Introduction: Stress induced by continuous swimming activates serotonergic activity in the brainstem and hippocampus. In several animal models of depression, exposure to stressful conditions results in observable behavioral changes, including a shift from proactive to reactive emotional responses, which are believed to reflect signs of depression. The primary aim of this experimental study was to evaluate serotonin levels in Wistar rats subjected to the forced swim test. Material and Methods: This experimental study spanned 22 days and involved twenty Wistar rats, divided into two groups of ten. The rats underwent the forced swim test for 21 days, swimming for 60 to 90 minutes each day. Serotonin levels were measured on days one, seven, fourteen, and twenty-one using an ELISA kit protocol. Results: The study found no discernible variation in serotonin levels in the control group (p > 0.20). After seven days of swimming, serotonin levels in the experimental group measured 0.095, which was lower than in the control group but not statistically significant. Following 14 days of swimming, the experimental rats showed no significant changes in serotonin production or release, with serotonin levels at 0.139—slightly higher than in the control group. After 21 days, serotonin levels increased to 0.249, higher than levels recorded on days seven and fourteen, yet still not significantly different from the control group (p > 0.20). Conclusion: These findings suggest that prolonged swimming does not significantly increase serotonin release or synthesis. Serotonin levels may instead be influenced by other factors, such as physiological adaptation, duration of exposure, behavioral immobility, or acute stress induced by the forced swim test.