scholarly journals Molybdate and tungstate act like vanadate on glucose metabolism in isolated hepatocytes

1992 ◽  
Vol 282 (3) ◽  
pp. 659-663 ◽  
Author(s):  
C Fillat ◽  
J E Rodríguez-Gil ◽  
J J Guinovart
Keyword(s):  
Glucose Metabolism ◽  
Glycogen Phosphorylase ◽  
Kinase Activity ◽  
Glycogen Synthase ◽  
Rat Hepatocytes ◽  
Isolated Hepatocytes ◽  
The Other ◽  
Co2 Production ◽  
Glycolytic Flux ◽  
Dependent Mechanism

In rat hepatocytes, molybdate and tungstate inactivate glycogen synthase by a mechanism independent of Ca2+ and activate glycogen phosphorylase by a Ca(2+)-dependent mechanism. On the other hand, both molybdate and tungstate increase fructose 2,6-bisphosphate levels and counteract the decrease in this metabolite induced by glucagon. These effectors do not directly modify 6-phosphofructo-2-kinase activity, even though they partially counteract the inactivation of this enzyme induced by glucagon. These effects are related to an increase on the glycolytic flux, as indicated by the increase in L-lactate and CO2 production and the decrease in glucose 6-phosphate levels in the presence of glucose. All these effects are similar to those previously reported for vanadate, although molybdate and tungstate are less effective than vanadate. These results could indicate that molybdate, tungstate and vanadate act on glucose metabolism in isolated hepatocytes by a similar mechanism of action.

Fatty acid and amino acid modulation of glucose cycling in isolated rat hepatocytes

2001 ◽  
Vol 358 (3) ◽  
pp. 665-671 ◽  
Author(s):  
Lori A. GUSTAFSON ◽  
Mies NEEFT ◽  
Dirk-Jan REIJNGOUD ◽  
Folkert KUIPERS ◽  
Hans P. SAUERWEIN ◽  
...  
Keyword(s):  
Amino Acids ◽  
Glycogen Phosphorylase ◽  
Glycogen Synthase ◽  
Lactate Production ◽  
Rat Hepatocytes ◽  
Glycolytic Flux ◽  
Isolated Rat Hepatocytes ◽  
Glycogen Accumulation ◽  
Glycogen Deposition ◽  
Intracellular Glucose

We studied the influence of glucose/glucose 6-phosphate cycling on glycogen deposition from glucose in fasted-rat hepatocytes using S4048 and CP320626, specific inhibitors of glucose-6-phosphate translocase and glycogen phosphorylase respectively. The effect of amino acids and oleate was also examined. The following observations were made: (1) with glucose alone, net glycogen production was low. Inhibition of glucose-6-phosphate translocase increased intracellular glucose 6-phosphate (3-fold), glycogen accumulation (5-fold) without change in active (dephosphorylated) glycogen synthase (GSa) activity, and lactate production (4-fold). With both glucose 6-phosphate translocase and glycogen phosphorylase inhibited, glycogen deposition increased 8-fold and approached reported in vivo rates of glycogen deposition during the fasted → fed transition. Addition of a physiological mixture of amino acids in the presence of glucose increased glycogen accumulation (4-fold) through activation of GS and inhibition of glucose-6-phosphatase flux. Addition of oleate with glucose present decreased glycolytic flux and increased the flux through glucose 6-phosphatase with no change in glycogen deposition. With glucose 6-phosphate translocase inhibited by S4048, oleate increased intracellular glucose 6-phosphate (3-fold) and net glycogen production (1.5-fold), without a major change in GSa activity. It is concluded that glucose cycling in hepatocytes prevents the net accumulation of glycogen from glucose. Amino acids activate GS and inhibit flux through glucose-6-phosphatase, while oleate inhibits glycolysis and stimulates glucose-6-phosphatase flux. Variation in glucose 6-phosphate does not always result in activity changes of GSa. Activation of glucose 6-phosphatase flux by fatty acids may contribute to the increased hepatic glucose production as seen in Type 2 diabetes.

