Abstract: Objective: To investigate the effects and mechanisms of Morinda officinalis How. on the gut microbiota of chronic restraint stress (CRS) depression model mice. Methods: Eight-week-old male Kunming mice were randomly divided into six groups based on their body weight: control, model, fluoxetine, and M. officinalis How. Low- (25 mg/kg/d), medium- (50 mg/kg/d), and high-dose (100 mg/kg/d) groups were established. CRS was used to establish a mouse model of depression. Depression-like behavior was observed using the sucrose preference (SPT) and tail suspension tests (TST). Morphological changes in the liver, kidneys, and testes were observed using hematoxylin and eosin (HE) staining. Serum 5-hydroxytryptamine (5-HT) levels were measured using enzyme-linked immunosorbent assay (ELISA). Finally, the gut microbiota were analyzed using 16S rRNA gene amplicon sequencing. Results: Compared with the control group, the sucrose preference rate of mice in the model group was significantly reduced, tail suspension time was significantly prolonged (P<0.05), and serum 5-HT level was significantly increased (P<0.01). This caused varying degrees of damage in the liver, kidneys, and testicles, as well as gut microbiota imbalance. After the intervention, 50 mg/kg/d M. officinalis How. showed good antidepressant effects; its sucrose preference rate was significantly increased; tail suspension time was significantly shortened (P<0.05); 5-HT levels were significantly reduced (P<0.01); liver, kidney, and testicular damage returned to normal levels; the abundance of Proteobacteria was significantly increased (P<0.05); and the abundance of Oscillibacter, Eubacterium_xylanophilum_group, Kurthia and unidentified bacterium was increased (P<0.05). Conclusion: These results showed that 50 mg/kg/d M. officinalis How. regulates gut microbiota imbalance and improves depression-like behavior in CRS mice.