The effects of glucose, acetate, lactate and insulin on protein degradation in the perfused rat heart.
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ABSTRACT:
SUBMITTER: Sugden PH
PROVIDER: S-EPMC1158612 | biostudies-other | 1982 Sep
REPOSITORIES: biostudies-other
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Project description:Compared with glucose, lactate + acetate stimulated ventricular protein synthesis in anterogradely perfused hearts from fed or 72 h-starved rats. Stimulation was greater on a percentage basis in starved rats. Atrial protein synthesis was not detectably stimulated by lactate + acetate. Insulin stimulated protein synthesis in atria and ventricles. The stimulation of protein synthesis by lactate + acetate and insulin was not additive, the percentage stimulation by insulin being less in the ventricles of lactate + acetate-perfused hearts than in glucose-perfused hearts. Perfusion of hearts from 72 h-starved or alloxan-diabetic rats with glucose + lactate + acetate + insulin did not increase protein-synthesis rates or efficiencies (protein synthesis expressed relative to total RNA) to values for fed rats, implying there is a decrease in translational activity in these hearts. In the perfused heart, inhibition of protein synthesis by starvation and its reversal by re-feeding followed a relatively prolonged time course. Synthesis was still decreasing after 3 days of starvation and did not return to normal until after 2 days of re-feeding.
| S-EPMC1146874 | biostudies-other
Project description:The effects of insulin on glucose uptake and lactate release in the perfused working rat heart have been investigated in three types of preparation: (i) a control low-workload preparation; (ii) an increased-pressure-workload preparation, simulating conditions of aortic pressure encountered in vivo; (iii) an increased-volume-workload preparation, where pumping work done is approximately the same as (ii) but coronary flow is restricted because of the decreased aortic pressure. Insulin stimulated glucose uptake and lactate release in preparations (i) and (ii), but failed to do so in preparation (iii). It was considered possible that preparation (iii) was hypoxic, thus necessitating a maximal stimulation of glucose uptake. This was confirmed by improving cardiac oxygenation by addition of stroma-free haemoglobin to the perfusate in preparation (iii). Under these conditions in the absence of insulin, glucose uptake and lactate release were decreased compared with perfusions in the absence of haemoglobin. Insulin stimulation of both processes was restored. We conclude that the failure of other workers to observe insulin effects on glucose uptake and lactate release under physiological workloads [preparation (ii)] may be a consequence of intracellular hypoxia in their preparations.
| S-EPMC1158613 | biostudies-other
Project description:Increasing the extracellular pH over the range pH 7.4-8.9 stimulated protein synthesis by about 60% in the rat heart preparation anterogradely perfused in vitro. Protein degradation was inhibited by this pH increase. The magnitudes of the effects at pH 8.9 on protein synthesis and degradation were similar to those of high concentrations of insulin. Cardiac outputs were increased, as were cardiac phosphocreatine contents, indicating that the alterations in extracellular pH did not adversely affect the physiological viability of the preparation. ATP contents were unaltered. The creatine kinase equilibrium was used to assess the magnitude of the change in intracellular pH induced by these treatments. The increase in intracellular pH was about 0.2 for a 1-unit increase in extracellular pH. Thus small changes in intracellular pH have dramatic effects on cardiac protein turnover.
| S-EPMC1138488 | biostudies-other
Project description:The effect of insulin on glucose transport and glucose transporters was studied in perfused rat heart. Glucose transport was measured by the efflux of labelled 3-O-methylglucose from hearts preloaded with this hexose. Insulin stimulated 3-O-methylglucose transport by: (a) doubling the maximal velocity (Vmax); (b) decreasing the Kd from 6.9 to 2.7 mM; (c) increasing the Hill coefficient toward 3-O-methylglucose from 1.9 to 3.1; (d) increasing the efficiency of the transport process (k constant). Glucose transporters in enriched plasma and microsomal membranes from heart were quantified by the [3H]cytochalasin-B-binding assay. When added to normal hearts, insulin produced the following changes in the glucose transporters: (a) it increased the translocation of transporters from an intracellular pool to the plasma membranes; (b) it increased (from 1.6 to 2.7) the Hill coefficient of the transporters translocated into the plasma membranes toward cytochalasin B, suggesting the existence of a positive co-operativity among the transporters appearing in these membranes; (c) it increased the affinity of the transporters (and hence, possibly, of glucose) for cytochalasin B. The data provide evidence that the stimulatory effect of insulin on glucose transport may be due not to the sole translocation of intracellular glucose transporters to the plasma membrane, but to changes in the functional properties thereof.
