The Regulation of Gluconeogenesis: THE EFFECT OF PENT-4-ENOIC ACID ON GLUCONEOGENESIS AND ON THE GLUCONEOGENIC METABOLITE CONCENTRATIONS OF ISOLATED PERFUSED RAT LIVER

Abstract

Abstract The mechanism of inhibition of gluconeogenesis by pent-4-enoic acid was studied in isolated perfused rat livers. One millimolar pent-4-enoic acid decreased gluconeogenesis from alanine by 80% and ketogenesis from linoleate by 95% and markedly increased the concentration of the gluconeogenic intermediates from lactate to 3-phosphoglycerate and decreased the concentrations of the intermediates from d-glyceraldehyde-P to glucose-6-P. This was associated with a marked decrease in the tissue lactate-pyruvate ratio and only a 20% decrease in the ATP concentration. These data suggest that gluconeogenesis was inhibited at glyceraldehyde phosphate dehydrogenase, probably as a result of decreased NADH generation from fatty acid oxidation. Another point of inhibition was present at pyruvate carboxylase, as indicated by the excessive elevation of the tissue pyruvate concentrations and by the marked decrease of the acetyl-CoA concentrations. The inhibition at pyruvate carboxylase was, however, not rate-limiting. When 8.0 mm ethanol was added after 10 min of prior perfusion with 1.0 mm pentenoic acid, ketone body production remained inhibited while gluconeogenesis from alanine was restored. It was thus possible to maintain relatively normal rates of gluconeogenesis from alanine in spite of almost complete inhibition of long chain fatty acid oxidation. Under these conditions the return toward normal of the lactate-pyruvate ratio showed that ethanol metabolism provided sufficient NADH for restoration of gluconeogenesis. When 8.0 mm ethanol was added after prior perfusion with 1.0 mm pentenoic acid for more than 10 min, the inhibition at glyceraldehyde phosphate dehydrogenase was still reversed, but gluconeogenesis remained inhibited because of inhibition at pyruvate carboxylase because of the decreased acetyl-CoA concentration. In experiments in which ethanol was added after a period of prior perfusion with pentenoic acid, the rate-limiting step for gluconeogenesis appeared to be at pyruvate carboxylase. Since varying rates of gluconeogenesis were obtained depending on the period of prior perfusion with pentenoic acid, it was possible to correlate the rate of gluconeogenesis with the tissue acetyl-CoA concentration. A sigmoidal relationship was obtained with an apparent activation constant for acetyl-CoA of 1.8 x 10-5 m, which agrees well with the value obtained for rat liver pyruvate carboxylase.