Regulation of prostaglandin EP(1) and EP(4) receptor signaling by carrier-mediated ligand reuptake

After synthesis and release from cells, prostaglandin E(2) (PGE(2)) undergoes reuptake by the prostaglandin transporter (PGT), followed by cytoplasmic oxidation. Although genetic inactivation of PGT in mice and humans results in distinctive phenotypes, and although experiments in localized environments show that manipulating PGT alters downstream cellular events, a direct mechanistic link between PGT activity and PGE(2) (EP) receptor activation has not been made. Toward this end, we created two reconstituted systems to examine the effect of PGT expression on PGE(2) signaling via two of its receptors (EP(1) and EP(4)). In human embryonic kidney cells engineered to express the EP(1) receptor, exogenous PGE(2) induced a dose-dependent increase in cytoplasmic Ca(2+). When PGT was expressed at the plasma membrane, the PGE(2) dose–response curve was right-shifted, consistent with reduction in cell surface PGE(2) availability; a potent PGT inhibitor acutely reversed this shift. When bradykinin was used to induce endogenous PGE(2) release, PGT expression similarly induced a reduction in Ca(2+) responses. In separate experiments using Madin–Darby Canine Kidney cells engineered to express the PGE(2) receptor EP(4), bradykinin again induced autocrine PGE(2) signaling, as judged by an abrupt increase in intracellular cAMP. As in the EP(1) experiments, expression of PGT at the plasma membrane caused a reduction in bradykinin-induced cAMP accumulation. Pharmacological concentrations of exogenous PGE(2) induced EP(4) receptor desensitization, an effect that was mitigated by PGT. Thus, at an autocrine/paracrine level, plasma membrane PGT regulates PGE(2) signaling by decreasing ligand availability at cell surface receptors.