SYNAPTIC BASIS OF SLEEP CYCLE CONTROL

这一系列研究的主要目的是利用动物的工作 了解其生理和药理机制 控制睡眠，从而为 对人类睡眠障碍的理解和治疗，两者都是初级的 医疗和物理方面的第二把交椅。以前的工作 极大地提高了我们对脑干机制的认识 控制睡眠的快速眼动(REM)阶段，建议 使用乙酰胆碱作为神经递质的脑干神经元 (胆碱能神经元)促进这一阶段的睡眠。此应用程序 建立在这项工作的基础上，并对其进行扩展。要使用的关键技术是 微透析与细胞外单位记录的新组合 在自由活动的猫中，自然睡眠的猫的细胞内记录 并进行大鼠体外切片制备。 有待研究的假设包括腺苷是否增加 长时间清醒(和增加代谢活动)后的行为 作为减少觉醒(W)和增加慢波的一个因素 睡眠(SWS或非REM)睡眠阶段。我们假设腺苷 升高对基底节区胆碱能神经元的作用最强。 前脑和桥中脑区促进W和An的激活 脑电(EEG)。新数据显示腺苷强劲 对中缝背核(DRN)的状态改变也有影响。 微透析法测定胞外腺苷及其递送 腺苷转运抑制剂和伴随单位的记录将 用于体内试验，而体外研究将检查机制 行动的一部分。 含5-羟色胺的DRN神经元抑制作用的假说 胆碱能神经元和允许快速眼动睡眠发生时，这些DRN SWS和REM期间的神经元缓慢放电也将被评估； 体内和体外技术将检测每一种 可能控制DRN放电减慢的四个因素是：GABA， 5-羟色胺的侧支反馈、腺苷与突触前障碍 肾上腺素能输入。我们将使用细胞内活体记录和 双标记法确定REM-ON神经元(=放电 对REM睡眠有选择性的活动，并可能控制这种状态) 以及觉醒和快速眼动神经元(W/R-ON，可能控制 在脑桥中记录W和REM的脑电活动 胆碱能区可确定为胆碱能区。vbl.使用 微透析和单元记录，我们将检验这样的假设 该胆碱能区的REM-ON神经元不同于W/R-ON神经元 通过作用于5HT1a受体的DRN输入的抑制作用。 最后，我们将研究视前区腹外侧区。 下丘脑，在那里早期对CFOS蛋白的研究表明 SWS过程中一组神经元的选择性激活。vbl.使用 微透析(MD)我们将评估自发性细胞外 GABA水平在SWS期间下降，提示去抑制，以及 MD灌流荷包牡丹碱是否促进SWS，就像在 初步数据。我们还将评估胆碱能 控制(从基底前脑)和组胺能控制(从 结节乳头状核)。体外工作将检查后和 这些神经递质和其他神经递质对VLPOA的突触前效应 用生物细胞蛋白和GAD免疫组织化学鉴定神经元 贴标签。

The broad purpose of this series of studies is to use work in animals to understand the physiological and pharmacological mechanism controlling sleep, and thereby provide a sound baiss for the understanding and treatment of human sleep disorders, both primary and secondard to medical and physchiatric conditions. Previous work has greatly advanced our knowledge of brainstem mechanisms controlling the rapid eye movement (REM) phase of sleep, suggesting that brainstem neurons using acetylcholine as a neurotransmitter (cholinergic neurons) promote this phase of sleep. This application builds on, and extends this work. The key techniques to be used are a novel combination of microdialysis and extramcellular unit recording in freely moving cats, intracellular recordings in naturally sleeping cats and the rat in vitro slice preparation. Hypotheses to be investigated include whether increases in adenosine following prolonged wakefulness (and increased etabolic activity) act as a factor reducing wakefulness (W) and increasing the Slow Wave Sleep (SWS or nonREM) phase of sleep. We hypothesize adenosine increases act most strongly on cholinergic neurons in the basal forebrain and the mesopontine area that promote W and an activated electroencephalogram (EEG). New data suggest strong adenosine state-altering effects on the dorsal raphe nucleus (DRN) also. Microdialysis measurements of extracellular adenosine and delivery of adenosine transport inhibitors and concomitant unit recordings will be used for in vivo tests, while in vitro studies will examine mechanisms of action. The hypothesis that serotonin-containing DRN neurons disinhibit cholinergic neurons and allow REM sleep to occur when these DRN neurons slow discharge during SWS and REM will also be evaluated; in vivo and in vitro techniques will examine the degree to which each of four factors may control the slowing of DRN discharge: GABA, 5HT collateral feedback, adenosine, and presynaptic disfacilitation of adrenergic input. We will use intracellular in vivo recording and double labeling to determine if the REM-on neurons (=discharge activity selective for REM sleep, and possibly controlling this state) and the Waking- and REM-on neurons (W/R-on, possibly controlling EEG activation in both W and REM) recorded in the mesopontine cholinergic zone can be positively identified as cholinergic. Using microdialysis and unit recording we will test the hypothesis that the REM-on neurons of this cholinergic zone differ from W/R-on neurons by their inhibtability by DRN input acting on 5HT1A receptors. Finally we will examine the Ventrolateral preoptic Area (VLPOA) of hypothalamus, where earlier work with cFos protein indicated a selective activation of a population of neurons during SWS. Using microdialysis (MD) we will evaluate whether spontaneous extracellular GABA levels decrease during SWS, suggesting disinhibition, and whether MD-perfused bicuculline promotes SWS, as it did in preliminary data. We will also evaluate the extent of cholinergic control (from basal forebrain) and of histaminergic control (from the tuberomammillary nucleus). In vitro work will examine the post- and pre-synaptic effects of these and other neurotransmitters on VLPOA neurons identified with biocytin and GAD immunohistochemical labeling.