Recent studies indicate that members of the multidrug-resistance protein (MRP) family belonging to ATP binding cassette type C (ABCC) membrane proteins extrude cyclic nucleotides from various cell types. This study aimed to determine whether MRP proteins regulate cardiac cAMP homeostasis. Here, we demonstrate that MRP4 is the predominant isoform present at the plasma membrane of cardiacmyocytes and that it mediates the efflux of cAMP in these cells. MRP4-deficient mice displayed enhanced cardiac myocyte cAMP formation, contractility, and cardiac hypertrophy at 9 mo of age, an effect that was compensated transiently by increased phosphodiesterase expression at young age. These findings suggest that cAMP extrusion via MRP4 acts together with phosphodiesterases to control cAMP levels in cardiac myocytes.—Sassi, Y., Abi-Gerges, A., Fauconnier, J., Mougenot, N., Reiken, S., Haghighi, K., Kranias, EG, Marks, AR, Lacampagne, A., Engelhardt, S., Hatem, SN, Lompre, A.-M., Hulot, JS Regulation of cAMP homeostasis by the efflux protein MRP4 in cardiac myocytes.