Rationale <p>Chronic unpredictable mild stress (CUMS) is a validated model of anxiety- and depression-like disorders and is associated with impaired neuroplasticity. Dysregulation of the plasminogen activator system, particularly elevated plasminogen activator inhibitor-1 (PAI-1), may disrupt tissue plasminogen activator (tPA)-dependent brain-derived neurotrophic factor (BDNF) signaling and contribute to stress-induced behavioral deficits. However, the therapeutic relevance of PAI-1 inhibition in mood disorders remains poorly defined.</p> Aim <p>This study examined whether pharmacological inhibition of PAI-1 using TM5441 could alleviate CUMS-induced anxiety- and depression-like behaviors by modulating the hippocampal PAI-1/tPA/BDNF axis.</p> Methods <p>Male and female C57BL/6 mice were exposed to CUMS for 3 weeks. TM5441, a selective SERPINE1 (PAI-1) inhibitor, was administered intraperitoneally (5&#xa0;mg/kg) 20&#xa0;min before behavioral testing, based on established efficacy and safety. Control mice received the vehicle (DMSO diluted in saline). Anxiety- and depression-like behaviors were assessed using the elevated plus maze, open field, sucrose preference, tail suspension, and forced swim tests. Hippocampal PAI-1, tPA, and BDNF levels were quantified, and correlations with behavioral outcomes were analyzed.</p> Results <p>TM5441 reversed CUMS-induced anxiety-like behaviors and depression-like responses. At the molecular level, TM5441 normalized stress-induced elevations in hippocampal PAI-1, restored tPA expression, and increased BDNF levels. Significant correlations were observed between PAI-1, tPA, and BDNF, as well as between hippocampal molecular markers and behavioral measures.</p> Conclusion <p>PAI-1 inhibition by TM5441 alleviates stress-induced behavioral deficits by restoring hippocampal tPA/BDNF signaling, highlighting the PAI-1/tPA axis as a promising therapeutic target for stress-related mood disorders.</p>