scholarly journals Phosphorylation state of cytosolic and mitochondrial adenine nucleotides and of pyruvate dehydrogenase in isolated rat liver cells

1976 ◽  
Vol 156 (1) ◽  
pp. 91-102 ◽  
Author(s):  
E A Siess ◽  
O H Wieland
Keyword(s):  
Pyruvate Dehydrogenase ◽  
Adenine Nucleotides ◽  
Active Form ◽  
Inverse Correlation ◽  
Liver Cells ◽  
Diabetic Rats ◽  
Pyruvate Dehydrogenase Kinase ◽  
Intact Cells ◽  
Phosphorylation State

1. Cytosolic and mitochondrial ATP and ADP concentrations of liver cells isolated from normal fed, starved and diabetic rats were determined. 2. The cytosolic ATP/ADP ratio was 6,9 and 10 in normal fed, starved and diabetic rats respectively. 3. The mitochondrial ATP/ADP ratio was 2 in normal and diabetic rats and 1.6 in starved rats. 4. Adenosine increased the cytosolic and lowered the mitochondrial ATP/ADP ratio, whereas atractyloside had the opposite effect. 5. Incubation of the hepatocytes with fructose, glycerol or sorbitol led to a fall in the ATP/ADP ratio in both the cytosolic and the mitochondrial compartment. 6. The interrelationship between the mitochondrial ATP/ADP ratio and the phosphorylation state of pyruvate dehydrogenase in intact cells was studied. 7. In hepatocytes isolated from fed rats an inverse correlation between the mitochondrial ATP/ADP ratio and the active form of pyruvate dehydrogenase (pyruvate dehydrogenase a) was demonstrable on loading with fructose, glycerol or sorbitol. 8. No such correlation was obtained with pyruvate or dihydroxyacetone. For pyruvate, this can be explained by inhibition of pyruvate dehydrogenase kinase. 9. Liver cells isolated from fed animals displayed pyruvate dehydrogenase a activity twice that found in vivo. Physiological values were obtained when the hepatocytes were incubated with albumin-oleate, which also yielded the highest mitochondrial ATP/ADP ratio.

scholarly journals Inhibition of lactate glucogneogenesis in rat kidney by dichloroacetate

1978 ◽  
Vol 170 (3) ◽  
pp. 551-560 ◽  
Author(s):  
J H Lacey ◽  
P J Randle
Keyword(s):  
Pyruvate Dehydrogenase ◽  
Rat Kidney ◽  
Active Form ◽  
Glucose Production ◽  
Inhibitory Effect ◽  
Diabetic Rats ◽  
Pyruvate Dehydrogenase Kinase ◽  
No Oxidation ◽  
Kidney Cortex

1. Sodium dichloroacetate (1mM) inhibited glucose production from L-lactate in kidney-cortex slices from fed, starved or alloxan-diabetic rates. In general gluconeogenesis from other substrates was no inhibited. 2. Sodium dichloracetate inhibited glucose production from L-lactate but no from pyruvate in perfused isolated kidneys from normal or alloxan-diabetic rats. 3. Sodium dichloroacetate is an inhibitor of the pyruvate dehydrogenase kinase reaction and it effected conversion of pyruvate dehydrogenase into its its active (dephosphorylated) form in kidney in vivo. In general, pyruvate dehydrogenase was mainly in the active form in kidneys perfused or incubated with L-lactate and the inhibitory effect of dichloroacetate on glucose production was not dependent on activation of pyruvate dehydrogenase. 4. Balance data from kidney slices showed that dichloroacetate inhibits lactate uptake, glucose and pyruvate production from lactate, but no oxidation of lactate. 5. The mechanism of this effect of dichloroactetate on glucose production from lactate has not been fully defined, but evidence suggests that it may involve a fall in tissue pyruvate concentration and inhibition of pyruvate carboxylation.

scholarly journals The metabolism of l-phenylalanine and l-tyrosine by liver cells isolated from adrenalectomized rats and from streptozotocin-diabetic rats

