Abstract: Objective: This study aims to identify the potential targets and signaling pathways of the rhizome of Polygonatum odoratum (Yuzhu in traditional Chinese medicine) in the treatment of depression, through bioinformatics and network pharmacology, and to preliminarily explore its efficacy through PC12 cell experiments. Methods: Screening for differential gene expression was conducted by downloading the gene expression profile dataset GSE98793 and relevant information from the GPL570 platform. Component screening was conducted through the TCMSP database. The "drug-component-target" and PPI networks were constructed using Cytoscape 3.7.1 software. GO and KEGG enrichment analyses were performed using the DAVID database. Molecular docking verification of the main active components and targets was performed using SYBYL-X 2.0 software. After inducing a PC12 cell model of depression with glutamic acid (Glu), alcohol extracts of Yuzhu were administered in low, medium, and high doses to examine their protective effects against PC12 cell damage. Results: Twenty-three antidepressant active components and 34 active targets were identified for Yuzhu. Among the active components, palmitic acid, lauric acid, and (+)-cedrol were identified as the primary active components through the "drug-component-target" network analysis. The PPI network analysis showed that PTGS2, PTGS1, SLC6A2, GABRA1, and TNF were key targets. A total of 209 GO entries were obtained through GO enrichment analysis, including 149 biological processes (BP), 23 cellular components (CC), and 37 molecular functions (MF), primarily involving the Chemical carcinogenesis-receptor activation, Small cell lung cancer, and Adipocytokine signaling pathway. Molecular docking analysis revealed that the main active components of Yuzhu exhibited high docking stability with major targets of depression. In vitro cell experiments demonstrated that 600 μg/mL alcohol extracts of Yuzhu significantly ameliorated Glu-induced damage in PC12 cells. Conclusion: Yuzhu may exert its antidepressant effects through inflammatory response, oxidative stress, and prostaglandin synthesis pathways.