Depression- and anxiety- like behaviors were evaluated using the sucrose preference test (SPT), open field test (OFT), and elevated plus maze (EPM). Microglial activation and inflammatory mediators were assessed by immunofluorescence, quantitative real-time PCR (qRT-PCR), and Western blot. Levels of kynurenic acid (KYNA), quinolinic acid (QA), and oxidative stress markers were measured using enzyme-linked immunosorbent assay (ELISA) and biochemical kits. In addition, QA-treated BV-2 microglial cells were employed to validate the cellular mechanisms. Liquid chromatography-mass spectrometry (LC-MS) analysis was performed to characterize the chemical profile of ZZCD.

ZZCD significantly improved depression- and anxiety- like behaviors in CUMS rats, suppressed M1-type activation and promoted M2 polarization of microglia, corrected the imbalance of KYNA/QA, and alleviated oxidative stress by reducing reactive oxygen species (ROS) and malondialdehyde (MDA) while enhancing superoxide dismutase (SOD) and catalase (CAT) activities. Moreover, ZZCD inhibited the NF-κB pathway and activated ERK/CREB signaling in both hippocampal tissues and BV-2 cells, accompanied by upregulation of brain-derived neurotrophic factor (BDNF) and synaptic plasticity-related proteins, including postsynaptic density protein 95 (PSD95) and synaptophysin.

ZZCD exerts antidepressant effects through multi-target mechanisms, including regulation of microglial activation, kynurenine pathway balance, oxidative stress reduction, and ERK/CREB pathway activation, which may contribute to the restoration of synaptic plasticity. These findings provide experimental evidence supporting the potential of ZZCD as a traditional Chinese medicine-based intervention for depression.