Implications of the simultaneous occurrence of hepatic glycolysis from glucose and gluconeogenesis from glycerol.

Glycolysis from [6-(3)H]glucose and gluconeogenesis from [U-(14)C]glycerol were examined in isolated hepatocytes from fasted rats. A 5 mm bolus of glycerol inhibited phosphorylation of 40 mm glucose by 50% and glycolysis by more than 60%, and caused cellular ATP depletion and glycerol 3-phosphate accumulation. Gluconeogenesis from 5 mm glycerol was unaffected by the presence of 40 mm glucose. When nonsaturating concentrations of glycerol (< 200 microm) were maintained in the medium by infusion of glycerol, cellular ATP concentrations remained normal. The rate of uptake of infused glycerol was unaffected by 40 mm glucose, but carbohydrate synthesis from glycerol was inhibited 25%, a corresponding amount of glycerol being diverted to glycolytic products, whereas 10 mm glucose had no inhibitory effect on conversion of infused glycerol into carbohydrate. Glycerol infusion depressed glycolysis from 10 mm and 40 mm glucose by 15 and 25%, respectively; however, the overall rates of glycolysis were unchanged because of a concomitant increase in glycolysis from the infused glycerol. These studies show that exposure of hepatocytes to glucose and low quasi-steady-state concentrations of glycerol result in the simultaneous occurrence, at substantial rates, of glycolysis from glucose and gluconeogenesis from the added glycerol. We interpret our results as demonstrating that, in hepatocytes from normal rats, segments of the pathways of glycolysis from glucose and gluconeogenesis from glycerol are compartmentalized and that this segregation prevents substantial cross-over of phosphorylated intermediates from one pathway to the other. The competition between glucose and glycerol implies that glycolysis and phosphorylation of glycerol take place in the same cells, and that the occurrence of simultaneous glycolysis and gluconeogenesis may indicate channelling within the cytoplasm of individual hepatocytes.     

Compartmentation between glycolysis and gluconeogenesis in rat liver

1. The specific radioactivity-time relationships of glucose, glucose 6-phosphate, glycerol 1-phosphate and UDP-glucose were determined in rat liver after the intravenous injection of [U-(14)C]fructose, and a kinetic analysis was carried out. The glucose 6-phosphate pool was found to be compartmented into gluconeogenic and glycolytic components, and evidence was obtained that the triose phosphates were similarly compartmented. The glycolytic pathway was fed by glycogenolysis and glucose phosphorylation. There was no direct evidence that glycogenolysis fed only the glycolytic pathway, but this interpretation would make the liver resemble other organs in this respect. 2. UDP-glucose was not formed solely from gluconeogenic glucose 6-phosphate, as there was some dilution of label in the intervening glucose 1-phosphate pool, probably from glycogenolysis, though other pathways cannot be excluded. 3. The data cannot be explained by isotopic exchange.     

The control of glycolysis and gluconeogenesis by protein phosphorylation

Fructose 2,6-bisphosphate has been discovered as a potent stimulator of liver phosphofructokinase. It is also an inhibitor of fructose 1,6-biphosphatase and a stimulator of PPi: fructose 6-phosphate phosphotransferase from higher plants. It is formed from fructose 6-phosphate and ATP by a 6-phosphofructo 2-kinase and hydrolysed by a fructose 2,6-bisphosphatase. These two enzymes have very similar physicochemical properties and could not be separated from each other. They are substrates for cyclic-AMP-dependent protein kinase, which inactivates the first enzyme and activates the second.     

A reassessment of glycolysis and gluconeogenesis in higher plants

Sung, S.-J. S., Xu, D.-P., Galloway, C. M. and Black, C. C., Jr. 1988. A reassessment of glycolysis and gluconeogenesis in higher plants. - Physiol. Plant. 72: 650–654.
Sucrose is the starting point of glycolysis and end point of gluconeogenesis in higher plants. During both glycolysis and gluconeogenesis alternative enzymes are present at various steps to carry out parallel pathways; alternatives are available for utilizing nucleotide triphosphates and pyrophosphate; fructose 2,6-bisphosphate serves as a strong internal regulator; and plants use these cytoplasmic alternatives as they develop and as their environments change.     

