Properties of (Na + + K +)-activated ATPase in rat liver plasma membranes enriched with bile canaliculi

Abstract

Liver plasma membranes enriched in bile canaliculi were isolated from rat liver by a modification of the technique of Song et al. (J. Cell Biol. (1969) 41, 124–132) in order to study the possible role of ATPase in bile secretion. Optimum conditions for assaying (Na + + K +)-activated ATPase in this membrane fraction were defined using male rats averaging 220 g in weight. (Na + + K +)-activated ATPase activity was documented by demonstrating specific cation requirements for Na + and K +, while the divalent cation, Ca 2+, and the cardiac glycosides, ouabain and scillaren, were inhibitory. (Na + + K +)-activated ATPase activity averaged 10.07 ± 2.80 μmol P i/mg protei per h compared to 50.03 ± 11.41 for Mg 2+-activated ATPase and 58.66 ± 10.07 for 5′-nucleotidase. Concentrations of ouabain and scillaren which previously inhibited canalicular bile secretion in the isolated perfused rat liver produced complete inhibition of (Na + + K +)-activated ATPase without any effect on Mg 2+-activated ATPase. Both (Na + + K +)-activated ATPase and Mg 2+-activated ATPase demonstrated temperature dependence but differed in temperature optima. Temperature induced changes in specific activity of (Na + + K +)-activated ATPase directly paralleled previously demonstrated temperature optima for bile secretion. These studies indicate that (Na + + K +)-activated ATPase is present in fractions of rat liver plasma membranes that are highly enriched in bile canaliculi and provide a model for further study of the effects of various physiological and chemical modifiers of bile secretion and cholestasis.