Purified cardiac sarcolemmal membrane vesicles were used to determine if specific prostaglandin (PG) receptors are present on the myocyte. Two binding sites for PGE2 were identified in isolated bovine sarcolemmal membranesa high-affinity site with a dissociation constant (K4) of 032 nM and a maximum binding (Bmax of 376 fmol/mg of protein and a lower-affinity site with a K4 of 3.41 nM and a B max of 2,112 fmol/mg of protein. In competition experiments, unlabeled PGE1 displaced HPGE2 from its membrane receptor at concentrations similar to those of unlabeled PGE2. Both PGF2 μ and PGD2 displaced HPGE2 from the membrane, but only at high concentrations (> 10 M and > 10 M, respectively). Digestion of sarcolemmal membrane with trypsin resulted in a threefold decrease in specific HPGE2 binding. Phosphorylation of the membrane with protein kinase A also decreased specific HJPGE2 binding. At concentrations of PGE2} that occupy the high-affinity site, sarcolemmal adenylate cyclase activity was inhibited in the presence of 5ʼ-guanylylimidodiphosphate Gpp(NH)p. We conclude that the isolated cardiac sarcolemmal membrane contains a high-affinity binding site for PGEj that is functionally coupled to adenylate cyclase. The binding site is stereospecific and probably recognizes the 9-keto, ll-hydroxyl portion of the ring structure of these prostaglandins.