scholarly journals Prostaglandins E2 and F2α affect glycogen synthase and phosphorylase in isolated hepatocytes

1989 ◽  
Vol 261 (1) ◽  
pp. 93-97 ◽  
Author(s):  
A M Gómez-Foix ◽  
J E Rodriguez-Gil ◽  
J J Guinovart ◽  
F Bosch
Keyword(s):  
Glycogen Phosphorylase ◽  
Glycogen Synthase ◽  
Rat Hepatocytes ◽  
Isolated Hepatocytes ◽  
Time Dependent ◽  
Dependent Manner ◽  
Metabolizing Enzymes ◽  
Prostaglandin F2 Alpha ◽  
Parenchymal Liver ◽  
Prostaglandins E2 And F2α

Prostaglandin E2 (PGE2) and prostaglandin F2 alpha (PGF2 alpha) inactivated glycogen synthase and activated glycogen phosphorylase in rat hepatocytes in a dose- and time-dependent manner. These effects were dependent on the presence of Ca2+ in the incubation medium. When glycogen synthase was immunoprecipitated from cells incubated with [32P]Pi and then treated with PGE2 or PGF2 alpha, there was increased phosphorylation of the 88 kDa subunit of the enzyme. This phosphorylation affected two CNBr fragments of the glycogen synthase, CB-1 and CB-2, the same fragments that are phosphorylated by different glycogenolytic hormones. No phosphorylation of glycogen synthase by prostaglandins was observed in the absence of Ca2+. Thus the effect of PGE2 and PGF2 alpha on these glycogen-metabolizing enzymes supports a role for regulation by prostaglandins of glucose metabolism in parenchymal liver cells.

scholarly journals Effects of C-1-substituted glucose analogue on the activation states of glycogen synthase and glycogen phosphorylase in rat hepatocytes

1995 ◽  
Vol 311 (3) ◽  
pp. 845-852 ◽  
Author(s):  
M Board ◽  
M Bollen ◽  
W Stalmans ◽  
Y Kim ◽  
G W J Fleet ◽  
...  
Keyword(s):  
Glycogen Phosphorylase ◽  
Kinase Inhibitor ◽  
Glycogen Synthase ◽  
Kinetic Studies ◽  
Rat Hepatocytes ◽  
Glycogen Metabolism ◽  
Isolated Hepatocytes ◽  
Effective Control ◽  
Subsequent Addition ◽  
Glucose Analogue

A series of glucose-analogue inhibitors of glycogen phosphorylase b (GPb) has been designed, synthesized and investigated in crystallographic binding and kinetic studies. The aim is to produce a compound that may exert more effective control over glycogen metabolism than the parent glucose molecule and which could alleviate hyperglycaemia in Type-II diabetes. N-Acetyl-beta-D-glucopyranosylamine (1-GlcNAc) has a Ki for muscle GPb in crude extracts of 30 microM, 367-fold lower than that of beta-D-glucose [Board, Hadwen and Johnson (1995) Eur. J. Biochem. 228, 753-761]. In the current work, the effects of 1-GlcNAc on the activation states of GP and glycogen synthase (GS) in cell-free preparations and in isolated hepatocytes are reported. In gel-filtered extracts of liver, which lack ATP for kinase activity, 1-GlcNAc produced a rapid and time-dependent inactivation of GP with a subsequent activation of GS. Effects of 1-GlcNAc on both enzymes were stronger than those of glucose, with 0.8 mM 1-GlcNAc being equipotent with 50 mM glucose. At 1 mM, 1-GlcNAc enhanced the dephosphorylation of exogenous GPa by liver extracts (600%) and by muscle extracts (75%). This represents an approximately 500-fold improvement on glucose for the liver activity and 40-fold for the muscle activity. In whole hepatocytes, 1-GlcNAc showed an approximately 5-fold enhancement of glucose effects for GP inactivation but failed to elicit activation of GS. Glucose-induced activation of GS in whole hepatocytes was reversed by subsequent addition of 1-GlcNAc. However, when GS activation was achieved via the adenosine analogue and kinase inhibitor, 5′-iodotubercidin (ITU), subsequent addition of 1-GlcNAc allowed continued activation of GS. Phosphorylation of 1-GlcNAc in rat hepatocytes was established using radiolabelled material. The rate of phosphorylation was 1.60 nmol/min per 10(6) cells at 20 mM 1-GlcNAc but was reduced by the presence of 50 microM ITU (0.775 nmol/min per 10(6) cells). It is suggested that the phosphorylated derivative of 1-GlcNAc formed in hepatocytes is 1-GlcNAc 6-phosphate and that the presence of this species is responsible for the failure of 1-GlcNAc to activate GS. The relative importance of the reduction in concentration of GPa versus increased glucose 6-phosphate levels for activation of GS is discussed.

