We previously described a method for regional myocardial cooling that reaches the target temperature within 4 min. The present study evaluated whether this method for regional myocardial cooling during reperfusion reduces myocardial infarct size (IS) in 75-kg pigs. Myocardial infarction was induced by inflation of an angioplasty balloon in the left anterior descendent artery for 45 min followed by 3 h reperfusion. First, 15 pigs were randomized to regional myocardial cooling during reperfusion (n = 8) or control (n = 7). As further control experiments, systemic hypothermia was induced prior to ischemia (n = 3) and during reperfusion (n = 3). IS and area at risk (AAR) were evaluated in vivo by single photon emission cardiac tomography (SPECT) and by standard histochemical staining. Regional cooling during reperfusion did not reduce IS/AAR as assessed by histochemistry (cooling: 0.71 +/- 0.8; control: 0.68 +/- 0.10; p = ns) and SPECT (cooling: 0.90 +/- 0.20; control: 0.88 +/- 0.32; p = ns). Systemic hypothermia during ischemia reduced IS/AAR (histochemistry: 0.09 +/- 0.11; SPECT: 0.25 +/- 0.22; p < 0.001 and p = 0.01 vs control, respectively). Induction of systemic hypothermia during reperfusion had no significant effect on IS/AAR (histochemistry: 0.63 +/- 0.07; SPECT: 0.74 +/- 0.09; p = ns vs control for both comparisons). In conclusion, hypothermia during ischemia is strongly myocardioprotective while hypothermia during reperfusion does not reduce myocardial infarct size in human-sized pigs.