Agonist-specific role for Na+-H+ antiport in prostaglandin release from microvessel endothelium

Abstract

Rabbit coronary microvascular endothelial (RCME) cells synthesize prostaglandin (PG) I2 and PGE2 in response to stimulation with human thrombin, ATP, and the Ca2+ ionophore, A23187. Replacement of extracellular Na+ with choline or N-methylglucamine reduced thrombin-stimulated PG secretion but did not significantly affect either ATP- or A23187-stimulated PG secretion. Pretreatment of RCME cells with Na+ channel or Na+ -Ca2+ exchange blockers did not alter PG release in response to any of these three agonists. Pretreatment of RCME cells with the specific Na+ -H+ antiport blockers 5-(N-ethyl-N-isopropyl)-amiloride (EIPA, 10 microM) and 5-(N,N-hexamethylene)-amiloride (HMA, 0.1 microM) significantly reduced thrombin but not A23187- or ATP-stimulated PG secretion. Studies with the intracellular pH indicator dye 2,7-bis(carboxyethyl)-5(6)-carboxyfluorescein demonstrated thrombin activation of Na+ -H+ antiport, an effect blocked by either HMA or EIPA. Since manipulations known to inhibit Na+ -H+ exchange (EIPA, HMA, replacement of Na+ with choline or N-methylglucamine) reduced thrombin-stimulated RCME cell PG release, we conclude that activation of Na+ -H+ exchange is involved in the coupling of thrombin interaction with RCME cells to subsequent phospholipase activation and PG release.