scholarly journals Further evidence for the classical pentose phosphate cycle in the liver

1982 ◽  
Vol 208 (3) ◽  
pp. 851-855 ◽  
Author(s):  
Robert Rognstad ◽  
Pat Wals ◽  
Joseph Katz
Keyword(s):  
Specific Radioactivity ◽  
Rat Hepatocytes ◽  
Total Radioactivity ◽  
Pentose Phosphate ◽  
Classical Type ◽  
Isolated Rat Hepatocytes ◽  
Pentose Phosphate Cycle ◽  
Fructose 1,6 Bisphosphatase ◽  
Phenazine Methosulphate ◽  
Substrate Concentrations

Isolated rat hepatocytes were incubated with [3-14C]xylitol or d-[3-14C]xylulose plus xylitol or glucose at substrate concentrations. The glucose formed was isolated and degraded to give the relative specific radioactivities in each carbon atom. C-4 of glucose had the highest specific radioactivity, followed by C-3, with half to one-fifth that of C-4. Only about 1% of the total radioactivity was in C-1. The data are compared with the predictions of the classical pentose phosphate cycle [Horecker, Gibbs, Klenow & Smyrniotis (1954) J. Biol. Chem.207, 393–403], and the proposed new version of the pentose phosphate cycle in liver [Longenecker & Williams (1980) Biochem. J.188, 847–857], which they denoted as the ‘L-type pentose cycle’. The Williams pathway predicts that the specific radioactivity of C-1 of glucose should be half that of C-4 (after correction for approximately equal labelling on C-3 and C-4 of hexose phosphate in the pathway involving fructose 1,6-bisphosphatase). The actual labelling in C-1 is 20–350-fold less than this. When the hepatocytes are incubated with phenazine methosulphate, to stimulate the oxidative branch of the pentose phosphate cycle, the predicted relationship between (C-2/C-3) and (C-1/C-3) ratios of specific radio-activities is nearly exactly in accord with the classical pentose phosphate cycle. Glucose and glucose 6-phosphate were isolated and degraded from an incubation of hepatocytes from starved/re-fed rats with [3-14C]xylitol. Although the patterns were of the classical type, there was more randomization of 14C into C-2 and C-1 in the glucose 6-phosphate isolated at the end of the incubation than in the glucose which was continuously produced.

scholarly journals The pentose phosphate pathway in rabbit liver. Studies on the metabolic sequence and quantitative role of the pentose phosphate cycle by using a system in situ

1971 ◽  
Vol 123 (5) ◽  
pp. 923-943 ◽  
Author(s):  
J. F. Williams ◽  
K. G. Rienits ◽  
P. J. Schofield ◽  
M. G. Clark
Keyword(s):  
Pentose Phosphate Pathway ◽  
Time Course ◽  
Specific Radioactivity ◽  
Glucose Production ◽  
Pentose Phosphate ◽  
Rate Of Change ◽  
Exchange Reactions ◽  
High Concentration ◽  
Pentose Phosphate Cycle ◽  
Definition Of

1. The reactions of the pentose phosphate cycle were investigated by the intraportal infusion of specifically labelled [14C]glucose or [14C]ribose into the liver of the anaesthetized rabbit. The sugars were confined in the liver by haemostasis and metabolism was allowed to proceed for periods up to 5min. Metabolism was assessed by measuring the rate of change of the specific radioactivity of CO2, the carbon atoms of glucose 6-phosphate, fructose 6-phosphate and tissue glucose. 2. The quotient oxidation of [1-14C]glucose/oxidation of [6-14C]glucose as measured by the incorporation into respiratory CO2 was greater than 1.0 during most of the time-course and increased to a maximum of 3.1 but was found to decrease markedly upon application of a glucose load. 3. The estimate of the pentose phosphate cycle from C-1/C-2 ratios generally increased during the time-course, whereas the estimate of the pentose phosphate cycle from C-3/C-2 ratios varied depending on whether the ratios were measured in glucose or hexose 6-phosphates. 4. The distribution of 14C in hexose 6-phosphate after the metabolism of [1-14C]ribose showed that 65–95% of the label was in C-1 and was concluded to have been the result of a rapidly acting transketolase exchange reaction. 5. Transaldolase exchange reactions catalysed extensive transfer of 14C from [2-14C]glucose into C-5 of the hexose 6-phosphates during the entire time-course. The high concentration of label in C-4, C-5 and C-6 of the hexose 6-phosphates was not seen in tissue glucose in spite of an unchanging rate of glucose production during the time-course. 6. It is concluded that the reaction sequences catalysed by the pentose phosphate pathway enzymes do not constitute a formal metabolic cycle in intact liver, neither do they allow the definition of a fixed stoicheiometry for the dissimilation of glucose.

scholarly journals Regulation of the oxidative phase of the pentose phosphate cycle in mussels

