Jieyu Wuwei Sinisan formula alleviates depression-like behaviors in CUMS mice by regulating synaptic plasticity through PLD2 pathway.

Abstract

ETHNOPHARMACOLOGICAL RELEVANCE: Jieyu Wuwei Sinisan Formula (JWS) is derived from "sinisan", a basic and classical formula of traditional Chinese medicine (TCM) for treating depression by regulating the liver. The disorder of the synaptic plasticity in brain regions is closely to the pathogenesis of depression, yet its mechanism remains to be further clarified. AIM OF THE STUDY: This study aims to investigate the antidepressant effect and precise mechanism of JWS in a validated rodent model of depression. MATERIALS AND METHODS: High performance liquid chromatography was applied to establish the fingerprint of the TCM formula JWS. Depressed mice model was built by chronic unpredictable mild stress (CUMS). Following the administration of multiple doses of JWS, parameters including various behavioral tests and organs indexes to identify the optimal dosage of JWS for treating depression. Effects of JWS on the pathological damage of the prefrontal cortex (PFC) were investigated by hematoxylin-eosin (HE) and Nissl staining. Non-targeted metabolomics analyses were conducted on extracellular fluid (ECF) and brain tissue from the PFC using LC-MS technology to identify differential metabolites and pathways associated with JWS treatment of CUMS-induced depression in mice. Ingenuity Pathway Analysis (IPA) was used to analyze potential key proteins. Potential targets were preliminarily validated using ELISA, WB and Pearson correlation analysis. Finally, WB, Golgi staining, and transmission electron microscopy (TEM) were used for target validation to elucidate the antidepressant mechanism of JWS. RESULTS: JWS was found to not only alleviate depression-like behaviors, but also decrease the anxiety index in elevated plus maze test of the CUMS depressed mice. The results of the organ index evaluation uncovered that the safety of JWS. HE and Nissl staining results indicated that JWS could reverse the pathological damage of the PFC in depressed mice. The results of non-targeted metabolomic analysis of ECF and the PFC were shown that glycerophospholipid metabolism pathway was the key differential metabolic pathway between the control, CUMS and JWS group. Combined with literature research and IPA analysis, the key protein target phospholipase D (PLD) was found, and the ELISA and WB results indicated the activity of PLD2 and the expression of PLD2 protein in the PFC were significantly decreased after CUMS, and increased after JWS treatment. Mechanism studies revealed the JWS exerted antidepressant effect by up-regulating PLD2 pathway, and adjusting synaptic plasticity. CONCLUSION: These findings proved the safety and effectiveness of JWS, revealed the antidepressant mechanism of JWS and provided a basis of JWS for clinical application of depression, especially depression with anxiety states.