FORMATION OF BILE CANALICULI IN LONG-TERM PRIMARY CULTURES OF ADULT-RAT HEPATOCYTES ON PERMEABLE MEMBRANE - AN ULTRASTRUCTURAL-STUDY

Adult rat hepatocytes were cultured for 15 days on type I collagen-coa ted permeable membranes in a hormonally defined Waxman's modified medi um supplemented with very low concentrations of insulin, glucagon and dexamethasone. Phase contrast examination showed that 15-day-old cultu res still formed a regular monolayer of polygonal cells. In similarly aged cultures, intracellular glycogen was abundant and evenly distribu ted, while steatosis remained very limited, Scanning and transmission electron microscopy showed that well developed bile canaliculi could b e observed on the lateral side of the hepatocyte membrane after 4 days of incubation and persisted for 2 weeks. These canalicular structures probably originated from coalescence of membrane invaginations observ ed in 1-day-old cultures. Transmission electron microscopy showed that the ultrastructure of the cells was very close to that of normal rat hepatocytes in the intact liver. These results suggest that rat hepato cytes cultured under these experimental conditions are able to develop and maintain tissue-specific cytochemical and morphological propertie s for at least 15 days.