1978 ◽  
Vol 170 (3) ◽  
pp. 577-585 ◽  
Author(s):  
S Rodriguez-Segade ◽  
M Freire ◽  
A Carrion
Keyword(s):  
Inorganic Ions ◽  
Pentose Phosphate ◽  
Oxidized Glutathione ◽  
Phosphogluconate Dehydrogenase ◽  
Pentose Phosphate Cycle ◽  
Intracellular Concentrations ◽  
Glucose 6 Phosphate Dehydrogenase ◽  
Substrate Concentrations

1. The mechanisms that control the oxidative phase of the pentose phosphate cycle in mussel hepatopancreas were investigated. 2. The effects of GSSG (oxidized glutathione) on the inhibition of glucose 6-phosphate dehydrogenase by NADPH [Eggleston & Krebs (1974) Biochem. J. 138, 425-435] extend to 6-phosphogluconate dehydrogenase. 3. The effect of GSSG on both enzymes increases as the [NADP+1]/[NADPH] ratio decreases; greater percentage deinhibition always was obtained for 6-phosphogluconate dehydrogenase. 4. Increasing concentration of GSSG increased the percentage deinhibition. This effect is more pronounced with 6-phosphogluconate dehydrogenase. 5. We confirmed the apparent imbalance between the activities of the two enzymes [sapag-Hagar, Lagunas & Sols (1973) Biochem. Biophys. Res. Commun, 50, 179-185] in the presence of 10mM-Mg2+. 6. The imbalance practically disappears when the substrate concentrations are less than saturating and Mg2+ approaches physiological concentrations. 7. The addition of GSSG at physiological concentrations allows the activities of both enzymes to be measured at high [NADPH]/[NADP+] ratios ratios and the co-operative action of GSSG and Mg2+ on the imbalance between the two enzymes to be verifed. 8. The control of the activity of the two enzymes of the pentose cycle could be carried out by deinhibition of the two dehydrogenases and by the intracellular concentrations of substrates and inorganic ions.

Effect of pentobarbital on fructose 2,6-bisphosphate metabolism in isolated rat hepatocytes

1985 ◽  
Vol 249 (5) ◽  
pp. E525-E533
Author(s):  
F. Nyfeler ◽  
M. R. el-Maghrabi ◽  
S. J. Pilkis
Keyword(s):  
Kinase Activity ◽  
Lactate Production ◽  
Rat Hepatocytes ◽  
Protein Kinase Activity ◽  
Dependent Protein Kinase ◽  
Isolated Rat Hepatocytes ◽  
Fructose 1,6 Bisphosphatase ◽  
Dependent Protein ◽  
Dose Dependent Decrease ◽  
Dose Dependent

Addition of the commonly used anesthetic pentobarbital to hepatocytes from fed rats resulted in a dose-dependent decrease in the level of fructose 2,6-bisphosphate. At a concentration of pentobarbital (0.4 mM) that lowered fructose 2,6-bisphosphate by 60%, there was no significant change in the level of fructose 6-phosphate, ATP, or L-glycerol 3-phosphate. Higher concentrations of pentobarbital (2 mM) enhanced both glycolysis and glycogenolysis and fructose 2,6-bisphosphate levels were reduced to less than 10% of the control. Concomitant with these changes there was a decrease in ATP, glucose 6-phosphate, and fructose 6-phosphate and a two- and fivefold increase in ADP and AMP, respectively. In hepatocytes from starved rats pentobarbital also lowered ATP levels and inhibited gluconeogenesis but had no effect on either lactate production or the already low level of sugar diphosphate. However, in the fasted case pentobarbital completely prevented the 10-fold elevation of fructose 2,6-bisphosphate brought about by 30 mM glucose. The anesthetic had no effect on cAMP-dependent protein kinase activity or on pyruvate kinase activity in hepatocytes from fed or starved rats but caused reciprocal changes in the activities of the bifunctional enzyme 6-phosphofructo-2-kinase/fructose 2,6-bisphosphatase. Kinase activity was decreased and bisphosphatase activity was increased. These results suggest that the effects of pentobarbital on gluconeogenesis and glycolysis are due to inhibition of energy metabolism with elevated AMP levels causing activation of 6-phosphofructo-1-kinase and inhibition of fructose 1,6-bisphosphatase.(ABSTRACT TRUNCATED AT 250 WORDS)

scholarly journals Effects of amino acids, ammonia and leupeptin on protein synthesis and degradation in isolated rat hepatocytes

1978 ◽  
Vol 174 (2) ◽  
pp. 469-474 ◽  
Author(s):  
P O Seglen
Keyword(s):  
Amino Acids ◽  
Protein Synthesis ◽  
Protein Degradation ◽  
Specific Radioactivity ◽  
Rat Hepatocytes ◽  
Amino Acid Mixture ◽  
Acid Mixture ◽  
Isolated Rat Hepatocytes ◽  
Lysosomal Protein ◽  
Isolated Rat

