Premenstrual syndrome (PMS), a prevalent neuroendocrine disorder in reproductive-age women, is characterized by emotional disturbances and hormonal imbalances. Honghua Xiaoyao Tablets (HXTs) exhibit clinical promise in PMS therapy. Nevertheless, the therapeutic mechanisms and active constituents underlying its efficacy have not been fully elucidated.

This study aimed to investigate the underlying mechanisms of HXT in ameliorating PMS-associated depressive behaviors through modulation of the hypothalamic-pituitary-adrenal (HPA) axis and ovarian-hippocampal signaling.

A progesterone-withdrawal rat model mimicking PMS symptoms was established to evaluate behavioral, hormonal, and histological changes with and without the treatment of HXT. Network pharmacology identified potential targets of HXT against PMS. Molecular docking and surface plasmon resonance (SPR) were employed to validate interactions between HXT constituents and key targets. Experimental validation included Western blot, immunoprecipitation, immunohistochemistry, and enzyme activity assays to dissect molecular mechanisms using ovarian and hippocampal tissues.

HXT administration significantly alleviated depressive behaviors in PMS rats, evidenced by improved exploratory activity in open-field tests, reduced immobility time in forced swim tests, and restored sucrose preference in sucrose preference tests. It also normalized serum levels of adrenocorticotropic hormone, corticosterone, corticotropin-releasing hormone, follicle-stimulating hormone/luteinizing hormone and norepinephrine, while increasing estradiol, prolactin and 5-hydroxytryptamine. Mechanistically, HXT enhanced ovarian estrogen synthesis by upregulating CYP19A1, thereby rebalancing glucocorticoid and estrogen receptor signaling, inhibited GSK3β overactivation, consequently attenuating neuroinflammation. Molecular docking and SPR analyses confirmed direct binding of key HXT constituents—saikosaponins A/D, gallic acid, benzoylpaeoniflorin, and ononin—to targets along the neuroendocrine-ovarian axis, with dissociation constants ranging from 2.52 to 950.10?μM.

HXT alleviates PMS-associated depression through the major effective constituents via multi-dimensional regulation of neuroendocrine-ovarian circuits, combining HPA axis normalization with SENP1/UBE2I-NR5A1-CYP19A1-mediated estrogen recovery and hippocampal GSK3β/NLRP3-pyroptosis inhibition. This study provides molecular evidence supporting HXT as a multitarget therapeutic agent for PMS-related depressive behaviors, bridging traditional herbal medicine with modern systems pharmacology approaches.