STUDIES ON THE METABOLISM OF METALLOTHIONEIN AND ALKALINE-PHOSPHATASEOF ADULT-RAT PRIMARY HEPATOCYTE CULTURES - ROLE OF FETAL CALF SERUM AND AGONISTS OF THE PHOSPHOINOSITIDE CASCADE

Adult rat primary hepatocytes maintained in DMEM/F12 (Ham) media were used as a model system for studying the role of fetal calf serum (FCS) and agonists of the phosphoinositide cascade in the metabolism of met allothionein (MT) and alkaline phosphatase (ALP). Experiments were per formed both after a 24 h preincubation with FCS and with bovine serum albumin (BSA). Hepatocytes were treated with dexamethasone (DEX), zinc (Zn) and with the agonists of the phosphoinositide cascade A 23187, 1 ,2-dioctanoyl-sn-glycerol (DiC8), 12-O-tetradecanoylphorbol-13-acetate (TPA), angiotensin II (AT), platelet activating factor (PAF), Arg8-va sopressin (VP) and were analyzed for MT and ALP activity in cell homog enates. Cell viability was evaluated by lactate dehydrogenase (LDH) li beration into culture medium, induction of tyrosine aminotransferase ( TAT) through DEX and by trypan blue exclusion. Overall, cell viability was improved by the FCS pretreatment and by DEX. Exposure of hepatocy tes to the established direct inducers Zn and DEX of MT resulted in a manifold increase in MT, independent of whether the cultures were FCS pretreated or not. The FCS preincubation produced a moderate elevation of ALP activity by stimulating cell viability. However, ALP was unalt ered in response to Zn and DEX. None of the experiments conducted with agonists of the phosphoinositide cascade led to an elevation of MT an d ALP. Only the incubation of hepatocytes with A 23187 resulted in a c oncentration dependent significant decrease of MT and ALP. This observ ation was due to a cytotoxic effect of A 23187, displayed by LDH leaka ge and an increase in the number of cells stained with trypan blue.