Control of glycolysis and gluconeogenesis in rat kidney cortex slices

1. Glucose uptake or glucose formation has been studied in kidney cortex slices to investigate metabolic control of phosphofructokinase and fructose-diphosphatase activities. 2. Glucose uptake is increased and glucose formation is decreased by anoxia, cyanide or an uncoupling agent. Under these conditions the intracellular concentrations of glucose 6-phosphate and ATP decreased whereas that of fructose diphosphate either increased or remained constant, and the concentrations of AMP and ADP increased. 3. Glucose uptake was decreased, and glucose formation from glycerol or dihydroxyacetone was increased, by the presence of ketone bodies or fatty acids, or after starvation of the donor animal. Under these conditions, the concentrations of glucose 6-phosphate and citrate were increased, whereas those of fructose diphosphate and the adenine nucleotides were unchanged (see also Newsholme & Underwood, 1966). 4. It is concluded that anoxia and cell poisons increase glucose uptake and decrease gluconeogenesis by stimulating phosphofructokinase and inhibiting fructose diphosphatase, whereas ketone bodies, fatty acids or starvation increase gluconeogenesis and decrease glucose uptake through the citrate inhibition of phosphofructokinase.     

Adenosine phosphates and the control of glycolysis and gluconeogenesis in yeast

1. Changes in dry weight, protein, RNA and DNA were measured in yeast during adaptation to glycolytic metabolism. 2. Only RNA increased significantly during the lag phase, but during the exponential phase all these cellular components increased in parallel. 3. The concentrations of ATP, ADP, AMP and glucose 6-phosphate were measured in respiring yeast and during the transition to glycolytic metabolism. 4. In respiring cells the concentration of AMP was at its highest and that of ATP was at its lowest; this relationship was reversed in glycolysing cells. 5. ADP concentration was similar in respiring and glycolysing cells, but glucose 6-phosphate concentration was much higher in the glycolysing cells. 6. A possible reason for mitochondrial repression is suggested. 7. It is concluded that adenosine phosphates do not control the direction of glycolytic flux in yeast and an alternative control of glycolysis and gluconeogenesis by enzyme activation and inactivation is suggested.     

ATP versus pyrophosphate: glycolysis revisited in parasitic protists

It is generally assumed that glycolysis is remarkably similar among all organisms, prokaryotic and eukaryotic. This view has been extended to protists. However, there is growing evidence that significant deviations from conventional glycolysis occur in protists. Very different species, some parasitic, rely on peculiar enzymes that use pyrophosphate as substrate. In this review, Emmanuel Mertens describes such unusual pyrophosphate metabolism in parasitic protists.     

Regulation of hepatic glycolysis and gluconeogenesis by atrial natriuretic peptide.

Recently we reported the presence of both the guanylyl cyclase-linked (116 kDa) and the ANF-C (66 kDa) atrial natriuretic peptide receptors in the rat liver. Since ANF 103-125 (atriopeptin II) stimulates cGMP production in livers and because cGMP has previously been shown to mimic the actions of cAMP in regulating hepatic carbohydrate metabolism, studies were performed to investigate the effects of atriopeptin II on hepatic glycolysis and gluconeogenesis. Additionally, employing analogs of atrial natriuretic hormone [des-(Q116, S117, G118, L119, G120) ANF 102-121 (C-ANF) and des-(C105,121) ANF 104-126 (analog I)] which bind only the ANF-C receptors, the role of the ANF-C receptors in the hepatic actions of atriopeptin II was evaluated. In perfused livers of fed rats atriopeptin II, but not C-ANF and analog I, inhibited hepatic glycolysis and stimulated glucose production. Moreover, analog I did not alter the ability of atriopeptin II to inhibit hepatic glycolysis. Atriopeptin II, but not C-ANF and analog I, also stimulated cGMP production in perfused rat livers. Furthermore, while atriopeptin II inhibited the activity ratio of pyruvate kinase by 30%, C-ANF did not alter hepatic pyruvate kinase activity. Finally, in rat hepatocytes, atriopeptin II stimulated the synthesis of [14C]glucose from [2-14C]pyruvate by 50% and this effect of atriopeptin II was mimicked by the exogenously supplied cGMP analog, 8-bromo cGMP. Thus atriopeptin II increases hepatic gluconeogenesis and inhibits glycolysis, in part by inhibiting pyruvate kinase activity, and the effects of atriopeptin II are mediated via activation of guanylyl cyclase-linked ANF receptors which elevate cGMP production.     

Catabolic efficiency of aerobic glycolysis: The Warburg effect revisited

Background  

Cancer cells simultaneously exhibit glycolysis with lactate secretion and mitochondrial respiration even in the presence of oxygen, a phenomenon known as the Warburg effect. The maintenance of this mixed metabolic phenotype is seemingly counterintuitive given that aerobic glycolysis is far less efficient in terms of ATP yield per moles of glucose than mitochondrial respiration.     

