Effects of cortisol on the levels of metabolites and enzymes controlling glucose production from pyruvate

Abstract

Listen

Translate article

Starvation and cortisol treatment of rats result in an almost 2-fold increase in the glucogenic capacity of kidney cortex slices when supplemented with an excess of pyruvate or succinate. A rise of both pyruvate carboxylase and phosphoenolpyruvate (PEP) carboxykinase was considered to be the cause of the increased glucogenic capacity observed. This assumption could be supported by assays of pyruvate carboxylase and PEP carboxykinase under the influence of cortisol and after starvation: the activities of both enzymes were found to be elevated in the liver 6 hr after treatment of rats with a single dose of cortisol and in starved animals. The activities of both enzymes from kidney behaved in a similar fashion. ▪ Studies on the intracellular location of pyruvate carboxylase revealed that 30–50% of the total activity present in liver and kidney may be solubilized by isotonic solutions supplemented with sodium or potassium ions. Acetyl-CoA can substitute for these cations. Glutamic dehydrogenase, an enzyme exclusively located in the mitochondria, is solubilized to 3% only by these treatments. This behavior of pyruvate carboxylase favors an extramitochondrial location of part of this enzyme which is apparently bound to the external surface of the mitochondrion or other subcellular particulate fractions. Assays of the activities of the soluble and the mitochondrial pyruvate carboxylase after administration of cortisol support a regulation of gluconeogenesis by control of the extramitochondrial enzyme levels. The stimulation of gluconeogenesis by glucocorticoids independent of de novo synthesis of enzymes in liver was studied. Two possible mechanisms were investigated: activation of glucogenic enzymes, and inhibition of enzymes involved in glucose degradation by metabolites. Activation of pyruvate carboxylase, a key enzyme of gluconeogenesis, could be supported by assays of acetyl-CoA in livers of rats with a sensitive isotope assay. Within 3–6 hr after treatment of adrenalectomized rats with a single dose of cortisol, the levels of acetyl-CoA, an activator of pyruvate carboxylase, were significantly elevated when compared with untreated animals. A negative control of glycolysis under the influence of cortisol was supported by assays of glucose-6-phosphate in liver. Concomitant with acetyl-CoA, glucose-6-phosphate levels were also elevated. Inhibition of phosphofructokinase by citrate as the cause of the glucose-6-phosphate rise could be excluded. The mechanism involved has still to be elucidated. An additional negative control of glucose degradation under the influence of glucocorticoids could be supported by a study of the effects of various metabolites on the activity of a 100-fold purified pyruvate kinase from liver. Among 32 metabolites investigated, alanine proved to be a potent inhibitor of the enzyme from liver. The inhibitory action of alanine was evident in a concentration range such as has been found in liver. Amino acid mobilization under cortisol may thus also control the glucogenic capacity of liver. Glycogenesis, fixation of CO 2 into glycogen, and the rise of the various metabolites after cortisol treatment could be blocked by actinomycin D. This finding is in agreement with an enzyme induction in adipose tissue as the cause of the cortisol-dependent mobilization of fatty acids and the concomitant rise of the metabolites investigated. The dose of actinomycin necessary for the suppression of the latter effects of the hormone, however, was found to be twice (400 μg) the dose which completely blocks de novo synthesis of enzymes in liver (150–200 μg). In order to obtain experimental evidence for a primary action in the control of enzyme synthesis, the effect of glucocorticoids on CO 2-fixation and on the activities of pyruvate carboxylase was studied in vitro. Since kidney cortex slices, in contrast to liver slices, did not show leakage of pyruvate carboxylase activities on prolonged incubation, this tissue was chosen for studies in vitro. Incubation of kidney cortex slices from adrenalectomized rats with cortisol or dexamethasone ( 2 × 10 −5 m ) resulted in a significant increase in both the activity of pyruvate carboxylase and CO 2-fixation into glucose and into di- and tricarboxylic acids. Puromycin completely abolished this effect in vitro. Reproducibility of the effects in vitro depended on the presence of an amino acid mixture. This finding is in accord with a “permissive” role of glucocorticoids in the control of enzyme synthesis, as suggested by others.