Substantial new information regarding the function of the liver and other secretory epithelia suggests that current concepts of canalicular bile formation need revision. Recent observations suggest specifically that a) the NaK-ATPase-generated electrochemical sodium gradient which exists across liver plasma membranes (LPM) may provide the driving force for hepatic uptake (and transcellular transport) of bile salts and inorganic electrolytes via sodium-coupled membrane transport systems, b) that bile salts may exist in membrane-bound compartments within the liver cell, c) that the transport of some organic anions is equilibrative rather than concentrative, and d) that water and solutes may enter the canaliculus via paracellular pathways. This research proposal focuses on the characterization of LPM transport mechanisms for organic and inorganic solutes, the intracellular localization of organic solutes such as bile salts, and the relationship between the function of LPM transport systems and LPM enzymes such as NaK-ATPase and the composition and physical properties of LPM lipids. LPM lipid composition will be determined by standard methods and lipid fluidity will be assessed by fluorescence polarization, electron spin resonance, and differential scanning calorimetry. Hepatic transport will be studied using the isolated perfused rat liver, isolated hepatocytes, and vesicles prepared from LPM. Intracellular localization will be studied by autoradiography. These studies will broaden our understanding of bile formation and cholestasis and will provide valuable information reqarding more general questions such as the working of secretory epithelia and the relationship between physical and functional properties of cell surface membranes.