To elucidate the first step in the vectorial transport of bile acids by the liver, plasma membrane vesicles were isolated from rat liver by differential and sucrose-Ficoll density gradient centrifugation. The membranes were selectively enriched 20-fold in Na',K'-ATPase activity, a marker of sinusoidal plasma membranes. Electron microscopy of pellets from sinusoidal membrane fraction did not reveal other organelles.
The initial rate of taurocholate uptake by the membrane vesicles was stimulated by a sodium gradient but not by a potassium gradient (Naouteide > Nainside). Sodium-dependent uptake of taurocholate was inhibited at low temperature (O"C), reduced by decreasing intravesicular space, and showed transstimulation in tracer exchange experiments indicating that uptake into vesicles, rather than binding to membranes, was occurring. Sodium-dependent taurocholate transport by the vesicles exhibited saturation kinetics with respect to taurocholate (apparent K, = 56 pM, V , , = 0.65 nmoles per mg X 15 sec at 100 mM NaN03 and 25Β°C) and sodium (apparent K, = 48 mM at 50 taurocholate). Other cations, such as lithium and choline, did not replace sodium in its stimulatory action. Sodium-dependent taurocholate uptake was selectively inhibited by cholic acid and probenecid. When the electrical potential difference across the vesicle membrane was altered by anion replacement, a more negative intravesicular potential inside the vesicles stimulated and a more positive potential inhibited sodium-dependent taurocholate transport. These data indicate the presence of a taurocholate-sodium cotransport system in the sinusoidal membranes of rat hepatocytes. This sodium-cotransport system probably participates in sodium-dependent uptake of bile acids into the hepatocyte. Bile acid secretion in the liver can be classified as secondary active transport.
Transport of organic anions, such as bile acids and bilirubin, is an important function of the liver. Based on kinetic analysis in intact animals (1-4), isolated perfused liver (5-9), and hepatocyte suspensions (10-14), carriermediated, sodium-dependent hepatic uptake of bile acids has been postulated. Vectorial transport of bile acids from plasma into bile is a sequence of many steps including uptake at the sinusoidal membrane, interaction with