1985 ◽  
Vol 228 (1) ◽  
pp. 249-255 ◽  
Author(s):  
J C Stanley ◽  
M J Fisher ◽  
C I Pogson
Keyword(s):  
Insulin Treatment ◽  
Maximal Activity ◽  
Liver Cells ◽  
Diabetic Rats ◽  
Steroid Treatment ◽  
Phosphorylation State ◽  
Tyrosine Metabolism ◽  
Streptozotocin Diabetic Rats ◽  
Adrenalectomized Rats

Flux through, and maximal activities of, key enzymes of phenylalanine and tyrosine degradation were measured in liver cells prepared from adrenalectomized rats and from streptozotocin-diabetic rats. Adrenalectomy decreased the phenylalanine hydroxylase flux/activity ratio; this was restored by steroid treatment in vivo. Changes in the phosphorylation state of the hydroxylase may mediate these effects; there was no significant change in the maximal activity of the hydroxylase. Tyrosine metabolism was enhanced by adrenalectomy; this was not related to any change in maximal activity of the aminotransferase. Steroid treatment increased the maximal activity of the aminotransferase. Both acute (3 days) and chronic (10 days) diabetes were associated with increased metabolism of phenylalanine; insulin treatment in vivo did not reverse these changes. Although elevated hydroxylase protein concentration was a major factor, changes in the enzyme phosphorylation state may contribute to differences in phenylalanine degradation in the acute and chronic diabetic states. Tyrosine metabolism, increased by diabetes, was partially restored to normal by insulin treatment in vivo. These changes can, to a large extent, be interpreted in terms of changes in the maximal activity of the aminotransferase.

scholarly journals Glucose metabolism in perfused skeletal muscle. Pyruvate dehydrogenase activity in starvation, diabetes and exercise

1976 ◽  
Vol 158 (2) ◽  
pp. 203-210 ◽  
Author(s):  
S A Hagg ◽  
S I Taylor ◽  
N B Ruberman
Keyword(s):  
Skeletal Muscle ◽  
Dehydrogenase Activity ◽  
Pyruvate Dehydrogenase ◽  
Preceding Paper ◽  
Diabetic Rats ◽  
Pyruvate Dehydrogenase Kinase ◽  
Lactate Oxidation ◽  
Pyruvate Oxidation ◽  
Measured Activity ◽  
Exercise Induced

1. The interconversion of pyruvate dehydrogenase between its inactive phosphorylated and active dephosphorylated forms was studied in skeletal muscle. 2. Exercise, induced by electrical stimulation of the sciatic nerve (5/s), increased the measured activity of (active) pyruvate dehydrogenase threefold in intact anaesthetized rated within 2 min. No further increase was seen after 15 min of stimulation. 3. In the perfused rat hindquarter, (active) pyruvate dehydrogenase activity was decreased by 50% in muscle of starved and diabetic rats. Exercise produced a twofold increase in its activity in all groups; however, the relative differences between fed, starved and diabetic groups persisted. 4. Perfusion of muslce with acetoacetate (2 mM) decreased (active) pyruvate dehydrogenase activity by 50% at rest but not during exercise. 5. Whole-tissue concentrations of pyruvate and citrate, inhibitors of (active) pyruvate dehydrogenase kinase and (inactive) pyruvate dehydrogenase phosphate phosphatase respectively, were not altered by excerise. A decrease in the ATP/ADP ratio was observed, but did not appear to be sufficient to account for the increase in (active) pyruvate dehydrogenase activity. 6. The results suggest that interconversion of the phosphorylated and dephosphorylated forms of pyruvate dehydrogenase plays a major role in the regulation of pyruvate oxidation by eomparison of enzyme activity with measurements of lactate oxidation in the perfused hindquarter [see the preceding paper, Berger et al. (1976)] suggest that pyruvate oxidation is also modulated by the concentrations of substrates, cofactors and inhibitors of (active) pyruvate dehydrogenase activity.

scholarly journals Studies of the long-term regulation of hepatic pyruvate dehydrogenase kinase

1998 ◽  
Vol 329 (1) ◽  
pp. 89-94 ◽  
Author(s):  
C. Mary SUGDEN ◽  
G. D. Lee FRYER ◽  
A. Karen ORFALI ◽  
A. David PRIESTMAN ◽  
Elaine DONALD ◽  
...  
Keyword(s):  
Fatty Acids ◽  
Pyruvate Dehydrogenase ◽  
Unsaturated Fatty Acids ◽  
Fold Increase ◽  
Membrane Lipid ◽  
Dietary Lipid ◽  
Pyruvate Dehydrogenase Kinase ◽  
Plasma Insulin Concentration

