Stimulation of ouabain-sensitive Rb-86(+) uptake and Na+,K+-ATPase alpha-subunit phosphorylation by a cAMP-dependent signalling pathway in intact cells from rat kidney cortex

We investigated in intact cortical kidney tubules the rot of PKA-mediated phosphorylation in the short-term control of Na+,K+-ATPase activity. The phosphorylation level of Na+,K+-ATPase was evaluated after immunoprecipitation of the enzyme from P-32-labelled cortical tubules and the cation transport activity of Na+,K+-ATPase was measured by ouabain-sensitive Rb-86(+) uptake. Incubation of cells with cAMP analogues (8-bromo-cAMP, dibutyryl-cAMP) or with forskolin plus 3-isobutyl-1-methylxanthine increased the phosphorylation level of the Na+,K+-ATPase alpha-subunit and stimulated ouabain-sensitive Rb-86(+) uptake. Inhibition of PKA by H-89 blocked the effects of dibutyryl-cAMP on both phosphorylation and Rb-86(+) uptake processes. The results suggest that phosphorylation by PKA stimulates the Na+,K+-ATPase activity.