Multidrug resistance protein 4 mediates cAMP efflux from rat preglomerular vascular smooth muscle cells.

1. Previous studies have shown that stimulation of adenylyl cyclase in preglomerular vascular smooth muscle cells (PGVSMC) increases extracellular cAMP; however, the mechanism by which PGVSMC transport intracellular cAMP into the extracellular milieu is unknown. 2. We hypothesize that multidrug resistance protein (MRP) 4 is the primary transporter mediating efflux of intracellular cAMP from PGVSMC. 3. Both reverse transcription-polymerase chain reaction and real-time polymerase chain reaction detected MRP4 mRNA in PGVSMC in culture. Moreover, western blotting using an antibody specific for MRP4 gave rise to a 150 kDa signal, consistent with the presence of MRP4 protein in PGVSMC. 4. Specifically designed short interference (si) RNA reduced MRP4 mRNA expression by 71% (P = 0.0075) and MRP4 protein by 80% (P = 0.0004). 5. Isoproterenol (1 micromol/L) increased intracellular cAMP, which resulted in efflux of cAMP into the medium. The siRNA knockdown of MRP4 significantly reduced basal extracellular cAMP and nearly abolished isoproterenol-induced increases in extracellular cAMP (P = 0.0143, interaction between isoproterenol and MRP4 siRNA in two-factor analysis of variance). In isoproterenol-treated cells, MRP4 siRNA decreased the ratio of extracellular cAMP to intracellular cAMP by 72% (P = 0.0019). 6. We conclude that MRP4 is the dominant cAMP transporter in PGVSMC.