Effect of cobalt on the liver glycogen content in the streptozotocin-induced diabetic rats

Cobalt decreases blood glucose in diabetic rats but the mechanisms involved are unclear. To determine the contribution of glycogen metabolism to glycemia-lowering effect, glycogen contents of liver and muscle in the streptozotocin-induced diabetic rats were determined. The liver glycogen was depleted in diabetic rats. But when cobalt was administered to the rats, the glycogen returned to the level of healthy rats, concomitantly with the decrease in blood glucose. The cobalt treatment had no effect on the muscle glycogen in the diabetic rats. The tissue-specific responses of glycogen metabolism suggest the involvement of suppressed glucagon signaling due to cobalt treatment.     

Glycogen synthase,glycogen phosphorylase and α-amylase activity in homogenates of islets of GK rats: Comparison with hepatic and pancreatic extracts

Glycogen accumulation in pancreatic islet cells in situations of sustained hyperglycaemia may participate in the phenomenon of so-called B-cell glucotoxicity. Unexpectedly, however, previously little if any glycogen was found in islet cells of non-insulin-dependent diabetic Goto-Kakizaki rats (GK rats). Therefore, the activities of glycogen synthase, glycogen phosphorylase and α-amylase were measured in islets of control and GK rats. No significant difference in enzymatic activity was observed between the control and diabetic animals. In the liver, the activity of glycogen synthase appeared even somewhat higher in GK rats than in control animals. It is concluded that the diabetic syndrome in the GK rats does not involve any major anomaly of glycogen synthase and glycogen phosphorylase activity in the liver of these animals, as well as α-amylase, in pancreatic islets.     

Post-exercise ketosis and the glycogen content of liver and muscle in rats on a high carbohydrate diet

Post-exercise ketosis is known to be suppressed by physical training and by a high carbohydrate diet. As a result it has often been presumed, but not proven, that the development of post-exercise ketosis is closely related to the glycogen content of the liver. We therefore studied the effect of 1 h of treadmill running on the blood 3-hydroxybutyrate and liver and muscle glycogen concentrations of carbohydrate-loaded trained (n = 72) and untrained rats (n = 72). Resting liver and muscle glycogen levels were 25%-30% higher in the trained than in the untrained animals. The resting 3-hydroxybutyrate concentrations of both groups of rats were very low: less than 0.08 mmol.l-1. Exercise did not significantly influence the blood 3-hydroxybutyrate concentrations of trained rats, but caused a marked post-exercise ketosis (1.40 +/- 0.40 mmol.l-1 h after exercise) in the untrained animals, the time-course of which was the approximate inverse of the changes in liver glycogen concentration. Interpreting the results in the light of similar data obtained after a normal and low carbohydrate diet it has been concluded that trained animals probably owe their relative resistance to post-exercise ketosis to their higher liver glycogen concentrations as well as to greater peripheral stores of mobilizable carbohydrate.     

In vivo effects of vanadate on hepatic glycogen metabolizing and lipogenic enzymes in insulin-dependent and insulin-resistant diabetic animals

The insulin-mimetic action of vanadate is well established but the exact mechanism by which it exerts this effect is still not clearly understood. The role of insulin in the regulation of hepatic glycogen metabolizing and lipogenic enzymes is well known. In our study, we have, therefore, examined the effects of vanadate on these hepatic enzymes using four different models of diabetic and insulin-resistant animals. Vanadate normalized the blood glucose levels in all animal models. In streptozotocin-induced diabetic rats, the amount of liver glycogen and the activities of the active-form of glycogen synthase, both active and inactive-forms of phosphorylase, and lipogenic enzymes like glucose 6-phosphate dehydrogenase and malic enzyme were decreased and vanadate treatment normalized all of these to near normal levels. The other three animal models (db/db mouse, sucrose-fed rats and fa/fa obese Zucker rats) were characterized by hyperinsulinemia, hypertriglyceridemia, increases in activities of lipogenic enzymes, and marginal changes in glycogen metabolizing enzymes. Vanadate treatment brought all of these values towards normal levels. It should be noted that vanadate shows differential effects in the modulation of lipogenic enzymes activities in type I and type II diabetic animals. It increases the activities of lipogenic enzymes in streptozotocin-induced diabetic animals and prevents the elevation of activities of these enzymes in hyperinsulinemic animals. The insulin-stimulated phosphorylation of insulin receptor subunit and its tyrosine kinase activity was increased in streptozotocin-induced diabetic rats after treatment with vanadate. Our results support the view that insulin receptor is one of the sites involved in the insulin-mimetic actions of vanadate.     

