The interaction between the cytosolic pyridine nucleotide redox potential and gluconeogenesis from lactate/pyruvate in isolated rat hepatocytes. Implications for investigations of hormone action.
By using very low concentrations of cells to minimize alterations in substrate concentrations, we demonstrated that the lactate/pyruvate ratio of the incubation medium, which determines the cytosolic NADH/NAD+ ratio, affects gluconeogenic flux in suspensions of isolated hepatocytes from fasted rats. At a fixed extracellular pyruvate concentration of 1 mM and with the lactate/pyruvate ratio varied from 0.6 to 10 and to 50, glucose production rates increased from 2.5 to 5.5 and then decreased to 1.8 nmol/mg of cell protein/min. This finding paralleled the observation of Sugano et al. (Sugano, T., Shiota, M., Tanaka, T., Miyamae, Y., Shimada, M., and Oshino, N. (1980) J. Biochem. (Tokyo) 87, 153-166) who noted a similar biphasic response in the perfused liver system when lactate was held constant and pyruvate varied. The biphasic relationship can be explained by the influence of the NADH/NAD+ ratio on the near-equilibrium reactions catalyzed by glyceraldehyde-3-phosphate dehydrogenase and malate dehydrogenase in the hepatocyte cytosol. By shifting the equilibrium of the glyceraldehyde-3-phosphate dehydrogenase reaction, a rise in the NADH/NAD+ ratio decreases the concentration of 3-phosphoglycerate which, because of the linkage of 3-phosphoglycerate to phosphoenolpyruvate through two near-equilibrium reactions, reduces the concentration of phosphoenolpyruvate and therefore causes a decline in flux through pyruvate kinase. This decrease in pyruvate kinase flux results in an enhanced gluconeogenic flux. At higher NADH/NAD+ ratios, however, the oxalacetate concentration drops to such an extent that the consequent decreased flux through phosphoenolpyruvate carboxykinase exceeds the decline in flux through pyruvate kinase, producing a decrease in gluconeogenic flux. The lactate/pyruvate ratio was found to influence the actions of three hormones thought to stimulate gluconeogenesis by different mechanisms. Except for an inhibition by glucagon seen at the lowest lactate/pyruvate ratio tested, the stimulations by this hormone were relatively insensitive to lactate/pyruvate ratios, while angiotensin II produced greater stimulations of gluconeogenesis as the lactate/pyruvate ratio was increased. Dexamethasone, added in vitro, stimulated gluconeogenesis significantly only at very low and very high lactate/pyruvate ratios.
D000804:Angiotensin II / Q000494:pharmacology*; D000818:Animals; D003600:Cytosol / Q000378:metabolism; D003907:Dexamethasone / Q000494:pharmacology*; D005934:Glucagon / Q000494:pharmacology*; D005943:Gluconeogenesis / Q000187:drug effects; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases / Q000378:metabolism; D007700:Kinetics; D007773:Lactates / Q000378:metabolism; D019344:Lactic Acid; D008099:Liver / Q000378:metabolism*; D008291:Malate Dehydrogenase / Q000378:metabolism; D008297:Male; D009243:NAD / Q000378:metabolism*; D010071:Oxaloacetates / Q000378:metabolism; D010084:Oxidation-Reduction; D011770:Pyruvate Kinase / Q000378:metabolism; D011773:Pyruvates / Q000378:metabolism; D019289:Pyruvic Acid; D051381:Rats; D011919:Rats, Inbred Strains
D007773:Lactates; D010071:Oxaloacetates; D011773:Pyruvates; D009243:NAD; D000804:Angiotensin II; D019344:Lactic Acid; D003907:Dexamethasone; D019289:Pyruvic Acid; D005934:Glucagon; D008291:Malate Dehydrogenase; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases; D011770:Pyruvate Kinase