scholarly journals Effects of lactation of ketogenesis from oleate or butyrate in rat hepatocytes

1977 ◽  
Vol 164 (3) ◽  
pp. 521-528 ◽  
Author(s):  
E Whitelaw ◽  
D H Williamson
Keyword(s):  
Ketone Bodies ◽  
Rat Hepatocytes ◽  
Isolated Hepatocytes ◽  
The Other ◽  
Glycolytic Flux ◽  
Control Group ◽  
Triacylglycerol Synthesis ◽  
Group 2 ◽  
Long Chain ◽  
Peak Lactation

1. Rates of ketogenesis from endogenous butyrate or oleate were measured in isolated hepatocytes prepared from fed rats during different reproductive states [virgin, pregnant, early-lactating (2-4 days) and peak-lactating (10-17 days)]. In the peak-lactation group there was a decrease (25%) in the rate of ketogenesis from butyrate, but there were no differences in the rates between the other groups. Wth oleate, the rate of ketogenesis was increased in the pregnant and in the early-lactation groups compared with the virgin group, whereas the rate was 50% lower in the peak-lactation group. 2. Experiments with [1-(14)C]oleate indicated that these differences in rates of ketogenesis were not due to alterations in the rate of oleate utilization, but to changes in the amount of oleoyl-CoA converted into ketone bodies. 3. Although the addition of carnitine increased the rates of ketogenesis from oleate in all groups of rats, it did not abolish the differences between the groups. 4. Measurements of the accumulation of glucose and lactate showed that hepatocytes from rats at peak lactation had a higher rate of glycolytic flux than did hepatocytes from the other groups. After starvation, the rate of ketogenesis from oleate was still lower in the peak-lactation group compared with the control group. This suggests that the alteration in ketogenic capacity in the former group is not merely due to a higher glycolytic flux. 5. It is concluded that livers from rats at peak lactation have a lower capacity to produce ketone bodies from long-chain fatty acids which is due to an alteration in the partitioning of long-chain acyl-CoA esters between the pathways of triacylglycerol synthesis and beta-oxidation. The physiological relevance of this finding is discussed.

Inhibition of glycogen synthase (casein) kinase-1 by divalent metal ions

1988 ◽  
Vol 66 (3) ◽  
pp. 238-243 ◽  
Author(s):  
Toolsee J. Singh
Keyword(s):  
Metal Ions ◽  
Kinase Activity ◽  
Glycogen Synthase ◽  
Divalent Metal ◽  
Casein Kinase ◽  
The Other ◽  
Divalent Metal Ions ◽  
Casein Kinase 1 ◽  
Competitive Inhibitors ◽  
Noncompetitive Inhibitors

