薰衣草精油及其有效促眠成分经嗅觉的作用机制与神经环路

Lavender essential oil (LEO) mainly contains linalool, linalyl acetate, 1,8-cineole, β-ocimene, terpinen-4-ol. It has been proved that inhalation of LEO improves sleep in humans. However the active ingredients and the mechanisms involved is not clear, which makes it difficult to be approved in clinic. Our recent studies found that the intranasal delivery of LEO volatile in mice shortened the sleep latency, increased the non-rapid eye movement (NREM) and rapid eye movement (REM) sleep time at beginning and late stage during inactive period respectively. It also inhibited the activity of orexinergic (ORX) neurons in the lateral hypothalamus. And orexinergic neurons were involved in the olfaction afferent projection pathways. Therefore, we hypothesize that LEO and its active ingredients may promote sleep by stimulating the main olfactory epithelium, which in turn inhibits the orexinergic neurons relayed by multi orders of neurons. To verify the hypothesis, a synthesized method including transgenic mice, fiber photometry, optogenetics, trans-synaptic tracing by neurotropic virus, brain tissue clarity and immunofluorescent labeling techniques, precise odor delivery and sleep state resolution and power spectral analysis in free moving mice will be used in this proposal project for revealing the characteristics of sleep states induced by the action of LEO and its active ingredients through olfactory and the possible neuronal circuits involved. The expected results of the project will enrich the knowledge of the sleep regulation by olfactory information, and provide new strategies for treatment of insomnia by LEO in clinic.

薰衣草精油（LEO）主要成分含芳樟醇、乙酸芳樟酯、1,8-桉树脑、β-罗勒烯和松油烯-4-醇。已有研究表明LEO鼻腔吸入可改善人类睡眠，但其有效作用成分和机制不明，难于获得临床认可。我们近期研究发现：小鼠鼻腔精准投递LEO缩短入睡潜伏期；增加非活动期初始与近末时间段非快动眼（NREM）和快动眼（REM）睡眠时间；抑制下丘脑Orexin（ORX）能神经元活动；ORX能神经元参与嗅觉传入投射环路。因此提出“LEO刺激主嗅上皮经多级神经元中继抑制ORX能神经元发挥促眠作用”的假说。本项目拟利用转基因小鼠、光纤记录、光遗传学，结合嗜神经病毒跨突触追踪、脑组织透明与免疫荧光标记等技术，联合自由活动条件下气味的精准投递和睡眠时相解析与能谱分析，揭示LEO及其有效成分通过嗅觉促眠作用的时相特征与神经环路。预期结果将丰富嗅觉信息对睡眠调控理论，为LEO临床治疗失眠症提供新策略。

薰衣草精油（lavender essential oil, LEO）含有芳樟醇（linalool）、乙酸芳樟酯（linalyl acetate）、1,8-桉树脑（1,8-cineole）、β-罗勒烯（β-ocimene）和4-松油醇（4-terpineol）等二十多个复杂单体成分。研究显示LEO鼻腔吸入虽有促进人类睡眠的作用，但其有效成分和作用机制不明，因而未被临床认可。依据我们提出的“LEO刺激主嗅上皮经多级神经元中继抑制Orexin（ORX）能神经元发挥促眠作用”科学假说，本项目分别开展了LEO促眠特征分析、LEO促眠成分筛选和LEO促眠神经环路解析。重要成果与关键数据：（1）光照期鼻腔吸入LEO具有显著缩短入睡潜伏期和促眠的作用。其中25%是具有显著促眠效果的最低浓度。（2）腹腔注射LEO（200 mg/kg）不具有显著的促眠作用，硫酸锌损毁嗅上皮后阻断了鼻腔吸入LEO的促眠作用。（3）LEO各主要成分中，具有显著促眠作用的包括芳樟醇、乙酸芳樟酯和4-松油醇，各促眠成分的作用存在差异。（4）嗅球（olfactory bulb, OB）向睡眠-觉醒调控中枢-下丘脑外侧区（lateral hypothalamus, LH）Orexin能神经元的投射环路中，终纹床核（bed nucleus of the stria terminalis, BNST）γ-氨基丁酸（γ-aminobutyric acid, GABA）能神经元是抑制ORX的关键上游，利用GABAA受体拮抗剂氟马西尼可阻断LEO的促眠作用。提示LEO通过嗅觉途径激活BNST（GABA），进而抑制LH（ORX）神经元发挥促眠作用。本项目成果丰富了嗅觉信息对睡眠的调控理论，为LEO临床治疗失眠症提供了新的策略。