Intracellular traffic of asialoglycoproteins depends on vacuolar pH in polarized isolated rat hepatocyte couplets

Abstract Asialoglycoproteins are internalized via specific receptors localized on the hepatic parenchymal cells. The role of vacuolar type H + -ATPase (v-ATPase) and vacuolar pH in intracellular transport of asialoglycoproteins was investigated in isolated rat hepatocyte couplets. Two specific inhibitors of v-ATPase, bafilomycin A1 and concanamycin B, were used to inhibit acidification of the vacuolar system. Intracellular transport of asialoglycoproteins was investigated using gold-conjugated asialofetuin with electron microscopy. Fluorescent staining by acridine orange showed that bafilomycin A1 and concanamycin B each inhibited acidification of endocytic compartments. Electron microscopy demonstrated that the number of autophagic vacuoles was increased, and the uptake of gold-conjugated asialofetuin was significantly inhibited by these agents. Vesicle budding and membrane fusion between endocytic structures in the peripheral area and intracellular transport of endocytosed asialofetuin to lysosomes were inhibited by these agents. These results suggest that the intracellular transport of endocytosed asialoglycoproteins to lysosomes depends on acidification of the vacuolar system (vacuolar pH) in hepatocytes.