Guangdong Provincial Key Laboratory of Clinical Research on Traditional Chinese Medicine Syndrome, The Second Affiliated Hospital of Guangzhou University of Chinese Medicine, Guangzhou, 510120, China; Joint Laboratory for Translational Cancer Research of Chinese Medicine of the Ministry of Education, International Institute for Translational Chinese Medicine, School of Pharmaceutical Science, Guangzhou University of Chinese Medicine, Guangzhou, 510006, China; School of Fundamental Medical Science, Guangzhou University of Chinese Medicine, Guangzhou, 510006, China.
Outpatient Department, Guangzhou University of Chinese Medicine, Guangzhou, 510405, China.
J Ethnopharmacol. 2024 Nov 15;334:118529. doi: 10.1016/j.jep.2024.118529. Epub 2024 Jul 6.
Sinisan formula (SNSF), documented in the classic books Shanghan Lun, is known for its ability to regulate liver-qi and treat depression. However, its underlying mechanism, particularly its effects on dynamic real-time neuron activity and circuits remains to be fully elucidated.
This study aimed to investigate the antidepressant effect of SNSF and its central nervous system mechanism on depression-like behaviors, focusing on the prefrontal cortex (PFC) to dorsal raphe nucleus (DRN) neural circuit in a stress-induced adolescent animal model.
SNSF comprised four herbs, the root of Bupleurum chinense DC., the root of Paeonia lactiflora Pall., the fruit of Citrus aurantium L., the rhizome of Glycyrrhiza uralensis Fisch., in equal propotions. The adolescent depression animal model was induced by maternal separation (MS) and chronic restraint stress (CRS). In-vivo multichannel physiological electrodes were implanted into the PFC on PND 28 and animals were recorded 5 times during PND 35-46. From PND 47, the behavioral tests were performed to evaluate the antidepressant efficacy of SNSF. Subsequently, brain tissue was collected for Western blot and immunofluorescence staining analysis. Retro virus was injected into the DRN to explore sources of projections received by serotonergic (5-HTergic) neurons. And the PFC-to-DRN circuit was activated or inhibited through chemogenetic techniques to investigate the effects of SNSF on depression-like behaviors.
Administration of SNSF for 18 days effectively alleviated depression-like behaviors in MS&CRS adolescent mice. The PFC emerged as the primary glutamatergic projection source of the DRN neurons. Following SNSF administration for 13/15/18 days, there was an increase in the firing rate of excitatory neurons and excitatory/inhibitory (E/I) ratio in the PFC. MS&CRS stress let to a reduction in the density of 5-HT+ and CaMKII + neurons in the DRN, accompanied by an increase in the density of GAD + neurons in the DRN, while SNSF administration reversed the alterations. Chemogenetic activation of the PFC-to-DRN circuit rescued the depression-like behaviors induced by MS&CRS, whereas suppression of this circuit attenuated the antidepressant effect of SNSF.
SNSF significantly mitigated depression-like behaviors in MS&CRS mice. SNSF exerts its antidepressant effects by increasing the E/I ratio in the PFC and enhancing glutamatergic projections from the PFC to the DRN.
《伤寒论》中记载的四逆散（SNSF）以调节肝气、治疗抑郁症而闻名。然而，其潜在机制，特别是对动态实时神经元活动和回路的影响，仍有待充分阐明。
本研究旨在探讨 SNSF 对抑郁样行为的抗抑郁作用及其对中枢神经系统的机制，重点关注应激诱导的青少年动物模型中的前额叶皮层（PFC）至背侧中缝核（DRN）神经回路。
SNSF 由四种草药组成，即柴胡、白芍、枳实、甘草，按等比例混合。青少年抑郁动物模型通过母婴分离（MS）和慢性束缚应激（CRS）诱导。在 PND 28 时将体内多通道生理电极植入 PFC，并在 PND 35-46 期间进行 5 次记录。从 PND 47 开始，进行行为测试以评估 SNSF 的抗抑郁疗效。随后，收集脑组织进行 Western blot 和免疫荧光染色分析。通过逆行病毒将 DRN 注射入 5-羟色胺能（5-HTergic）神经元，以探索接受 5-HTergic 神经元投射的来源。通过化学遗传技术激活或抑制 PFC-至-DRN 回路，以研究 SNSF 对抑郁样行为的影响。
SNSF 连续给药 18 天可有效缓解 MS&CRS 青少年小鼠的抑郁样行为。PFC 是 DRN 神经元的主要谷氨酸能投射源。SNSF 给药 13/15/18 天后，PFC 中兴奋性神经元的放电率和兴奋/抑制（E/I）比值增加。MS&CRS 应激导致 DRN 中 5-HT+和 CaMKII+神经元密度降低，同时 DRN 中 GAD+神经元密度增加，而 SNSF 给药可逆转这些改变。化学遗传激活 PFC-至-DRN 回路可挽救 MS&CRS 诱导的抑郁样行为，而抑制该回路则减弱了 SNSF 的抗抑郁作用。
SNSF 可显著减轻 MS&CRS 小鼠的抑郁样行为。SNSF 通过增加 PFC 中的 E/I 比值并增强 PFC 至 DRN 的谷氨酸能投射来发挥抗抑郁作用。