Effects of vitamin C on muscle glycogen and oxidative events in experimental diabetes

Streptozotocin (STZ) is an agent used in creating experimental diabetes. Varying findings have been reported about the striated muscle glycogen levels in diabetes. In this study, it was planned to observe interaction of vitamin C (AA), of which deficiency has been shown in diabetics, with soleus muscle glycogen levels and oxidative events on STZ-diabetic subjects. Material and Method: In the study, 38 male adult Wistar Albino rats with weights 200 ± 20 g were used by separating them into four groups: Control, Vitamin C, Diabetes, Diabetes + Vitamin C. Body weights and fasting blood glucose were measured at the beginning and end of the experiment. AA, TBARS, GSH, NOx and glycogen levels of soleus muscles, and AA level of blood were measured. The results were compared using Anova variance and Mann-Whitney U tests. Results showed that AA levels in blood increased with vitamin C administration; AA, GSH and NOx levels in the muscle were low and MDA and glycogen levels were high in diabetics; and that vitamin C in the given dosage partially corrected these values. These results indicate that higher dosage than daily 20 mg/kg Vitamin C is required for being effective on metabolic and oxidizing events in diabetic rats.     

Heart glycogen content and isoprenaline-induced myocardial lesions

The role of glycogen content in the heart for the development of isoprenaline-induced myocardial lesions (IML) was studied in Wistar rats and in two inbred rat strains: In IR rats (resistant to the development of IML) and in IS rats (sensitive to IML development).Glycogen content in the heart can be dramatically lowered or increased by various interventions. IML develop during the period of very low heart glycogen content (about 0.6 mg.g^–1) induced by isoprenaline administration. In animals with increased resistance to IML, either due to genetic factors or induced by isoprenaline pretreatment a high glycogen content in the heart is found (up to 7.5 mg.g^–1).The increase of resistance to IML development and increased glycogen content induced by isoprenaline pretreatment were accompanied by lower basal or ISO-, guanylylimidodiphosphate- (Gpp/NH/p) and forskolin-stimulated activities of adenylyl cyclase. On the other hand, these parameters did not differ between IR and IS rats in spite of the presence of significant differences in the resistance to the development of IML and in heart glycogen content in these two rats strains.These results suggest that genetically determined differences between two inbred rat strains in the resistance of the heart to the development of IML and in the heart glycogen content are caused by factors which are independent of the receptor-adenylyl cyclase complex and are therefore different from those involved in the increase of resistance and glycogen content due to isoprenaline pretreatment.     

Liver glycogen content and activity of several hepatic enzymes in repeatedly immobilized rats

Liver glycogen content and activity of several hepatic enzymes in repeatedly immobilized rats. Acta Physiol. Pol., 1978, 29 (1): 55-60. In adult, male rats subjected to immobilization for 2.5 h daily, on the 7th day, 24 h after a sixth trial, an increased activity of the liver aspartate aminotransferase was found as shown by comparing their data with those of normally fed controls and also of the animals pair fed to the immobilized. Moreover, a net increasing effect of daily immobilization on the liver glycogen content was described.     