The administration of a low-carbohydrate/high-saturated-fat (LC/HF) diet for 28 days or starvation for 48 h both increased pyruvate dehydrogenase kinase (PDHK) activity in extracts of rat hepatic mitochondria, by approx. 2.1-fold and 3.5-fold respectively. ELISAs of extracts of hepatic mitochondria, conducted over a range of pyruvate dehydrogenase (PDH) activities, revealed that mitochondrial immunoreactive PDHKII (the major PDHK isoform in rat liver) was significantly increased by approx. 1.4-fold after 28 days of LC/HF feeding and by approx. 2-fold after 48 h of starvation. The effect of LC/HF feeding to increase hepatic PDHK activity was retained through hepatocyte preparation, but was decreased on 21 h culture with insulin (100μ-i.u./ml). A sustained (24 h) 2-4-fold elevation in plasma insulin concentration in vivo (achieved by insulin infusion via an osmotic pump) suppressed the effect of LC/HF feeding so that hepatic PDHK activities did not differ significantly from those of (insulin-infused) control rats. The increase in hepatic PDHK activity evoked by 28 days of LC/HF feeding was prevented and reversed (within 24 h) by the replacement of 7% of the dietary lipid with long-chain ω-3 fatty acids. Analysis of hepatic membrane lipid revealed a 1.9-fold increase in the ratio of total polyunsaturated ω-3 fatty acids to total mono-unsaturated fatty acids. The results indicate that the increased hepatic PDHK activities observed in livers of LC/HF-fed or 48 h-starved rats are associated with long-term actions to increase hepatic PDHKII concentrations. The long-term regulation of hepatic PDHK by LC/HF feeding might be achieved through an impaired action of insulin to suppress PDHK activity. In addition, the fatty acid composition of the diet, rather than the fat content, is a key influence.

Increased hepatic pyruvate dehydrogenase kinase activity in fed hyperthyroid rats: studies in vivo and with cultured hepatocytes

1996 ◽  
Vol 119 (2) ◽  
pp. 219-224 ◽  
Author(s):  
Mary C. Sugden ◽  
Lee G.D. Fryer ◽  
David A. Priestman ◽  
Karen A. Orfali ◽  
Mark J. Holness
Keyword(s):  
Pyruvate Dehydrogenase ◽  
Kinase Activity ◽  
Pyruvate Dehydrogenase Kinase ◽  
Cultured Hepatocytes

Overexpression of pyruvate dehydrogenase kinase 4 in heart perturbs metabolism and exacerbates calcineurin-induced cardiomyopathy

2008 ◽  
Vol 294 (2) ◽  
pp. H936-H943 ◽  
Author(s):  
Guixiang Zhao ◽  
Nam Ho Jeoung ◽  
Shawn C. Burgess ◽  
Kimberly A. Rosaaen-Stowe ◽  
Takeshi Inagaki ◽  
...  
Keyword(s):  
Transgenic Mice ◽  
Pyruvate Dehydrogenase ◽  
Marked Decrease ◽  
Active Form ◽  
Pyruvate Dehydrogenase Kinase ◽  
Metabolic Flexibility ◽  
Energy Demands ◽  
Pyruvate Dehydrogenase Kinase 4 ◽  
Fatty Acid Catabolism

The heart adapts to changes in nutritional status and energy demands by adjusting its relative metabolism of carbohydrates and fatty acids. Loss of this metabolic flexibility such as occurs in diabetes mellitus is associated with cardiovascular disease and heart failure. To study the long-term consequences of impaired metabolic flexibility, we have generated mice that overexpress pyruvate dehydrogenase kinase (PDK)4 selectively in the heart. Hearts from PDK4 transgenic mice have a marked decrease in glucose oxidation and a corresponding increase in fatty acid catabolism. Although no overt cardiomyopathy was observed in the PDK4 transgenic mice, introduction of the PDK4 transgene into mice expressing a constitutively active form of the phosphatase calcineurin, which causes cardiac hypertrophy, caused cardiomyocyte fibrosis and a striking increase in mortality. These results demonstrate that cardiac-specific overexpression of PDK4 is sufficient to cause a loss of metabolic flexibility that exacerbates cardiomyopathy caused by the calcineurin stress-activated pathway.

