Abstract: OBJECTIVE To analyze the targets and related pathways of paeonol in the treatment of premenstrual dysphoric disorder(PMDD) by network pharmacology and whole animal experiments. METHODS The action targets of paeonol were obtained from the SwissTargetPrediction database; the pathogenesis targets of PMDD were obtained from the OMIM, Drug Bank, Gene Cards and DisGeNET databases; the protein-protein interaction network analysis of the paeonol-PMDD intersection targets was performed by using the STRING database; Gene Ontology(GO) and Kyoto Encyclopedia of Genes and Genomes(KEGG) enrichment analysis for intersection targets using DAVID website; and molecular docking verification for core targets. The PMDD rat model was replicated, and the mRNA expression levels of 5-HT1A, MAOA, and MAOB in the hippocampus as well as 5-HT levels were detected, the action pathways and targets of paeonol in the treatment of PMDD were verified. RESULTS Network pharmacology found that paeonol and PMDD had 16 intersection targets, mainly involving genes in the 5-HT signaling pathway, such as 5-HT1A, MAOA, MAOB, etc; the overall animal test found that paeonol could increase the mRNA expression levels of 5-HT1A, MAOA and MAOB decreased the level of 5-HT in the hippocampus. CONCLUSION In this study, network pharmacology is used to successfully predict the target of paeonol in the treatment of PMDD. Combined with the overall animal experiment, it is confirmed that the mechanism of paeonol in the treatment of PMDD is related to the promotion of 5-HT decomposition and the increase of 5-HT1A expression, which provide theoretical and experimental basis for the mechanism study of paeonol in clinical treatment of PMDD.