The gut-brain axis is dysregulated as a consequence of alterations in the gut microbiota. These alterations increase toxic microbial metabolites, endotoxemia, and the release of immune mediators and contribute to the development of depression. Cubebin is a dibenzyl butyrolactone lignan, and its stem is also known as Agaru in Tibetan areas, it is commonly used as a sedative and tranquilizing medicine. This study aimed to investigate the effects of cubebin on chronic stress-induced depression-like behavior in mice. Cubebin was observed to mitigate depressive-like behavior in chronic unpredictable mild stress (CUMS) mice, influence the restoration of their cerebral cortex and hippocampal tissue morphology, and enhance the abundance of relevant intestinal flora in depression model mice, particularly by decreasing the abundance of Clostridium, Dorea, and Ruminococcus. The final protein function expression was normalized by regulating depression-related metabolic pathways. Concomitantly, the concentrations of neurotransmitters serotonin (5-HT), norepinephrine (NE), and dopamine (DA) in the brains of mice in the model group were enhanced, and their depressive symptoms were mitigated. Our study findings suggest that cubebin may ameliorate CUMS-induced depression in mice by modulating the microbe-gut-brain axis, elucidating the key effect of gut metabolites on depressive symptoms.