The importance of arachidonic acid metabolites on the control of cell growth, particularly those derived from cyclooxygenase pathway has long been recognized. Recently, we observed that prostaglandin E 2 (PGE 2 ) interaction with EP 1 and EP 4 receptors is involved in serum-induced 3T6 fibroblast growth due to their effect at various levels of the cell cycle machinery. This study shows that prostanoid EP 3 receptor was expressed in 3T6 fibroblast. We studied the role of EP 3 receptor agonist GR 63799X in serum-induced 3T6 cell proliferation. This was concentration-dependent inhibit (IC 50 ∼10 μM) to a complete inhibition without any cytotoxic or proapoptotic effect. The prostanoid EP 3 receptor agonist treatment decreased the G 0 /G 1 and G 2 /M populations whereas cells were accumulated in S phase. This arrest in S phase was associated with a decrease in cyclin B levels and the enhancement of p21 expression. Our data show that EP 3 agonist decreases cAMP levels in our experimental conditions. Interestingly, the S arrest caused by prostanoid EP 3 receptor agonist seems to be cAMP dependent, at least in part, because forskolin treatment allowed S-arrested cells to progress through cell cycle and consequently growth. Thus, our results suggest that PGE 2 EP 3 receptor interaction may be involved in serum-induced 3T6 fibroblast growth due to their effects on cAMP levels and on cell cycle machinery of the S phase.