Background: The pathogenesis of depression is related to the NOD-like receptor protein 3 (NLRP3) inflammasome activation and low level of mitophagy. In traditional Chinese medicine, Paeonia lactiflora Pall is a common herb as a possible treatment for depression. As a main and active constituent of P. lactiflora Pall, paeoniflorin (PF)'s mechanisms of antidepression effects are not evidently unstated. Therefore, this study intended to explore whether PF can prevent depression-like behavior by conquering NLRP3 inflammasome activation and whether PF prevents NLRP3 inflammasome activation via upregulating mitophagy. Materials and Methods: Ten of a total of 50 rats were selected randomly as the control group. After establishment of the chronic unpredictable mild stress (CUMS) model, CUMS rats were randomly divided into four groups: CUMS group, fluoxetine hydrochloride group, PF group, and PF + cyclosporine A group. After 3 weeks of drug involvement, behavioral tests were measured. The protein expressions of PINK-1, Parkin, Beclin-1, LC3B, NLRP3, apoptosis-associated speck-like protein containing CARD (ASC), caspase-1 p20, interleukin-1β (IL-1β), and IL-18 were spotted with Western Blot method. Results: PF upturned stress persuaded behavioral changes and PF treatment augmented sucrose consumption rates (P < 0.01) in sucrose preference test and reduced the immobility time (P < 0.01) in forced swimming test of CUMS rats. PF also improved the levels of mitophagy-related proteins PINK-1, Parkin, Beclin-1, and LC3B (P < 0.01) in the hippocampus. Moreover, PF decreased the levels of NLRP3 inflammasome-related proteins (NLRP3, ASC, caspase-1 p20 antibody, IL-1β, and IL-18 [P < 0.01]) tempted by stress. Conclusion: PF advances depression in CUMS rats, reduced the inflammatory injury in the hippocampus of CUMS rats. Hence, based on those facts, NLRP3 inflammasome activation is accomplished by inhibiting its effect on mitophagy.