Sensitivity of pathway rate to activities of substrate-cycle enzymes: application to gluconeogenesis and glycolysis

In a study of metabolic regulation, it is frequently useful to consider the degree to which an enzyme can influence the rate of its pathway. The most productive expression of rate-controlling influence is the fractional change in pathway rate per fractional change in enzyme activity (called control strength or sensitivity coefficient). We have developed a system for considering how a substrate-cycle enzyme's control strength depends on its flux and reaction order and on related features of other enzymes of its pathway. We have applied this system to the gluconeogenic pathway of rat liver and the glycolytic pathway of bovine sperm, where enough fluxes and reaction orders have been published to allow valid estimates of several control strengths. In normal fed animals where gluconeogenesis is slow and unidirectional substrate-to-product and product-to-substrate fluxes are comparable, all substrate-cycle limbs have very high and similar control strengths regardless of their flux rates and positions in the pathway. The activity of a step affects all substrate-cycle control strengths similarly as it affects unidirectional end-to-end fluxes relative to net rate. Control strengths of non-substrate-cycle enzymes are negligible compared to those of substrate cycles. In fasting animals, on the other hand, where unidirectional Pyr----Glc flux is much greater than Glc----Pyr flux, upstream enzymes (near Pyr) have a regulatory advantage over downstream enzymes (near Glc). In this circumstance, control strength of each substrate-cycle enzyme is inversely related to rate limitingness between its substrate and the pathway substrate. Because the Pyr/PEP cycle is significantly rate limiting, the control strength of the Pyr----PEP limb is much greater than that of pyruvate kinase and all downstream enzymes. In the glycolytic pathway of bovine sperm, strong product inhibition of hexokinase detracts greatly from its rate limitingness and control strength, which are very small despite its position at the beginning of the pathway and its large free energy. Because the glucose-transport-hexokinase segment is not rate limiting, phosphofructo 1-kinase has almost as much control strength as it would have as the first enzyme of the pathway, and because the F6P/FDP cycle is only moderately rate limiting, Fru-1,6-P2ase and enzymes further downstream have substantial control strengths. When glycolysis is accelerated by stimulation of phosphofructo 1-kinase, control strength shifts from phosphofructo-1-kinase and all downstream enzymes to the transporthesokinase segment.(ABSTRACT TRUNCATED AT 400 WORDS)     

Control of pyruvate kinase activity during glycolysis and gluconeogenesis in Propionibacterium shermanii

The concentrations of glycolytic intermediates and ATP and the activities of certain glycolytic and gluconeogenic enzymes were determined in Propionibacterium shermanii cultures grown on a fully defined medium with glucose, glycerol or lactate as energy source. On all three energy sources, enzyme activities were similar and pyruvate kinase was considerably more active than the gluconeogenic enzyme pyruvate, orthophosphate dikinase, indicating the need for regulation of pyruvate kinase activity. The intracellular concentration of glucose 6-phosphate, a specific activator of pyruvate kinase in this organism, changed markedly according to both the nature and the concentration of the growth substrate: the concentration (7-10 mM) during growth with excess glucose or glycerol was higher than that (1-2 mM) during growth with lactate or at growth-limiting concentrations of glycerol or glucose. Other glycolytic intermediates, apart from pyruvate, were present at concentrations below 2 mM. Glucose 6-phosphate overcame inhibition of pyruvate kinase activity by ATP and inorganic phosphate. With 1 mM-ATP and more than 10 mM inorganic phosphate, a change in glucose 6-phosphate concentration from 1-2 mM was sufficient to switch pyruvate kinase from a strongly inhibited to a fully active state. The results provide a plausible mechanism for the regulation of glycolysis and gluconeogenesis in P. shermanii.     