Purification and partial characterization of glycogen synthase kinase-3 from rabbit liver

Summary Glycogen synthase kinase-3 (GSK-3) was purified from rabbit liver to homogeneity by ultracentrifugation, ion-exchange chromatography on DEAE-cellulose, Cellulose phosphate, CM-Sephadex and Fast Protein Liquid Chromatography (FPLC) on Mono-S column. The enzyme was purified approximately 20,000 fold with an approximate 2% recovery. The purified enzyme showed a single band on SDS-polyacrylamide gel electrophoresis. GSK-3 is a monomeric enzyme with a molecular weight of 50,000–52,000 as derived from SDS-polyacrylamide gel electrophoresis and gel filtration. The purified enzyme was indeed a GSK-3 since it phosphorylated three sites, i.e., 3a, 3b, and 3c on liver glycogen synthase. GSK-3 incorporated up to 2.6 mol Pi/mol glycogen synthase subunit with a concomitant inactivation of glycogen synthase activity.     

Pancreatic fate of D-[3H] mannoheptulose

D-Mannoheptulose was recently postulated to be transported into cells by GLUT2. The validity of such an hypothesis was assessed by comparing the uptake of tritiated D-mannoheptulose by pancreatic islets versus pieces of pancreas and, in the latter case, by comparing results obtained in control rats versus animals injected with streptozotocin (STZ). The uptake of D-[3H] mannoheptulose by islets represents a time-related and temperature-sensitive process, inhibited by cytochalasin B and enhanced by D-glucose. The uptake of the tritiated heptose was much lower in pieces of pancreatic tissue and inhibited by D-glucose, at least in the STZ rats. Whether in pieces of pancreas exposed in vitro to D-[3H] mannoheptulose or after intravenous injection of the tritiated heptose, the radioactive content of the pancreatic tissue was lower in STZ rats than in control animals. This contrasted with an unaltered radioactive content of liver and muscle in the STZ rats, at least when treated with insulin. Suitably radiolabelled D-mannoheptulose or an analogue of the heptose could thus conceivably be used for quantification of the endocrine pancreatic mass.     

Determination of glycogen and enzymes of glycogen metabolism in human hair follicles

The skin epithelium and its organelles use glycogen as well as glucose as source of energy. Therefore the characterisation of glycogen metabolism and the enzymes involved is important in the study of mechanisms regulating the normal or abnormal differentiation of skin organelles such as sebaceous glands and hair follicles.The present paper describes fluorimetric methods for the determination of glycogen and for the measurements of phosphorylase and phosphorylase kinase activity in one and the same lysate of minute tissue samples. The methods were tested for their suitability on freshly isolated human hair follicles and cultured hair follicle cells. The possible use of these techniques for studies on the pathophysiology of acne and hirsutism is discussed.     

Observations on the role of smooth endoplasmic reticulum in glucocorticoid-induced hepatic glycogen deposition

We have studied by quantitative electron microscopy the relationship of specific hepatic cellular organelles to glycogen synthesis using dexamethasone, a potent synthetic glucocorticoid, to induce glycogen deposition in livers of adrenalectomized rats. Chemical and ultrastructural glycogen determinations revealed that the livers of fasted adrenalectomized rats had very low glycogen levels. Dexamethasone caused a time-related increase in hepatic glycogen which was the result of increases in the number of hepatocytes depositing glycogen and the amount of glycogen in each cell. The surface density of smooth endoplasmic reticulum (SER) in centrilobular and periportal hepatocytes also increased after treatment with dexamethasone; this increase preceded glycogen deposition. The newly deposited glycogen was spatially associated with membranes of SER, and a continued increase in SER surface density was correlated temporally with the increasing glycogen volume density. In both centrilobular and periportal hepatocytes, the suface density of rough endoplasmic reticulum (RER) initially decreased after dexamethasone administration but later increased. These data support the hypothesis that dexamethasone-induced enhancement of SER is functionally associated with the increase in glycogen, and that although the initial increase in SER may occur through transformation of RER to SER, later increases in SER require synthesis of new membranes.     