The specificity of glycogen synthase (casein) kinase-1 (CK-1) for different divalent metal ions was explored in this study. Of nine metal ions (Mg2+, Mn2+, Zn2+, Cu2+, Ca2+, Ba2+, Ni2+, Co2+, Fe2+) tested, only Mg2+ supported significant kinase activity. Several of the other metals, however, inhibited the Mg2+-stimulated kinase activity. Half-maximal inhibitions by Mn2+, Zn2+, Co2+, Fe2+, and Ni2+ were observed at 55, 65, 110, 125, and 284 μM, respectively. Kinetic analyses indicate that the metal ions are acting as competitive inhibitors of CK-1 with respect to the protein substrate (casein) and as noncompetitive inhibitors with respect to the nucleotide substrate (ATP). The inhibition of CK-1 by the different metal ions can be reversed by EGTA.

scholarly journals Effects of vanadate on protein kinases in rat hepatocytes

1989 ◽  
Vol 262 (2) ◽  
pp. 563-567 ◽  
Author(s):  
C Villar-Palasi ◽  
J J Guinovart ◽  
A M Gómez-Foix ◽  
J E Rodriguez-Gil ◽  
F Bosch
Keyword(s):  
Protein Kinase ◽  
Cyclic Amp ◽  
Kinase Activity ◽  
Activity Ratio ◽  
Glycogen Synthase ◽  
Rat Hepatocytes ◽  
Casein Kinase ◽  
Optimal Time ◽  
Dependent Protein Kinase ◽  
Dependent Protein

In rat hepatocytes, vanadate modifies neither the intracellular concentration of cyclic AMP nor the -cyclic AMP/+cyclic AMP activity ratio for cyclic AMP-dependent protein kinase. Vanadate can, however, counteract the increase in cyclic AMP and the increase in the -cyclic AMP/+cyclic AMP activity ratio of cyclic AMP-dependent protein kinase induced by glucagon. On the other hand, vanadate treatment of hepatocytes can produce a time- and concentration-dependent increase in cyclic AMP- and Ca2+-independent casein kinase activity. Maximal activation at the optimal time with 5 mM-vanadate was about 70% over control. A clear relationship was observed between the activation of casein kinase and the inactivation of glycogen synthase after vanadate treatment. These results suggest that casein kinase activity may be involved in vanadate actions in rat hepatocytes.

scholarly journals Characterization of the liver P2-purinoceptor involved in the activation of glycogen phosphorylase

1986 ◽  
Vol 240 (2) ◽  
pp. 367-371 ◽  
Author(s):  
S Keppens ◽  
H De Wulf
Keyword(s):  
Binding Sites ◽  
Glycogen Phosphorylase ◽  
Plasma Membranes ◽  
Specific Binding ◽  
Rank Order ◽  
Rat Hepatocytes ◽  
Isolated Hepatocytes ◽  
Isolated Rat Hepatocytes ◽  
P2 Purinoceptors

Evidence has been presented for the existence in rat liver of P2-purinoceptors which are involved in the control of glycogenolysis. Isolated rat hepatocytes and purified liver plasma membranes have been used to study the binding of the ATP analogue adenosine 5′-[alpha- [35S]thio]triphosphate (ATP alpha [35S]) to these postulated P2-purinoceptors. The nucleotide analogue behaves as a full agonist for the activation of glycogen phosphorylase in isolated hepatocytes, 0.3 microM being required for half-maximal activation. Specific binding of ATP alpha [35S] to hepatocytes and plasma membranes occurs within 1 min and is essentially reversible. The analysis of the dose-dependency at equilibrium indicates the presence of binding sites with Kd of 0.23 microM with hepatocytes and Kd of 0.11 microM with plasma membranes. The relative affinities of 10 nucleotide analogues were deduced from competition experiments for ATP alpha [35S] binding to hepatocytes, and these correlated highly with their biological activity (activation of glycogen phosphorylase in hepatocytes). For all the agonists, binding occurs in the same concentration range as the biological effect. These data clearly suggest that the detected binding sites correspond to the physiological P2-purinoceptors involved in the regulation of liver glycogenolysis. The rank order of potency of some ATP analogues suggests that liver possesses the P2Y-subclass of P2-purinoceptors.

