2,3,7,8-TETRACHLORODIBENZO-P-DIOXIN BLOCKS THE PHYSIOLOGICAL REGULATION OF HEPATIC PHOSPHOENOLPYRUVATE CARBOXYKINASE ACTIVITY IN PRIMARY RAT HEPATOCYTES

It has been previously reported that TCDD dose-dependently reduces the activity of PEPCK, the rate-limiting enzyme of hepatic gluconeogenesi s. To further investigate the mechanism, whereby TCDD decreases PEPCK activity, we studied the effect of TCDD on PEPCK activity in primary r at hepatocytes (PRH). PRH were isolated from male Sprague-Dawley rats by collagenase perfusion and incubated on collagen-coated culture dish es in medium M199 containing 1 nM insulin. Cells were pretreated with dexamethasone (100 nM) 8 h before PEPCK induction was initiated by add ition of glucagon (10 nM) and concurrent withdrawal of insulin. This h ormonal treatment induced the enzymatic activity of PEPCK in control c ells about 2-fold within 8 h. This PEPCK induction regimen was used to perform two sets of experiments. In the first set of experiments, rat s were pretreated with TCDD (125 mu g/kg p.o. in corn oil, 4 ml/kg) 4 days prior to isolation of PRH. This resulted in a complete block of t he glucagon-dependent induction of PEPCK in PRH from TCDD-pretreated a nimals. In the second set of experiments, TCDD (100 nM) was added dire ctly to the PRH either 24 or 48 h prior to the induction regimen. Incu bation of PRH with TCDD 24 h prior to initiation of the induction regi men resulted in a slight decrease in the degree of PEPCK induction whe n compared to controls. However, treatment of PRH with TCDD 48 h prior to initiation of the induction regimen almost completely blocked PEPC K induction. It is, therefore, suggested that the effect of TCDD on li ver PEPCK activity is due to a direct effect on liver cells and is not mediated by factors from outside the liver.