CAMP EVOKES A RISE IN INTRACELLULAR NA+ MEDIATED BY NA+ PUMP INHIBITION IN RAT AORTIC SMOOTH-MUSCLE CELLS

The effect of adenosine 3',5'-cyclic monophosphate (cAMP) on intracellular Na+ concentration ([Na+](i)) was studied in primary cultured vascular smooth muscle cells from rat aorta [Na+](i) was measured using digital imaging of cells loaded with the Na+-sensitive fluorescent dye sodium-bonding benzofuran isophthalate. The cAMP level was raised by I) the membrane-permeable cAMP derivative 8-bromoadenosine 3',5'-cyclic monophosphate, 2) the combination of the adenylate cyclase activator forskolin and the phosphodiesterase inhibitor 3-isobutyl-1-methylxanthine, and 3) the beta-adrenoceptor agonist isoproterenol. All three methods caused a dose-dependent continuous rise in [Na+](i) during 40-60 min of observations. A rise in [Na+](i) may be caused by stimulation of the Naf influx and/or inhibition of Na+ efflux; therefore, the involvement of both mechanisms was studied. Elevation of the cAMP level had no effect on Na+ influx, measured as the rate of rise of [Na+](i) when Na+ efflux was inhibited with 1 mM ouabain. In contrast, elevation of the cAMP level attenuated Na+ efflux, measured as the rate of decline of [Na+](i) in Na+-loaded cells exposed to Na+-free medium. cAMP-induced inhibition of Na+ efflux was not observed when the Na+ pump was inhibited; therefore, cAMP inhibits the Na+ pump-mediated component of Na+ efflux. Agents that raise the cAMP level also inhibited, in a dose-dependent fashion, ouabain-sensitive Rb-86 uptake in rat aortic rings. The latter observation confirms that the cAMP-induced inhibition of the Na+ pump occurs both in cultured cells and in the native tissue. Changes in cellular cAMP levels evoked by the test compounds were measured and correlated with changes in [Na+](i) and Na+ pump activity: both the increase in [Na+](i) in cells and the inhibition of the Na+ pump in aortic rings showed a similar dependence on the net gain in cAMP. Taken together, the results suggest that, in rat aortic smooth muscle cells, elevation of the cAMP level inhibits the ouabain-sensitive Na+ pump and thus causes [Na+](i) to rise.