Owen H. Filsell

Co-ordinated regulation of muscle glycolysis and hepatic glucose output in exercise by catecholamines acting via α-receptors

Recent findings indicate that glucose uptake by contracting hindlimb [Acta Physiol. Scand. (1982) 116, 215‐222] and heart [Biochem. Biophys. Res. Commun. (1982) 108, 124–131] of the rat is stimulated by epinephrine acting through α‐adrenergic mechanisms. Since in exercise hepatic glucose output may be increased markedly by activation of α‐adrenergic receptors and matched by the increase in muscle glucose uptake (maintaining blood glucose levels relatively constant), it is now proposed that a general coordination of glucose metabolism may operate via α‐adrenergic receptor mechanisms. The basis for this proposal is discussed.

An effect of diet on the activity of phosphofructokinase in rat heart

Summary High-carbohydrate diet and high-fat diet lead to an increase and a decrease, respectively in the activity of heart extract phosphofructokinase when determined at 10 μM-hexose 6-phosphate and 1 mM-ATP. Neither the maximal catalytic activity nor the total immunoprecipitable phosphofructokinase was significantly altered by these diets. Dialysis of the heart extracts from rats on either high-carbohydrate or high-fat diet decreased the activity of phosphofructokinase in each case but the proportional difference between the two activities remained essentially the same. It appears unlikely that the changes in activity result from changes in intracellular effector concentration that may be introduced by the diets.

Epinephrine-mediated stimulation of glucose uptake and lactate release by the perfused rat heart. Evidence for α- and β-adrenergic mechanisms

Abstract Epinephrine treatment of the perfused rat heart led to an increase in the rate of glucose uptake and lactate release as well as increases in the rate of beating and the activity ratio of phosphofructokinase. The dose of epinephrine required for half maximal increases in the rate of beating, and glucose uptake and the activity ratio of phosphofructokinase was approx.10−7M. Glucose uptake, lactate release and the activity ratio of phosphofructokinase were increased by the α-agonists methoxamine and phenylephrine, and the β agonist, isoproterenol. Propranolol and phenoxybenzamine each partially blocked the stimulatory effects of epinephrine on glucose uptake and lactate production. Phenoxybenzamine blocked the stimulatory effects of methoxamine but had no effect on those produced by isoproterenol which were blocked by propranolol. It is concluded that dual α and β adrenergic control of glycolysis occurs in cardiac muscle. It is proposed that the previously reported α-adrenergic control of phosphofructokinase plays a key role in the control of heart muscle glycolysis.

Epinephrine activation of phosphofructokinase in perfused rat heart: Regulation via α-adrenergic receptor mechanism independent of phosphorylation

Epinephrine treatment of the perfused rat heart leads to the activation of phosphofructokinase (PFK) via an a-adrenergic receptor mechanism, independent of changes in the intracellular concentration of cyclic AMP [ 1 ]. The activation is characterized by a loss in sensitivity to the inhibitors, ATP [ 1 ] and citrate [2], and is stable to gel-filtration [2,3]. Reconversion of the activated form to the non-activated form is catalyzed in heart extracts but does not appear to involve phosphoprotein phosphatase [2]. Here, the enzyme assay from [ 1,4] and SDS-polyacrylamide gel electrophoresis of immunoprecipitates of PFK were used to examine the relationship between c~-receptormediated activation and phosphorylation. 5 vol. 100 mM Tris-HC1 (pH 7.4) containing 1 mM dithiothreitol, 100 mM phosphate and 30 mM NaF. Supernatants (100 000 × g for 15 min) were mixed with an equal volume of reconstituted antiserum (100 mg/ml Tris-HCl-dithiothreitol) and allowed to stand at 4°C for 17 h. Immunoprecipitates (8000 × g for 10 min) were washed twice with isotonic saline and solubilized by incubation for 10 min at 95°C in 200/21 63 mM Tris-HC1 (pH 7.0), 1% SDS, 2.5% 2-mercaptoethanol, 5% glycerol and 0.02% bromophenol blue. Electrophoresis on 6% gels (10 cm) was done as in [8]. The specific radioactivity of [,),.32p]. ATP in heart powders was determined as in [9]. The activity ratio of PFK [ 1 ] was determined in hearts to which [32p] phosphate had not been added. Other experimental details were as in [ 1,3,4].

Obesity and the Regulation of Phosphofructokinase in Heart: An Apparent Insensitivity to Adrenergic Activation in Mature-Age Genetically Obese Rats

The activity ratio of phosphofructokinase in perfused rat heart and its activation by epinephrine was examined in non-obese, fat-fed obese, and genetically obese rats. For non-obese colony rats there was an age-dependent increase in the activity ratio of phosphofructokinase from 0.2 at 40 days to 0.4 at mature age (greater than 200 days). Epinephrine (10 microM) treatment of the heart for 5 min increased the ratio at all ages but the proportional increase diminished with age. For mature-age lean Zucker rats carrying the genetic determinant for obesity the results were similar to those obtained for comparable non-obese colony rats. For fat-fed obese rats the activity ratio of phosphofructokinase at 200 days of age was 0.2 and was increased to 0.6 by epinephrine treatment. For mature-age obese Zucker rats the activity ratio was 0.2 and no significant response to epinephrine occurred. The activity ratio of glycogen phosphorylase and its response to epinephrine (beta-adrenergic receptor mediated) in heart was unaffected by age, diet or the gene for obesity. The present findings indicate a specific defect in the adrenergic regulatory mechanism for phosphofructokinase in genetically obese rats.

Intracellular redox state and stimulation of gluconeogenesis and glycogenolysis in isolated hepatocytes from the pig (Sus scrofa)

Abstract 1. 1. Isolated pig hepatocytes were prepared and the effects of changes in the cytoplasmic [NADH]/[NAD + ] ratio on the efficacy of glucagon to alter rates of metabolism were examined. 2. 2. With hepatocytes from fed pigs 1 μM-glucagon stimulated glucose output. The response to glucagon was similar in magnitude regardless of whether 10 mM-lactate or 10 mM-pyruvate was present in incubations. 3. 3. With hepatocytes from 72-hr fasted pigs, glucagon (1 μM) increased the rate of gluconeogenesis from 10 mM-pyruvate but was without effect on the rate from 10 mM-lactate. These results differed from those obtained using rat hepatocytes where 1 μM-glucagon increased gluconeogenesis from 10 mM-lactate and inhibited gluconeogenesis from 10 mM-pyruvate. 4. 4. Intracellular concentrations of lactate and pyruvate were measured following 10 min incubations of pig hepatocytes with 10 mM-lactate or 10 mM-pyruvate. Comparisons with similar experiments conducted using rat hepatocytes indicated that both lactate and pyruvate entered the cells of both species and significantly altered the lactate/pyruvate ratio. 5. 5. Properties of the membrane-bound low K m cyclic AMP phosphodiesterase from pig and rat liver were compared. The activity of the enzyme in each species was similar and was inhibited to the same extent by NADH. 6. 6. The inability of pyruvate to inhibit the stimulatory effect of glucagon on glucose output and gluconeogenesis in pig hepatocytes does not appear to result from differences in the permeation of substrate into the cells or the sensitivity of cyclic AMP phosphodiesterase to altered cytoplasmic [NADH]/[NAD + ] ratio mediated by pyruvate or lactate addition.