Skeletal muscle glycogen content, structure, and metabolism are normal in rats with hepatic glycogen phosphorylase kinase deficiency

Skeletal muscle glycogen content and structure, and the activities of several enzymes of glycogen metabolism are reported for the hepatic glycogen phosphorylase b kinase deficient (gsd/gsd) rat. The skeletal muscle glycogen content of the fed gsd/gsd rat is 0.50 +/- 0.11% tissue wet weight, and after 40 hours of starvation this value is lowered 40% to 0.30 +/- 0.05% tissue wet weight. In contrast the gsd/gsd rat liver has an elevated glycogen content which remains high after starvation. The skeletal muscle phosphorylase b kinase, glycogen phosphorylase, glycogen synthase and acid alpha-glucosidase activities are 17.2 +/- 2.9 units/g tissue, 119.9 +/- 6.4 units/g tissue, 12.2 +/- 0.4 units/g tissue and 1.4 +/- 0.4 milliunits/g tissue, respectively, with approx. 20% of phosphorylase and approx. 24% of synthase in the active form (at rest). These enzyme activities resemble those of Wistar skeletal muscle, and again this contrasts with the situation in the liver where there are marked differences between the Wistar and the gsd/gsd rat. Fine structural analysis of the purified glycogen showed resemblance to other glycogens in branching pattern. Analysis of the molecular weight distribution of the purified glycogen indicated polydispersity with approx. 66% of the glycogen having a molecular weight of less than 250 X 10(6) daltons and approx. 25% greater than 500 X 10(6) daltons. This molecular weight distribution resembles those of purified Wistar liver and skeletal muscle glycogens and differs from that of the gsd/gsd liver glycogen which has an increased proportion of the low molecular weight material.(ABSTRACT TRUNCATED AT 250 WORDS)     

Hypoglycemic and antihyperglycemic effect of Brassica juncea diet and their effect on hepatic glycogen content and the key enzymes of carbohydrate metabolism

Plants in the form of food or other remedial forms have been used for the treatment of various human ailments. Diabetes is one such disorder in which various form of plants and herbal remedies have been tried. Brassica juncea (BJ) seeds (Rai) are consumed in India as a spice in various food items. Previous studies have shown the hypoglycemic effect of this plant in rats. The present study was undertaken to study the hypoglycemic and anti-hyperglycemic of various strengths (5, 10 and 15%) of BJ seed diet in alloxan and streptozotocin induced diabetes in albino rats. In addition, key enzymes of carbohydrate metabolism (glucokinase – EC 2.7.1.1, hexokinase – EC 2.7.1.1, and phosphofructokinase – EC 2.7.1.11) and hepatic glycogen content was also assessed to understand the mechanism of action. BJ diet (10 and 15%) showed significant anti-hyperglycemic effect in alloxan (35 mg/kg) but not in STZ (60 mg/kg) rats. It also failed to modulate the hepatic glycogen content and enzyme activities.     

Mechanism of glycogen supercompensation in rat skeletal muscle cultures

A model to study glycogen supercompensation (the significant increase in glycogen content above basal level) in primary rat skeletal muscle culture was established. Glycogen was completely depleted in differentiated myotubes by 2 h of electrical stimulation or exposure to hypoxia during incubation in medium devoid of glucose. Thereafter, cells were incubated in medium containing glucose, and glycogen supercompensation was clearly observed in treated myotubes after 72 h. Peak glycogen levels were obtained after 120 h, averaging 2.5 and 4 fold above control values in the stimulated- and hypoxia-treated cells, respectively. Glycogen synthase activity increased and phosphorylase activity decreased continuously during 120 h of recovery in the treated cells. Rates of 2-deoxyglucose uptake were significantly elevated in the treated cells at 96 and 120 h, averaging 1.4–2 fold above control values. Glycogenin content increased slightly in the treated cells after 48 h (1.2 fold vs. control) and then increased considerably, achieving peak values after 120 h (2 fold vs. control). The results demonstrate two phases of glycogen supercompensation: the first phase depends primarily on activation of glycogen synthase and inactivation of phosphorylase; the second phase includes increases in glucose uptake and glycogenin level.     

