Steven E. Mayer

Effect of Glucagon on Cyclic 3',5'-AMP, Phosphorylase Activity and Contractility of Heart Muscle of the Rat

The purpose of this investigation was to contrast the effect of glucagon and that of epinephrine on the concentration of cyclic adenosine 3′,5′-monophosphate (cyclic AMP), the activity of phosphorylase a and the contractile amplitude of isolated perfused rat hearts. The two drugs were about equally effective except that the maximal augmentation of contractility by epinephrine (5 × 10−9 moles) was twice that produced by an equivalent dose of glucagon with a fourfold greater increase in cyclic AMP concentration. Combination of large doses of the two drugs caused increases in the cyclic nucleotide considerably greater than those required for maximal phosphorylase activation or associated with a maximal inotropic response. The effects of glucagon also developed more slowly than those of epinephrine. An increase in cyclic AMP was not detectable until after phosphorylase a and contractile amplitude had increased. The beta-receptor-blocking agents dichloroisoproterenol and pronethalol did not block the biochemical responses to glucagon in doses which abolished the epinephrine-induced increases in cyclic AMP and phosphorylase a. These results, along with those obtained by other investigators, indicate that glucagon can elicit the same biochemical responses in intact heart as have been obtained with epinephrine, but by action at a different receptor site.

Regulation of the phosphorylase activating pathway in intact cardiac and skeletal muscle

Abstract Regulation of the phosphorylase activating pathway in striated muscles may take place at several sites. In heart there appear to be at least two receptor sites for adenyl cyclase activation: one for epinephrine, which was blocked by β-adrenergic blocking agents, and one sensitive to glucagon. The catalytic activity of phosphorylase kinase is probably dependent upon the availability of free intracellular calcium ion. After removal of calcium from the perfusion medium of rat hearts, epinephrine caused cyclic AMP formation, but not b to a transformation. Excess calcium produced b to a conversion in the absence of epinephrine. Small doses of this amine produced activation of kinase in intact dog hearts without b to a conversion again suggesting a requirement for an additional factor in the control of kinase activity. Regulation of phosphorylase activation by calcium is also suggested by the experiments of Drummond et al. (10) in which electrical stimulation of skeletal muscle produced rapid formation of phosphorylase a without either an increase in the concentration of cyclic AMP or activation of phosphorylase kinase. Thus the release of intracellular calcium ion by a drug, hormone, or consequent to the depolarization of the muscle fiber membrane, may provide a second mechanism of regulation of the activation of the phosphorylase system. The effectiveness of phosphorylase a in catalyzing glycogenolysis may be subject to further regulation in intact heart muscle. Maximal formation of phosphorylase a by epinephrine caused no net decrease in glycogen in intact dog heart as long as the concentration of high energy phosphate compounds and inorganic phosphate did not vary from controls. On the other hand, anoxia was a potent and rapid stimulus for glycogenolysis in the presence of rapid hydrolysis of high energy phosphates. Thus, under certain conditions glycogenolysis may be regulated by alterations in muscle metabolites. Intracellular compartmentalization may also play a role in the regulation of the system. Phosphorylation of the two phosphoproteins, phosphorylase kinase and phosphorylase a, appears to involve separate pools of muscle ATP. The experiments discussed in this paper suggest that the phosphorylase activating pathway can be stimulated and controlled at several points. This conclusion is in agreement with the experiments and hypotheses on the regulation of glycolysis in muscle presented by Helmreich and Cori in an earlier volume of this series of symposia (11).

Action of epinephrine on glucose uptake and glucose-6-phosphate in the dog heart in situ.

Abstract These experiments were designed to test the hypotheses: (1) that epinephrine inhibits glucose utilization as a result of the inhibition of phosphorylation by glucose-6-phosphate (glucose-6-P); (2) that the increase in the concentration of this ester is correlated with the positive inotropic action of the amine. The investigations were conducted on anesthetized dogs to which saline or epinephrine (1 or 10 μg/kg min −1 ) were infused for 15 min. The chloride and glucose space of the right ventricle and its content of glucose-6-P and the activity of glycogen phosphorylase were measured. In the control series 6% of the total phosphorylase was in the a form, the glucose space was 14% and glucose-6-P was 0.4 mmolc/kg. After 1 μg epinephrine, left ventricular-contractile force increased 76% and the enzyme was 19% in the a form, but the glucose space and glucose-6-P remained unchanged. After 10 μg of the drug, contractile force did not increase further and phosphorylase was 93% active, the glucose space rose to 67% and glucose-6-P concentration was 0.74 mmole/kg. Similar results were found in fed and fasted dogs and after parcreatectomy. These results indicate that inhibition of glucose utilization was associated with glucose-6-P accumulation, but this was not correlated with augmentation of contractile force of the heart.