Studies on the Mechanism of Action of Glucagon on Gluconeogenesis

Abstract

Abstract The mechanism by which glucagon stimulates gluconeogenesis was studied in livers perfused with 1 mm lactate, 0.1 mm pyruvate, and several different 14C-labeled substrates. In livers perfused for 2½ min with [14C]lactate plus [14C]-pyruvate, glucagon increased [14C]glucose synthesis 3-fold, increased the labeling of tissue P-pyruvate, aspartate, alanine, and glycogen, and decreased the labeling of pyruvate, oxalacetate, malate, citrate, α-ketoglutarate, and glutamate. The hormone also decreased the tissue levels of pyruvate, malate, citrate, α-ketoglutarate, and glutamate and increased those of aspartate and P-pyruvate. These changes are interpreted as indicating that glucagon stimulated the conversion of oxalacetate to aspartate and P-pyruvate. Tryptophan produced changes in livers perfused with [14C]lactate plus [14C]pyruvate which were consistent with inhibition of gluconeogenesis at P-enolpyruvate carboxykinase. It blocked completely the effects of glucagon on the labeling of glucose and P-pyruvate at 2½ or 20 min, and caused the hormone to induce accumulation of malate and aspartate. The extra malate and aspartate which accumulated under these circumstances were derived from unlabeled sources (e.g. endogenous protein) and not from pyruvate as indicated by their decreased or unchanged radioactivity. In livers perfused with H14CO3 for 1 min, glucagon decreased the radioactivities of oxalacetate, malate, citrate, and α-ketoglutarate and increased those of aspartate, P-pyruvate, succinate, and fumarate. The specific radioactivities of intermediates were consistent with fixation of CO2 predominately into malate, aspartate, and citrate. The changes induced by glucagon indicated stimulation of aspartate and P-pyruvate formation from oxalacetate and of succinate formation from α-ketoglutarate. Further evidence for an effect of glucagon at a site between oxalacetate and P-pyruvate was provided by perfusions with [14C]aspartate. These showed that glucagon decreased the radioactivities of aspartate, malate, oxalacetate, and citrate and increased the radioactivity of P-pyruvate at both 2½ and 8½ min, despite the fact that the specific radioactivities of P-pyruvate precursors were reduced. Although the interpretation of these data is subject to reservation because of the existence of multiple pools of metabolic intermediates, they provide consistent support for the view that P-pyruvate synthesis from oxalacetate is a major site of action of glucagon on gluconeogenesis in the liver.