Anesthesia and Sleep: Mechanisms of Generating Two Similar Yet Distinct Unconscious States in the Medulla

尽管麻醉剂导致昏迷的确切机制尚不清楚，但有 一些麻醉剂激活调节睡眠的神经回路并抑制神经系统促进睡眠的证据 醒着的。尽管全身麻醉和睡眠都激活了看似相似，如果不是相同的子集， 神经元，这两种无意识状态之间有明显的差异，包括唤醒的程度 阈值变化和状态转换的时间尺度。这些基础的神经机制是相关的，但 人们对不同的潜意识状态知之甚少。 副面部区域(PZ)最近被认为是一种非快速眼动(Non-REM)促进睡眠 具体地说，在非REM睡眠期间，PZ(PZ-GABA)中的GABA能神经元是活跃的。我的初选 数据表明，在异氟醚暴露期间，PZ-GABA也是活跃的，并且PZ-GABA的消融增加 对异氟醚的抗药性。结果还表明，PZ内的非GABA能神经元也参与其中 在异氟醚诱导的催眠中。最重要的问题是，驱动不同状态的神经电路是如何 通过首先检查PZ-GABA神经元，非REM睡眠和异氟醚麻醉的汇聚和发散，以及 然后扩展到PZ之外，考虑延髓中的所有细胞类型。据推测，这些不同的 内源性和药物诱导的无意识状态是由部分重叠的共享回路产生的，但 关键状态的差异源于不同的细胞激活模式。 我们在这项提案中将解决的三个关键问题是：1)急性可逆激活/抑制 促进睡眠的PZ神经元如何改变麻醉敏感性？2)PZ的细胞构成是什么，以及 在每种无意识状态下，哪些细胞被激活？3)哪些元素有重叠和不同之处 异氟醚暴露时的脑干神经回路与非快速眼动时的脑干神经回路之间的关系 睡了吗？这些问题将通过麻醉和睡眠表型分析来解决，单细胞水平 转录本分析采用单核RNA测序和多重原位杂交。 通过在活性群体中定向重组(TRAP)对活性神经元集合的旁侧比较。 拟议的项目将揭示脑干神经回路的潜在机制，包括 PZ，调节这两种不同的无意识状态。了解大脑是如何控制状态的 昏迷对于临床实践是至关重要的。它可以带来更有效和更安全的睡眠激素和新的潜力 镇静剂、催眠药、麻醉剂，有朝一日可能被用于治疗失眠和发作性睡病等睡眠障碍。

Although the exact mechanisms by which anesthetics induce unconsciousness remain unknown, there is evidence that some anesthetics activate neural circuits regulating sleep and inhibit neural systems promoting waking. Despite general anesthesia and sleep both activating a subset of seemingly similar, if not identical, neurons, there are clear differences between the two unconscious states, including the degree of arousal threshold changes and the timescale of state transition. The neural mechanisms underlying these related, yet distinct unconscious states are poorly understood. The parafacial zone (PZ) has recently been identified as a non-rapid-eye-movement (non-REM) sleep-promoting region; specifically, GABAergic neurons in the PZ (PZ-GABA) are active during non-REM sleep. My preliminary data demonstrate that PZ-GABA are also active during isoflurane exposure, and ablation of PZ-GABA increases resistance to isoflurane. The results also suggest that non-GABAergic neurons within the PZ are also involved in isoflurane-induced hypnosis. The overarching question asks how the neural circuitry driving distinct states of non-REM sleep and isoflurane anesthesia converge and diverge by first examining in PZ-GABA neurons, and then expanding beyond the PZ to consider all cell types in the medulla. It is hypothesized that these distinct endogenous and drug-induced unconscious states are generated by partially overlapping shared circuits but that key state differences arise from distinctive cellular activation patterns. The three key questions we will address during this proposal are: 1) Does acute reversible activation/inhibition of the PZ sleep-promoting neurons alter anesthetic sensitivity? 2) What is the cellular makeup of the PZ, and which cells are activated during each unconscious state? and 3) What are the overlapping and different elements between the brainstem neural circuits engaged during isoflurane exposure and those engaged during non-REM sleep? These questions will be addressed by anesthetic and sleep phenotyping assays, the single-cell level transcriptomic analysis by single nucleus RNA sequencing followed by multiplex in situ hybridization, and side- by-side comparison of ensembles of active neurons by Targeted Recombination in Active Population (TRAP). The proposed projects will uncover the underlying mechanism of how the brainstem neural circuits, including PZ, mediate these two different unconscious states. Understanding how the brain controls states of unconsciousness is vital for clinical practice. It can lead to more effective and safer somnogens and new potential sedative hypnotic anesthetics that may one day be used for sleep disorders such as insomnia and narcolepsy.