Objective: α-Melanocyte-stimulating hormone (α-MSH), β-endorphin and other pro-opiomelanocortin- (POMC) derived peptides have been detected in the heart, but it is uncertain whether they are synthesized by cardiomyocytes or by cardiac nerves innervating the heart. The objective of this study was to determine whether POMC peptides are synthesized by cardiomyocytes. Methods: Pro-opiomelanocortin peptides were localized in rat heart by immunohistochemistry using antisera against α-MSH, β-endorphin and αN-acetyl-β-endorphin, the predominant POMC peptides found in heart. Pro-opiomelanocortin mRNA was investigated by reverse transcription polymerase chain reaction (RT-PCR) using primers that discriminate between full-length POMC mRNA and a 5′ truncated POMC transcript that is presumed to be non-functional. Results: α-Melanocyte-stimulating hormone, β-endorphin and αN-acetyl-β-endorphin immunoreactivities were localized in atrial myocytes, particularly in the atrial appendages, but not to a significant extent in ventricular myocytes. Cardiac nerves were not immunostained. Atrial natriuretic peptide (ANP) immunoreactivity was similarly distributed in the adult heart. In neonatal heart, POMC-peptide and ANP immunoreactivities were present in both atrial and ventricular myocytes. RT-PCR amplification showed that full-length POMC mRNA transcripts were present in both atrial and ventricular tissue and provide evidence that 5′ truncated POMC mRNA is expressed in heart. Conclusions: These results support the hypothesis that cardiomyocytes synthesize POMC peptides.