STIMULATION OF PROTEIN-KINASE-C RAPIDLY REDUCES INTRACELLULAR NA+ CONCENTRATION VIA ACTIVATION OF THE NA+ PUMP IN OK CELLS

Na+ reabsorption is regulated in proximal tubules by hormones that sti mulate protein kinase C (PKC). To determine whether stimulation of PKC causes a reduction in intracellular Na+ concentration ([Na+](i)) that might link Na+ pump activation to increased Na+ reabsorption, [Na+](i ) was measured in kidney cells loaded with the Na+-sensitive fluoresce nt indicator SBFI. Rapid digital imaging fluorescence microscopy deter minations were performed in epithelial kidney cells transfected with t he rodent Na+ pump alpha 1 cDNA. In 42 determinations, the basal [Na+] (i) was 19.7 +/- 2.4 mM. Stimulation of PKC reduced the [Na+](i) to 5. 6 +/- 0.6 mM in similar to 10 sec. This drastic change in [Na+](i) req uires a transient 74-120-fold increase in Na+ pump activity. After the new steady state [Na+](i) is reached, the Na+ pump is 58% activated. The entry of Na+ into the cells is not affected by stimulation of PKC; therefore, the reduction in [Na+](i) is exclusively dependent on acti vation of the Na+ pump. Accordingly, PKC stimulation does not affect t he [Na+](i) of cells expressing a mutant Na+ pump that is not stimulat ed by PKC. The decrease in [Na+](i) observed in cells transfected with the rodent Na+ pump alpha 1 cDNA is large and sufficiently fast that it is expected to stimulate rapidly passive Na+-influx into the cells, thereby accounting for the observed PKC-induced stimulation of Na+ re absorption.