The apoptotic loss of cardiomyocytes contributes to numerous cardiovascular disorders. Evidence suggests that free fatty acids induce cellular apoptosis, and recent studies have shown that free fatty acids dramatically elevate mRNA levels of uncoupling protein 2 (UCP2) in some cell lines. In this study, we investigated the possibility that free fatty acids induce the expression of UCP2 through the peroxisome proliferator-activated receptor pathway, thereby increasing cell apoptosis in adult rat cardiomyocytes. Primary cultured adult rat cardiomyocytes exposed to free fatty acids exhibited a dose-dependent increase in apoptosis. Quantitative real-time reverse transcription-polymerase chain reaction and Western blotting showed significant increases in the level of UCP2 expression at 6, 12, and at 24 hours after treatment of adult rat cardiomyocytes with free fatty acids. Expression of UCP2 was suppressed with RNA interference, and knockdown of UCP2 attenuated free fatty acid-induced apoptosis in the cardiomyocytes. In summary, free fatty acids induced UCP2 expression through peroxisome proliferator-activated receptor alpha in adult rat cardiomyocytes.