| S-EPMC1148844 | biostudies-other
Project description:1. Protein degradation was studied in the glucose (5 mM)-perfused working rat heart preparation of Taegtmeyer, Hems & Krebs [(1980) Biochem. J. 186. 701-711]. 2. The effects of cardiac workload were investigated in three different preparations: (a) control (low workload), (b) increased pressure workload (simulating conditions of aortic pressure in vivo) and (c) increased volume workload. There was no effect of increased workload on protein degradation in preparation (b) or (c) when compared with preparation (a). Insulin inhibited protein degradation in all three preparations. Significantly greater inhibition by insulin was observed in the increased-pressure-workload preparation (b). 3. Hypoxia was induced by the partial replacement of O2 in the gaseous phase by N2. Hearts maintained their cardiac output when O2 content was decreased from 95% to 55% by volume, but the stability of the preparation was less at 50% O2. Lactate output was significantly increased at O2 contents of 65% or less. The rate of protein degradation was not different from control values (95% O2) in perfusions with 65, 55 or 50% O2. 4. We conclude that acutely increased workload or acute hypoxia does not affect protein degradation in the perfused working rat heart when cardiac output is relatively stable.
| S-EPMC1154189 | biostudies-other
Project description:We investigated the isolated working rat heart as a model to study early transcriptional remodeling induced by cardiac surgery with cardiopulmonary bypass (CPB). Because the mechanisms linking stress hyperglycemia and insulin resistance to increased peri-operative morbidity and mortality in the setting of open-heart surgery are still unclear, we also used this model to determine the transcriptional effects of exogenous glucose supply in absence or in presence of a preexisting state of myocardial insulin resistance. Male Sprague Dawley rats (200-224g) were kept on chow diet for 8 to 10 weeks, or fed a high-sucrose diet to induce myocardial insulin resistance. Following the feeding period, hearts were recovered, arrested in ice-cold Krebs-Henseleit (KH) buffer and subjected to 60 minutes normothermic reperfusion in the working mode with KH buffer supplemented with non-carbohydrate substrates plus glucose (25 mM) or mannitol (25 mM; osmotic control). Compared to non-perfused hearts, perfused hearts from chow-fed rats displayed a more than twofold increased expression for 71 genes (mannitol group) and 103 genes (glucose group) connected to inflammation, cell proliferation, and apoptosis. The same biological pathways were activated in hearts from insulin resistant rats, with 109 genes (mannitol group) and 70 genes (glucose group) up-regulated more than twofold when compared to the non-perfused hearts from high-sucrose diet-fed rats. The changes were highly similar to gene alterations occurring in the right atrium and left ventricle of open heart surgery patients, and included the up-regulation of the three master regulators of metabolic reprogramming MYC, NR4A1 and NR4A2. Targeted pathway analysis revealed an up-regulation of metabolic processes associated with the proliferation and activation of tissue-resident macrophages and fibroblasts. Although the remodeling occurred independently from glucose, glucose further increased the expression of a subset of genes associated with polarization of tissue reparative M2 macrophages. However, glucose failed to enhance the expression of M2-related markers in the isolated heart from rats rendered insulin resistant by high-sucrose feeding. These results expose the cellular components of the heart as a significant source of proinflammatory mediators released in response to stress associated with cardiac surgery with CPB, and suggest a major role for glucose as a signal in the determination of resident cardiac macrophage polarization.
2016-08-19 | GSE85811 | GEO
Project description:1. The metabolic pattern of [U-(14)C]glucose in the isolated rat heart has been studied, with both retrograde aortic (Langendorff) and atrially (working) perfused preparations in the presence and absence of insulin, in normal animals, animals rendered insulin-deficient (by injection of anti-insulin serum 1hr. before excision of the heart) and animals rendered diabetic by streptozotocin injection 7 days before use. 2. Radioautochromatograms of heart extracts show that the pattern of glucose metabolism in heart muscle is more complex than in diaphragm muscle. In addition to (14)CO(2), glycogen, oligosaccharides, phosphorylated sugars and lactate (the main metabolites formed from [(14)C]glucose in diaphragm muscle), (14)C label from [(14)C]glucose appears in heart muscle in glutamate, glutamine, aspartate and alanine, and in tricarboxylic acid-cycle intermediates. 3. By a quantitative scanning technique of two-dimensional chromatograms it was found that a mechanical work load stimulates glucose metabolism, increasing by a factor of 2-3 incorporation of (14)C into all the metabolites mentioned above except lactate and phosphorylated sugars, into which (14)C incorporation is in fact diminished; (14)CO(2) production is equally stimulated. 4. Addition of insulin to the perfusion fluid of the working heart causes increases in (14)C incorporation, by a factor of about 1.5 into (14)CO(2), by a factor of about 3-5 into glycogen, lactate and phosphorylated sugars, by a factor of about 2-3 into glutamate and tricarboxylic acid-cycle intermediates and by a factor of about 0.5 into aspartate, whereas incorporation into alanine and glutamine is not affected. The effect of a work load on the pattern of glucose metabolism is thus different from that of insulin. 5. Increasing the concentration of glucose in the perfusion fluid from 1 to 20mm leads to changes of the pattern of glucose metabolism different from that brought about by insulin. (14)CO(2) production steadily increases whereas [(14)C]lactate and glycogen production levels off at 10mm-glucose, at values well below those reached in the presence of insulin. 6. In Langendorff hearts of animals rendered insulin-deficient by anti-insulin serum or streptozotocin, glucose uptake, formation of (14)CO(2) and [(14)C]lactate, and (14)C incorporation into glycogen and oligosaccharides are decreased. In insulin-deficient working hearts, however, glucose uptake and (14)CO(2) production are normal, whereas incorporation of (14)C into glycogen and [(14)C]lactate production are greatly decreased. 7. Insulin added to the perfusion fluid restores (14)C incorporation from glucose into (14)CO(2), glycogen and lactate in the Langendorff heart from animals rendered insulin-deficient by anti-insulin serum; in hearts from streptozotocin-diabetic animals addition of insulin restores (14)C incorporation into glycogen and lactate, but (14)CO(2) production remains about 50% below normal. 8. The bearing of these results on the problem of the mode of action of insulin is discussed.
