Evidence for a Na+/K+/Cl- cotransport system in basolateral membrane vesicles from the rabbit parotid

Sodium (22Na) transport was studied in a basolateral membrane vesicle preparation from rabbit parotid. Sodium uptake was markedly dependent on the presence of both K+ and Cl- in the extravesicular medium, being reduced 5 times when K+ was replaced by a nonphysiologic cation and 10 times when Cl was replaced by a nonphysiologic anion. Sodium uptake was stimulated by gradients of either K+ or Cl (relative to nongradient conditions) and could be driven against a sodium concentration gradient by a KCl gradient. No effect of membrane potentials on KCl-dependent sodium flux could be detected, indicating that this is an electroneutral process. A KCl-dependent component of sodium flux could also be demonstrated under equilibrium exchange conditions, indicating a direct effect of K+ and Cl on the sodium transport pathway. KCl-dependent sodium uptake exhibited a hyperbolic dependence on sodium concentration consistent with the existence of a single-transport system with Km = 3.2 mM at 80 mM KCl and 23 degrees C. Furosemide inhibited this transporter with K0.5 = 2 X 10(4) M (23 degrees C). When sodium uptake was measured as a function of potassium and chloride concentrations a hyperbolic dependence on [K] (Hill coefficient = 0.87 +/- 0.09) and a sigmoidal dependence on [Cl] (Hill coefficient = 1.31 +/- 0.07) were observed, consistent with a Na/K/Cl stoichiometry of 1:1:2. Taken together these data provide strong evidence for the electroneutral coupling of sodium and KCl movements in this preparation and strongly support the hypothesis that a Na+/K+/Cl cotransport system thought to be associated with transepithelial chloride and water movements in many exocrine glands is present in the parotid acinar basolateral membrane.