Serotonin (5-hydroxytryptamine; 5-HT) is known to be activated during ischemia–reperfusion and triggers contractile dysfunction and pathological apoptosis. Here, the beneficial effects of the selective serotonin reuptake inhibitor (SSRI) fluvoxamine was demonstrated on ischemia–reperfusion injury in guinea-pig hearts perfused using the Langendorff technique. The recovery (%) of left ventricular developed pressure (LVDP) by fluvoxamine (5×10⁻⁸ M ) was 95.4% (control: 32%), which was consistent with the inhibition of mitochondrial Ca²⁺([Ca²⁺]_m) uptake induced by changes in the Ca²⁺ content and acidification of the perfusate, and similar to reperfusion following global ischemia in Langendorff-perfused hearts. Fluvoxamine inhibited the increase in [Ca²⁺]_m induced by changes in the Ca²⁺ content of the perfusate in perfused preparations of mitochondria, which was similar to the results obtained with the mitochondrial permeability transition pore (MPTP) opener atractyroside. The terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick end labeling (TUNEL)-positive cells were significantly less in fluvoxamine-treated hearts than in control hearts, with decreases in caspase-3 activity. These results suggest that SSRI inhibits opening of the MPTP by preventing [Ca²⁺]_m overload-induced apoptosis related to the endogenous accumulation of 5-HT in ischemia–reperfusion hearts.