Neuronal Basis Underlying Volatile Anesthetic Induced Hypnosis

项目总结 尽管在破译麻醉剂对个体离子通道的影响方面取得了很大进展， 麻醉剂产生行为效应的神经底物的鉴定仍然存在 很有挑战性。在表征麻醉状态的关键成分中，我们专注于挥发性麻醉剂- 诱导催眠，定义为对非有害刺激缺乏意识。了解麻醉剂如何 催眠的产生已经不仅仅是神经科学的中心问题，正如过去的多份报告所述 十年表明，现有的全麻药每年可能会伤害美国4000万患者中的一部分 需要麻醉的人。麻醉剂诱导的无意识的一个假设替代方案是产生一种 一种可逆的生理无意识状态，如睡眠，在这种状态下病人被锁在门外 处于清醒状态的人还能从自然睡眠中恢复体力。在我们最初的获奖期间，我们 发现挥发性麻醉剂确实直接使促进睡眠的视前前部的一部分去极化 下丘脑(POAH)神经元。我们的实验室和其他实验室也表明，每一种测试的全身麻醉剂 (氯胺酮除外)去极化腹外侧视前(VLPO)神经元。然而，重叠的确切程度 在触发内源性睡眠的神经元和因麻醉暴露而激活的神经元之间以及 麻醉剂“劫持”内源性睡眠回路的后果尚不清楚。中心假说 这种更新的特点是，挥发性麻醉剂将通过增强活性来传递部分催眠特性。 内源性POAH神经元，但神经元激活关键差异将区分内源性POAH神经元 在麻醉催眠状态下的NREM睡眠。在目标1中，我们将使用一种新的遗传编码的钙检测器来 在活体中永久标记睡眠活跃的POAH神经元，并随后确定 也被挥发性麻醉剂去极化。同样，我们将测定麻醉剂去极化的比例 在NREM和REM睡眠期间活跃的POAH神经元。在目标2中，我们将在体内采用开创性的 内窥镜显微镜观察和确定POAH神经元在小鼠周期中的实时神经元活动 自然地跨越觉醒、NREM和REM睡眠以及在相同的POAH神经元中 仔细滴定的挥发性麻醉剂暴露。最后，在目标3中，我们将在体内调节睡眠活跃的活动 POAH神经元通过神经元回路标记和新近发现的组织实现精确控制 区分睡眠活跃神经元和状态淡漠神经元或觉醒活跃神经元的特异性表达标志物 邻里。病毒驱动的“Designer受体”的条件性表达 药物“(DREADDS)将允许我们去极化或超极化POAH神经元以确定随后的 激活或抑制睡眠激活的POAH神经元对麻醉状态稳定性的影响。 累积起来，这些目标将使我们能够评估催眠引起的神经元汇聚/发散。 挥发性麻醉剂和睡眠。

PROJECT SUMMARY Although much progress has been made deciphering the effects of anesthetics upon individual ion channels, identification of the neural substrates upon which anesthetics act to produce their behavioral effects remains challenging. Of the key components that characterize the anesthetized state, we focus on volatile anesthetic- induced hypnosis, defined as a lack of awareness to non-noxious stimuli. Understanding how anesthetics produce hypnosis has become more than a central question for neuroscience, as multiple reports over the past decade suggest that existing general anesthetics may annually harm a subset of the 40 million US patients who require anesthesia. One hypothetical alternative to anesthetic-induced unconsciousness is to generate a state of reversible physiological unconsciousness, such as sleep, in which the patient is locked out of access to the state of wakefulness yet recoups restorative benefits of natural sleep. In our original award period, we discovered that volatile anesthetics do directly depolarize a subset of sleep-promoting, preoptic anterior hypothalamic (POAH) neurons. Our lab and others have also shown that every tested general anesthetic (except for ketamine) depolarizes ventrolateral preoptic (VLPO) neurons. However, the exact degree of overlap among neurons triggering endogenous sleep and those activated by anesthetic exposure as well as precise consequences of anesthetic “hijacking” of endogenous sleep circuitry remain unknown. The central hypothesis of this renewal is that volatile anesthetics will impart a portion of their hypnotic properties by enhancing activity in endogenous POAH neurons, but that key differences in neuronal activation will distinguish endogenous NREM sleep from anesthetic hypnosis. In Aim 1, we will use a novel genetically encoded calcium detector to permanently mark sleep-active POAH neurons in vivo and subsequently determine the exact fraction that is also depolarized by volatile anesthetics. Similarly, we will determine the fraction of anesthetic-depolarized POAH neurons that active during NREM and REM sleep. In Aim 2, we will employ groundbreaking in vivo endoscopic microscopy to visualize and determine real time neuronal activity of POAH neurons as mice cycle naturally across wakefulness, NREM, and REM sleep as well as in these same POAH neurons during a carefully titrated volatile anesthetic exposure. Finally, in Aim 3 we will modulate in vivo activity of sleep-active POAH neurons achieving precise control through neuronal circuit labeling and by recently discovered tissue specific expression markers that distinguish sleep-active neurons from state-indifferent or wake-active neighbors. Conditional expression of virally driven “designer receptors exclusively activated by designer drugs” (DREADDs) will permit us to depolarize or hyperpolarize POAH neurons to determine the ensuing effects of activating or inhibiting sleep-active POAH neurons upon the stability of the anesthetic state. Cumulatively, these aims will enable us to assess the neuronal convergence/divergence of hypnosis caused by volatile anesthetics and sleep.