| S-EPMC1185135 | biostudies-other
Project description:A modified anterogradely perfused rat heart preparation is described in which all the cardiac output passes through the coronary circulation. Such a preparation develops hypertensive aortic pressures. Hypertensive aortic pressures or insulin stimulate the rate of cardiac protein synthesis and inhibit the rate of protein degradation. Aortic pressure and insulin may be important in the regulation of cardiac nitrogen balance in vivo. By abolishing cardiac prostaglandin synthesis with 4-biphenylacetate, we were able to investigate the possible involvement of prostaglandins in the modulation of protein turnover by pressure overload or insulin. There was no evidence of any involvement. However, insulin stimulated and cycloheximide inhibited cardiac prostaglandin synthesis. These findings are consonant with an enzyme involved in prostaglandin synthesis being short-lived and prostaglandin synthesis being rapidly influenced by activators and inhibitors of protein synthesis and degradation.
| S-EPMC1147879 | biostudies-other
Project description:An isotope-dilution method is described for the measurement of N tau-methylhistidine release from the perfused rat heart. We argue that release of N tau-methylhistidine is indicative of cardiac actin degradation. N tau-Methylhistidine release is compared with phenylalanine release in the presence of cycloheximide (phenylalanine release being a measure of degradation of mixed proteins). In hearts perfused with glucose plus acetate, the rate of actin degradation was increased by starvation and was not inhibited by insulin. In contrast, the rate of mixed-protein degradation was decreased by starvation and was inhibited by insulin. The fractional rate of degradation of mixed proteins in hearts from fed or starved rats was greater than that for actin. It is suggested that there are at least two pools of intracellular protein, the degradation rates of which differ in terms of their response to insulin and starvation.
| S-EPMC1146998 | biostudies-other
Project description:1. The extractions of glucose, lactate, pyruvate and free fatty acids by dog heart in vivo were calculated from measurements of their arterial and coronary sinus blood concentration. Elevation of plasma free fatty acid concentrations by infusion of intralipid and heparin resulted in increased extraction of free fatty acids and diminished extractions of glucose, lactate and pyruvate by the heart. It is suggested that metabolism of free fatty acids by the heart in vivo, as in vitro, may impair utilization of these substrates. These effects of elevated plasma free fatty acid concentrations on extractions by the heart in vivo were reversed by injection of dichloroacetate, which also improved extraction of lactate and pyruvate by the heart in vivo in alloxan diabetes. 2. Sodium dichloroacetate increased glucose oxidation and pyruvate oxidation in hearts from fed normal or alloxan-diabetic rats perfused with glucose and insulin. Dichloroacetate inhibited oxidation of acetate and 3-hydroxybutyrate and partially reversed inhibitory effects of these substrates on the oxidation of glucose. In rat diaphragm muscle dichloroacetate inhibited oxidation of acetate, 3-hydroxybutyrate and palmitate and increased glucose oxidation and pyruvate oxidation in diaphragms from alloxan-diabetic rats. Dichloroacetate increased the rate of glycolysis in hearts perfused with glucose, insulin and acetate and evidence is given that this results from a lowering of the citrate concentration within the cell, with a consequent activation of phosphofructokinase. 3. In hearts from normal rats perfused with glucose and insulin, dichloroacetate increased cell concentrations of acetyl-CoA, acetylcarnitine and glutamate and lowered those of aspartate and malate. In perfusions with glucose, insulin and acetate, dichloroacetate lowered the cell citrate concentration without lowering the acetyl-CoA or acetylcarnitine concentrations. Measurements of specific radioactivities of acetyl-CoA, acetylcarnitine and citrate in perfusions with [1-(14)C]acetate indicated that dichloroacetate lowered the specific radio-activity of these substrates in the perfused heart. Evidence is given that dichloroacetate may not be metabolized by the heart to dichloroacetyl-CoA or dichloroacetylcarnitine or citrate or CO(2). 4. We suggest that dichloroacetate may activate pyruvate dehydrogenase, thus increasing the oxidation of pyruvate to acetyl-CoA and acetylcarnitine and the conversion of acetyl-CoA into glutamate, with consumption of aspartate and malate. Possible mechanisms for the changes in cell citrate concentration and for inhibitory effects of dichloroacetate on the oxidation of acetate, 3-hydroxybutyrate and palmitate are discussed.
| S-EPMC1177916 | biostudies-other