Protein synthesis in isolated rat hepatocytes, as measured by the incorporation of [14C]-valine at constant specific radioactivity, proceeded at a rate of 0.3-0.5%/h in an unsupplemented medium, i.e. only about one-tenth the rate of protein degradation (4%/h). Leupeptin, which inhibits lysosomal protein degradation (previously found to be 75% of the total degradation in hepatocytes), had no effect on protein synthesis, showing that endogenous protein degradation supplied amino acids in excess of the substrate requirements for protein synthesis. The inhibition of protein synthesis by NH4Cl (another inhibitor of lysosomal protein degradation) as well as the stimulation by a physiological amino acid mixture must therefore represent indirect effects, either on general energy metabolism, or on unknown regulatory processes.

scholarly journals Interrelationship and control of glucose metabolism and lipogenesis in isolated fat-cells. Control of pentose phosphate-cycle activity by cellular requirement for reduced nicotinamide adenine dinucleotide phosphate

1972 ◽  
Vol 128 (5) ◽  
pp. 1097-1102 ◽  
Author(s):  
H. Kather ◽  
M. Rivera ◽  
K. Brand
Keyword(s):  
Fatty Acid ◽  
Glucose Metabolism ◽  
Fatty Acid Synthesis ◽  
Nicotinamide Adenine Dinucleotide Phosphate ◽  
Acid Synthesis ◽  
Pentose Phosphate ◽  
Fat Cells ◽  
Isolated Fat Cells ◽  
Pentose Phosphate Cycle ◽  
Phenazine Methosulphate

By using inhibitors and stimulators of different metabolic pathways the interdependence of the pentose phosphate cycle and lipogenesis in isolated fat-cells was studied. Rotenone, which is known to inhibit electron transport in the respiratory chain, blocked glucose breakdown at the site of pyruvate dehydrogenase. Consequently, because of the lack of acetyl-CoA, fatty acid synthesis was almost abolished. A concomitant decrease in pentose phosphate-cycle activity was observed. Phenazine methosulphate stimulated pentose phosphate-cycle activity about five- to ten-fold without a considerable effect on fatty acid synthesis. The influence of rotenone on both the pentose phosphate cycle and lipogenesis could be overcome by addition of phenazine methosulphate, indicating that rotenone has no direct effect on these pathways. The decreased rate of the pentose phosphate cycle in the presence of rotenone therefore has to be considered as a consequence of decreased fatty acid synthesis. The rate of glucose catabolism via the pentose phosphate cycle in adipocytes appears to be determined by the requirement of NADPH for lipogenesis. Treatment of cells with 6-aminonicotinamide caused an accumulation of 6-phosphogluconate, indicating an inhibition of 6-phosphogluconate dehydrogenase. The rate of glucose metabolism via the pentose phosphate cycle as well as the rate of fatty acid synthesis, however, was not affected by 6-aminonicotinamide treatment and could still be stimulated by addition of insulin. Since even in cells from starved animals, in which the pentose phosphate-cycle activity is extremely low, no accumulation of 6-phosphogluconate was observed, it is concluded that the control of this pathway is achieved by the rate of regeneration of NADP at the site of glucose 6-phosphate dehydrogenase.

Glucose handling by hepatocytes from obese Zucker rats

1991 ◽  
Vol 11 (5) ◽  
pp. 285-292 ◽  
Author(s):  
Neus Carbó ◽  
Francisco J. López-Soriano ◽  
Josep M. Argilés
Keyword(s):  
Fatty Acids ◽  
Lipid Synthesis ◽  
Pentose Phosphate ◽  
High Rate ◽  
Zucker Rats ◽  
Esterified Fatty Acids ◽  
Obese Zucker Rats ◽  
Glucose 6 Phosphate Dehydrogenase ◽  
Phenazine Methosulphate ◽  
Substrate Concentrations

Hepatocytes isolated from obese Zucker rats showed a significantly higher rate of both [U-14C]glucose and [U-14C]lactate incorporation into [14C]lipid than those from their lean counterparts. This was associated with a marked increase in the lipogenic rate measured by the incorporation of3H2O into the cell esterified fatty acids. Although there were no changes in the incorporation of the tracer into either [14C]glycogen or14CO2, the [14C] total uptake was significantly higher in the obese animals. The high rate of [14C]lipid synthesis from glucose was observed both at 15 and 30 mM substrate concentrations and was linked to an enhanced uptake of the tracer into the cell as measured using the decarboxilation of [1-14C]glucose in the presence of phenazine methosulphate. The presence of insulin in the incubation medium had no effect on the uptake of glucose by the liver cells. However, the large uptake of glucose by the hepatocytes from the obese animals was not related to an enhanced rate of transport as measured using 3-O-methyl[U-14C]glucose. The activity of glucose-6-phosphate dehydrogenase together with a higher [1-14C]glucose/[U-14C]glucose descarboxylation ratio indicate a predominant very active pentose phosphate pathway which may be responsible for the enhanced glucose uptake observed in the hepatocytes from the obese animals.

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