scholarly journals Mechanism of activation of pyruvate dehydrogenase by mitogens in pig lymphocytes

1983 ◽  
Vol 216 (2) ◽  
pp. 359-367 ◽  
Author(s):  
E Baumgarten ◽  
M D Brand ◽  
T Pozzan
Keyword(s):  
Dehydrogenase Activity ◽  
Concanavalin A ◽  
Pyruvate Dehydrogenase ◽  
Pyruvate Dehydrogenase Kinase ◽  
Ionophore A23187 ◽  
Intact Cells ◽  
Permeabilized Cells ◽  
Maximal Activation ◽  
Pyruvate Dehydrogenase Phosphatase

The activity of pyruvate dehydrogenase in extracts of pig mesenteric lymphocytes was measured under different preincubation conditions. The mitogens concanavalin A and ionophore A23187 both increased pyruvate dehydrogenase activity. In both cases activation required extracellular Ca2+. Digitonin-permeabilized cells required 0.5 microM free Ca2+ for half-maximal activation of pyruvate dehydrogenase. The stimulation by concanavalin A in intact cells was probably not due to changes in effectors of pyruvate dehydrogenase kinase. This evidence suggests that activation of pyruvate dehydrogenase is by Ca2+ activation of pyruvate dehydrogenase phosphatase and supports the view that the cytoplasmic free [Ca2+] rises to something less than 1 microM on stimulation with mitogens.

Influence of caveolin on constitutively activated recombinant eNOS: insights into eNOS dysfunction in BDL rat liver

2003 ◽  
Vol 285 (3) ◽  
pp. G652-G660 ◽  
Author(s):  
H. Hendrickson ◽  
S. Chatterjee ◽  
S. Cao ◽  
M. Morales Ruiz ◽  
W. C. Sessa ◽  
...  
Keyword(s):  
Rat Liver ◽  
Hepatic Stellate Cells ◽  
Fluorescent Protein ◽  
Stellate Cell ◽  
Active Form ◽  
Stellate Cells ◽  
Liver Cells ◽  
No Production

Diminished endothelial nitric oxide (NO) synthase (eNOS)-derived NO production from the hepatic vascular endothelium contributes to hepatic vasoconstriction in portal hypertension. The aim of this study was to examine the mechanism of this process by testing the influence of a constitutively active form of eNOS (S1179DeNOS) in both primary and propagated liver cells in vitro and in the sham and bile duct ligated (BDL) rat liver in vivo, using an adenoviral vector encoding green fluorescent protein (AdGFP) and S1179DeNOS (AdS1179DeNOS). AdS1179DeNOS transduction augmented basal and agonist-stimulated NO generation in nonparenchymal liver cells. Sham rats transduced in vivo with AdS1179DeNOS evidenced a decreased pressor response to incremental doses of the vasoconstrictor methoxamine compared with sham rats transduced with AdGFP. However, BDL rats transduced with AdS1179DeNOS did not display improved vasodilatory responses as evidenced by similar flow-dependent pressure increases to that observed in BDL rats transduced with AdGFP, despite similar levels of viral transgene expression. We next examined the influence of the eNOS inhibitory protein caveolin on S1179DeNOS dysfunction in cirrhotic liver. Immunogold electron microscopic analysis of caveolin in BDL liver demonstrated prominent expression not only in liver endothelial cells, but also in hepatic stellate cells. In vitro studies in the LX2 hepatic stellate cell line demonstrate that caveolin precipitates recombinant S1179DeNOS in LX2 cells, that recombinant S1179DeNOS coprecipitates caveolin, and that binding is enhanced in the presence of overexpression of caveolin. Furthermore, caveolin overexpression inhibits recombinant S1179DeNOS activity. These studies indicate that recombinant S1179DeNOS protein functions appropriately in normal liver cells and tissue but evidences dysfunction in the cirrhotic rat liver and that caveolin expression and inhibition in BDL nonparenchymal cells, including hepatic stellate cells, may account for this dysfunction.

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