Preserved GLP-I Effects on Glycogen Synthase a Activity and Glucose Metabolism in Isolated Hepatocytes and Skeletal Muscle From Diabetic Rats

Diabetes ◽  
1997 ◽  
Vol 46 (8) ◽  
pp. 1264-1269 ◽  
Author(s):  
M. Morales ◽  
M. I. Lopez-Delgado ◽  
A. Alcantara ◽  
M. A. Luque ◽  
F. Clemente ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Glucose Metabolism ◽  
Glycogen Synthase ◽  
Isolated Hepatocytes ◽  
Diabetic Rats

scholarly journals Prostaglandins E2 and F2α increase fructose 2,6-bisphosphate levels in isolated hepatocytes

1991 ◽  
Vol 274 (1) ◽  
pp. 309-312 ◽  
Author(s):  
A M Gómez-Foix ◽  
J E Rodríguez-Gil ◽  
J J Guinovart ◽  
F Bosch
Keyword(s):  
Rat Hepatocytes ◽  
Isolated Hepatocytes ◽  
Glycolytic Flux ◽  
Dependent Manner ◽  
Hepatic Glucose Metabolism ◽  
Prostaglandin F2 Alpha ◽  
Hepatic Glucose ◽  
Dose Dependent ◽  
Prostaglandins E2 And F2α ◽  
Stimulation Of

In hepatocytes isolated from fed rats, prostaglandin E2 (PGE2) and prostaglandin F2 alpha (PGF2 alpha) increased, in a time- and dose-dependent manner, fructose 2,6-bisphosphate [Fru(2,6)P2] levels and stimulated the glycolytic flux. The rise in Fru(2,6)P2 was related to an increase in glucose 6-phosphate levels which resulted from the stimulation of glycogenolysis. In cells obtained from 24 h-starved rats, no effects of either PGE2 or PGF2 alpha could be observed. In addition, when the stimulation of glycogenolysis was abolished by incubation of fed-rat hepatocytes in a Ca2(+)-depleted medium, Fru(2,6)P2 levels did not increase. Furthermore, no effects of PGs on 6-phosphofructo-2-kinase activity could be observed. These results indicate that PGE2 and PGF2 alpha show similar actions to Ca2(+)-dependent hormones on hepatic glucose metabolism.

Characterization of glucagon and catecholamine effects on isolated sheep hepatocytes

1988 ◽  
Vol 255 (4) ◽  
pp. R539-R546
Author(s):  
C. Morand ◽  
C. Yacoub ◽  
C. Remesy ◽  
C. Demigne
Keyword(s):  
Rat Liver ◽  
Glycogen Phosphorylase ◽  
Plasma Membranes ◽  
Rat Hepatocytes ◽  
Isolated Hepatocytes ◽  
Cyclic Monophosphate ◽  
Sheep Liver ◽  
Beta Agonists ◽  
Rat Liver Cells

The purpose of this study was to characterize the glycogenolytic response to catecholamines and glucagon in isolated sheep hepatocytes. In this species, epinephrine appeared to exert its action on hepatic glycogenolysis by altering the cytosolic concentrations of both adenosine 3',5'-cyclic monophosphate (cAMP) and Ca2+. In contrast to results obtained in rat hepatocytes, glucagon failed to induce a rise in free cytosolic Ca2+ in sheep liver. Experiments on isolated hepatocytes or on liver plasma membranes showed that in sheep, glucagon was more efficient than epinephrine in promoting the production of cAMP. In the presence of glucagon or epinephrine, the activation of the glycogen phosphorylase a always appeared greater in sheep than in rat liver cells, whereas the variations in cellular cAMP were quite limited in sheep. The alpha 1- and beta-agonists (phenylephrine and isoproterenol) were alone as efficient as epinephrine in promoting phosphorylase a activation in sheep hepatocytes. All these results indicate the existence in sheep liver of a glycogen phosphorylase highly responsive to hormones.

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