EFFECTS OF INTRACELLULAR SIGNALS ON NA+ K+-ATPASE PUMP ACTIVITY IN THE FROG-SKIN EPITHELIUM

The effects of intracellular signals (pH(i), Na(i)+, Ca(i)2+, and the electrical membrane potential), on Na+ transport mediated by the Na+/K+ pump were investigated in the isolated Rana esculenta frog skin. In particular we focussed on pH(i) sensitivity since protons act as an intrinsic regulator of transepithelial Na+ transport (J(Na)) by a simultaneous control of the apical membrane Na+ conductance (g(Na)) and the basolateral membrane K+ conductance (g(K)). pH(i) changes which modify J(Na), g(Na) and g(K), do not affect the Na+ transport mediated by the pump as shown by kinetic and electrophysiological studies. In addition, no changes were observed in the number of H-3-ouabain binding sites in acid-loaded epithelia. Our attempts to modify cellular Ca2+ (by using Ca2+-free/EGTA Ringer solution or A23187 addition) also failed to produce any significant effects in the Na+ pump turnover rate or the number of H-3-ouabain binding sites. The Na+ pump current was found to be sensitive to the basolateral membrane potential, saturating for very positive (cell) potentials and a reversal potential of - 160 mV was calculated from I-V relationships of the pump. Changes in Na(i)+ considerably affected the Na+ pump rate. A saturating relationship was found between pump rate and Na(i)+ with maximal activation at Na(i)+ > 40 mmol/l; a high dependence of the pump rate and of the number of H-3-ouabain binding sites was observed in the physiological range of Na(i)+. We conclude that protons (in the physiological pH range) which act directly and simultaneously on the passive transport pathways (g(Na) and g(K)), have no direct effect on the Na+/K+ pump rate. After an acid load, the inhibition of J(Na) is primarily due to the reduction of g(Na). This results in a reduction of Na(i) and the pump turnover rate then becomes dependent on other pathways of Na+ entry such as the basolateral membrane Na+/H+ exchanger.