Influence of tamoxifen on gluconeogenesis and glycolysis in the perfused rat liver

The actions of tamoxifen, a selective estrogen receptor modulator used in chemotherapy and chemo-prevention of breast cancer, on glycolysis and gluconeogenesis were investigated in the isolated perfused rat liver. Tamoxifen inhibited gluconeogenesis from both lactate and fructose at very low concentrations (e.g., 5 μM). The opposite, i.e., stimulation, was found for glycolysis from both endogenous glycogen and fructose. Oxygen uptake was unaffected, inhibited or stimulated, depending on the conditions. Stimulation occurred in both microsomes and mitochondria. Tamoxifen did not affect the most important key-enzymes of gluconeogenesis, namely, phosphoenolpyruvate carboxykinase, pyruvate carboxylase, fructose 1,6-bisphosphatase and glucose 6-phosphatase. Confirming previous observations, however, tamoxifen inhibited very strongly NADH- and succinate-oxidase of freeze–thawing disrupted mitochondria. Tamoxifen promoted the release of both lactate dehydrogenase (mainly cytosolic) and fumarase (mainly mitochondrial) into the perfusate. Tamoxifen (200 μM) clearly diminished the ATP content and increased the ADP content of livers in the presence of lactate with a diminution of the ATP/ADP ratio from 1.67 to 0.79. The main causes for gluconeogenesis inhibition are probably: (a) inhibition of energy metabolism; (b) deviation of intermediates (malate and glucose 6-phosphate) for the production of NADPH required in hydroxylation and demethylation reactions; (c) deviation of glucosyl units toward glucuronidation reactions; (d) secondary inhibitory action of nitric oxide, whose production is stimulated by tamoxifen; (e) impairment of the cellular structure, especially the membrane structure. Stimulation of glycolysis is probably a compensatory phenomenon for the diminished mitochondrial ATP production. The multiple actions of tamoxifen at relatively low concentrations can represent a continuous burden to the overall hepatic functions during long treatment periods.     

Long-term effects of physiological oxygen concentrations on glycolysis and gluconeogenesis in hepatocyte cultures

Primary cultures of adult rat hepatocytes were kept for 46 h with either insulin ('insulin cells') or glucagon ('glucagon cells') as the dominant hormone under different oxygen concentrations with 13% (v/v) O2 mimicking arterial and 4% hepatovenous levels. Thereafter metabolic rates were measured for a 2 h period under the same ('overall long-term O2 effects') or a different ('short-term O2 effects') oxygen concentration. From the differences of the two effects the 'intrinsic long-term O2 effects' were derived. Glycolysis, as measured in 'insulin-cells', was stimulated by low O2 levels. It was about threefold faster in cells cultured and tested under 4% O2 as compared to cells cultured and tested under 13% O2, indicating the overall long-term effect. Glycolysis was about twofold faster in cells cultured and tested under 4% O2 as compared to cells cultured under 4% O2 but tested under 13% O2, demonstrating the short-term effect. Glycolysis was about 1.5-fold faster in cells cultured and tested under 4% O2 as compared to cells cultured under 13% O2 but tested under 4% O2, showing the intrinsic long-term effect. This difference was roughly parallel to the difference in levels of glucokinase and pyruvate kinase. Gluconeogenesis, as measured in 'glucagon cells', was stimulated by high O2 levels. Similar to glycolysis overall long-term, short-term and intrinsic long-term effects could be distinguished. The intrinsic long-term effects determined under 13% O2 corresponded to a 1.5-fold stimulation and paralleled the difference in phosphoenolpyruvate carboxykinase levels. The present results show that physiological oxygen concentrations also modulate hepatic carbohydrate metabolism by long-term effects and that the O2 gradient over the liver parenchyma thus contributes to the metabolic differences between periportal and perivenous hepatocytes in vivo.     

The selective control of glycolysis,gluconeogenesis and glycogenesis by temporal insulin patterns

Insulin governs systemic glucose metabolism, including glycolysis, gluconeogenesis and glycogenesis, through temporal change and absolute concentration. However, how insulin‐signalling pathway selectively regulates glycolysis, gluconeogenesis and glycogenesis remains to be elucidated. To address this issue, we experimentally measured metabolites in glucose metabolism in response to insulin. Step stimulation of insulin induced transient response of glycolysis and glycogenesis, and sustained response of gluconeogenesis and extracellular glucose concentration (GLC _ex ). Based on the experimental results, we constructed a simple computational model that characterises response of insulin‐signalling‐dependent glucose metabolism. The model revealed that the network motifs of glycolysis and glycogenesis pathways constitute a feedforward (FF) with substrate depletion and incoherent feedforward loop (iFFL), respectively, enabling glycolysis and glycogenesis responsive to temporal changes of insulin rather than its absolute concentration. In contrast, the network motifs of gluconeogenesis pathway constituted a FF inhibition, enabling gluconeogenesis responsive to absolute concentration of insulin regardless of its temporal patterns. GLC _ex was regulated by gluconeogenesis and glycolysis. These results demonstrate the selective control mechanism of glucose metabolism by temporal patterns of insulin.