Kinetics of active sodium transport in rat proximal tubules and its variation by cardiac glycosides at zero net volume and ion fluxes. Evidence for a multisite sodium transport system.

1. Transepithelial Na concentration difference, deltaCNa, across proximal tubules of rat kidney was measured at varying intraluminal Na concentrations (CNainfinity) under conditions of zero net volume and Na flux. Simultaneous stopped-flow intratubular and artificial peritubular capillary perfusion techniques were used together with intratubular raffinose to achieve zero net fluxes. Under these conditions in rat proximal tubules, deltaCNa represents active transport, JactNa, factored by permeability, PNa, plus an electrical factor depending on transepithelial potential difference. 2. The relationship between CNainfinity and deltaCNa appeared sigmoidal with saturation being reached when intratubular Na was above 80 m-mole/kg. In the presence of ouabain (10(-2)M) and scilliroside (10(-3)M) the relationship remained the same. The maximum deltaCNa was reduced by approximately 50% by cardiac glycoside inhibition whereas the half-saturation constant was essentially unchanged. These changes from the control represent simple non-competitive inhibition by the cardiac glycosides. 3. Absence of potential difference (p.d.) measurements precludes exact description of the relation between true active transport and substrate concentration but much evidence indicates that the apparently sigmoid relation in the presence and absence of cardiac glycoside inhibition, would be retained if correction of deltaCNa values were possible. Such results could then be explained if there are at least three or more sites for Na on the pump system, of which at least two are not cardiac glycoside sensitive. They would also unequivocally exclude the presence of a single-site single-pump system or the simple algebraic addition of two such units since the kinetic curves for both would be hyperbolic rather than sigmoidal.