Hepatocellular transport of bile acids. Evidence for distinct subcellular localizations of electrogenic and ATP-dependent taurocholate transport in rat hepatocytes

To investigate whether electrogenic and ATP-dependent taurocholate transport activities are both mediated by the same bile acid-transporting polypeptide in rat liver, we further purified isolated canalicular membrane vesicles by free flow electrophoresis. Removal of most of the contaminating endoplasmic reticulum resulted in a complete loss of electrogenic taurocholate transport from an ecto-ATPase-enriched canalicular membrane subfraction. In contrast, ATP-dependent taurocholate transport remained associated with both an ecto-ATPase-enriched and an ecto-ATPase-free canalicular membrane subfraction. Microsomes containing 64% of total endoplasmic reticulum exhibited saturable electrogenic (Km approximately 270 microM), but no ATP-dependent taurocholate uptake. Golgi membrane vesicles were devoid of any taurocholate transport activity. These results indicate that electrogenic taurocholate transport resides entirely in the endoplasmic reticulum, whereas ATP-dependent bile acid transport is an intrinsic function of the canalicular membrane as well as of a so far unidentified intracellular membrane bound compartment. Hence, the two transport activities are most probably mediated by two different bile acid transporting polypeptides. Furthermore, the finding of ATP-dependent taurocholate transport in virtually ecto-ATPase-free vesicles argues against the concept of primary active bile acid transport being exclusively mediated by the canalicular ecto-ATPase.