Prostaglandin E2 (PGE2) is a potent lipid mediator in a diverse range of biological processes. This study examined the hypertrophic effect of PGE2 in primary cultured rat neonatal cardiomyocytes. PGE2 increased total protein synthesis in a dose-dependent manner, as measured by [3H]-phenylalanine uptake. PGE2 increased the cell size and surface area and induced the reorganization of myofilaments. Phosphorylation of the p42/44 and p38 mitogen-activated protein kinases (MAPK) was also induced by PGE2, and U0126 [a mitogen-activated extracellular signal regulated kinase kinase (MEK) 1/2 inhibitor] significantly inhibited the PGE2-induced protein synthesis. Expression of the hypertrophic marker genes, atrial natriuretic peptide and brain natriuretic peptide, was increased by PGE2, but expression of the alpha-skeletal actin gene was significantly attenuated. Transcripts for all 4 PGE2 receptor subtypes (EP1, EP2, EP3, and EP4) were detected in the cardiomyocytes. AE3-208 (an EP4-selective antagonist) significantly inhibited the alpha-skeletal actin gene suppression induced by PGE2, whereas SC51322 (an EP1-selective antagonist) did not. In conclusion, PGE2 induced hypertrophic changes in cardiomyocytes and attenuated alpha-skeletal actin gene expression in part via EP4.