睡眠不足相关的认知障碍。SD引起注意力和认知障碍以及诱导稳态睡眠反应的中心机制对于设计治疗范例以减轻睡眠缺失的有害影响至关重要，这与NIMH的使命很接近。其中一个大脑区域，基底前脑（BF），除了在促进清醒方面发挥重要作用外，还被认为在平衡睡眠调节以及注意力和认知方面发挥作用。BF由多种利用乙酰胆碱、GABA或谷氨酸的神经元组成。长期以来，BF中神经元组成的复杂性阻碍了对每一种神经元亚型在细胞外神经化学改变和皮层活动调节中的因果作用的清晰理解，这些变化是在SD期间延长神经元激活后嗜睡增加和警觉性降低的基础。最近的研究表明，表达GABA能（PARV-pos GABA）的胆碱能和小白蛋白（PARV）神经元在清醒时都是活跃的，并且能够调节皮层的激活。然而，它们在睡眠稳态中的独特作用尚不清楚。虽然胆碱能和gaba能神经元的神经毒性病变强调了这些神经元在清醒和稳态睡眠反应中的重要性，但通过选择性地操纵每种神经递质特异性神经元细胞类型，可以最好地研究直接的因果评估。该应用程序的总体目标是辨别胆碱能和PARV-pos gaba能神经元激活在稳态睡眠调节中的功能作用的差异。利用最先进的光遗传学技术选择性地操纵胆碱能和PARV-GABA神经元的活动，结合同步多导睡眠图记录来监测皮质脑电图的变化和体内微透析来测量细胞外神经化学变化，我们将测试以下模型：延长SD->BF胆碱能神经元激活->增加细胞外NO和腺苷->抑制清醒活性胆碱能和非胆碱能神经元->增加嗜睡。这些探索性研究的成功完成将(1)验证光遗传学与体内微透析的新组合方法，(2)扩展我们对特定BF神经元亚型在调节皮质脑电图和稳态睡眠反应中的因果作用的理解。

DESCRIPTION (provided by applicant): The biological underpinnings of homeostatic sleep regulation are an important aspect of sleep research given the relevance of sleep deprivation (SD) to human health, well-being, and cognitive performance. More and more people either are forced to, due to vocational demands, or by choice, stay awake for long periods or at biologically non-optimal times of the day. Moreover, sleep loss-associated cognitive impairments are often observed in conditions such as depressive disorders, post-traumatic stress disorder, Alzheimer's and Parkinson's diseases. Central mechanisms that mediate the effects of SD causing attention and cognitive impairment as well as induction of homeostatic sleep response are critical to be understood to design treatment paradigms for alleviate such deleterious effects of sleep loss, and is close to the NIMH mission. One of the brain regions, the basal forebrain (BF), in addition to its important role in promoting wakefulness, is also recognized for its role i homeostatic sleep regulation as well as in attention and cognition. The BF consists of a variety of neurons that utilize acetylcholine or GABA or glutamate. The complexity of the neuronal composition in BF has long prevented a clear understanding of the causal role of each neuronal subtype on extracellular neurochemical alterations and modulation of cortical activity that underlies increased sleepiness and reduced alertness following prolonged neuronal activation during SD. Recent studies demonstrated that both cholinergic and parvalbumin (PARV) expressing GABAergic (PARV-pos GABA) neurons are active during wakefulness and are capable of modulating cortical activation. However, their distinct roles in sleep homeostasis are not clear. While neurotoxic lesions of cholinergic and GABAergic neurons have underlined the importance of these neurons in wakefulness and homeostatic sleep response, a direct cause and effect evaluation is best studied by selective manipulation of each neurotransmitter-specific neuronal cell types. The overall goals of this application is to discern the differences in the functional role of cholinergic and PARV-pos GABAergic neuronal activation in homeostatic sleep regulation. Using the state-of-the-art optogenetic technology to selectively manipulate the activities of cholinergic and PARV-GABA neurons combined with simultaneous polysomnographic recordings to monitor changes in cortical EEG and in vivo microdialysis for measuring extracellular neurochemical changes we will test the following model: Prolonged SD->BF cholinergic neuronal activation->increase extracellular NO and adenosine->inhibition of wake active cholinergic and non-cholinergic neurons->increased sleepiness. Successful completion of these exploratory studies will (1) validate a novel combinatorial method of performing optogenetics with in vivo microdialysis, (2) extend our understanding of the causal role of specific BF neuronal subtypes in modulating cortical EEG and homeostatic sleep response. PUBLIC HEALTH RELEVANCE: Sleep deprivation is a wide spread problem with negative impacts on performance efficiency, health and cognition. The current application proposes to use in vivo microdialysis with the state-of-the-art method of optogenetics to selectively manipulate two major wake-active neuronal cell types to discern their role in modulating the homeostatic sleep factors that increase the propensity to sleep and decrease attention and cognition. Understanding the neuronal components involved in mediating neurochemical changes is important towards designing targeted treatment plans to counteract the effects of sleep deprivation.