Hepatic glycogen metabolism in the db/db mouse

Summary Knowledge of the metabolic changes that occur in insulin-resistant type 2 diabetes is relatively lacking compared to insulin-deficient type 1 diabetes. This paper summarizes the importance of the C57BL/KsJ-db/db mouse as a model of type 2 diabetes, and illustrates the effects that insulin-deficient and insulin-resistant states have on hepatic glycogen metabolism. A longitudinal study of db/db mice of ages 2–15 weeks revealed that significant changes in certain parameters of hepatic glycogen metabolism occur during this period. The liver glycogen levels were similar between diabetic and control mice. However, glycogen particles from db/db mice were on average smaller in mass and had shorter exterior and interior chain lengths. Total phosphorylase and phosphorylase a activities were elevated in the genetically diabetic mice. This was primarily due to an increase in the amount of enzymic protein apparently the result of a decreased rate of degradation. It was not possible to find a consistent alteration in glycogen synthase activity in the db/db mice. Glycogen synthase and phosphorylase from diabetic liver revealed some changes in kinetic properties in the form of a decrease in V_max, and altered sensitivity to inhibitors like ATP. The altered glycogen structure in db/db mice may have contributed to changes in the activities and properties of glycogen synthase and phosphorylase. The exact role played by hormones (insulin and glucagon) in these changes is not clear but further studies should reveal their contributions. The db/db mouse provides a good model for type 2 diabetes and for fluctuating insulin and glucagon ratios. Its use should clarify the regulation of hepatic glycogen metabolism and other metabolic processes known to be controlled by these hormones. The other animal models of type 2 diabetes, ob/ob mouse and fatty Zucker (fa/fa) rat, show similar impairment of hepatic glycogen metabolism. The concentrations of glycogen metabolizing enzymes are high and in vitro studies indicate enhanced rate of glycogen synthesis and breakdown. However, streptozotocin-induced diabetic animals and BB rats which resemble insulin-deficient type 1 diabetes are characterized by decreased glycogen turnover as a result of reduction in the levels of glycogen metabolizing enzymes.     

Alzheimer-like changes in protein kinase B and glycogen synthase kinase-3 in rat frontal cortex and hippocampus after damage to the insulin signalling pathway

The insulin-resistant brain state is related to late-onset sporadic Alzheimer's disease, and alterations in the insulin receptor (IR) and its downstream phosphatidylinositol-3 kinase signalling pathway have been found in human brain. These findings have not been confirmed in an experimental model related to sporadic Alzheimer's disease, for example rats showing a neuronal IR deficit subsequent to intracerebroventricular (i.c.v.) treatment with streptozotocin (STZ). In this study, western blot analysis performed 1 month after i.c.v. injection of STZ showed an increase of 63% in the level of phosphorylated glycogen synthase kinase-3alpha/beta (pGSK-3alpha/beta) protein in the rat hippocampus, whereas the levels of the unphosphorylated form (GSK-3alpha/beta) and protein kinase B (Akt/PKB) remained unchanged. Three months after STZ treatment, pGSK-3alpha/beta and Akt/PKB levels tended to decrease (by 8 and 9% respectively). The changes were region specific, as a different pattern was found in frontal cortex. Structural alterations were also found, characterized by beta-amyloid peptide-like aggregates in brain capillaries of rats treated with STZ. Similar neurochemical changes and cognitive deficits were recorded in rats treated with i.c.v. 5-thio-d-glucose, a blocker of glucose transporter (GLUT)2, a transporter that is probably involved in brain glucose sensing. The IR signalling cascade alteration and its consequences in rats treated with STZ are similar to those found in humans with sporadic Alzheimer's disease, and our results suggest a role for GLUT2 in Alzheimer's pathophysiology.     

Amino acid anthranilamide derivatives as a new class of glycogen phosphorylase inhibitors

A series of amino acid anthranilamide derivatives identified from a high-throughput screening campaign as novel, potent, and glucose-sensitive inhibitors of human liver glycogen phosphorylase a are described. A solid-phase synthesis using Wang resin was also developed which provided efficient access to a variety of analogues, and resulted in the identification of key structure–activity relationships, and the discovery of a potent exemplar (IC₅₀ = 80 nM). The SAR scope, synthetic strategy, and in vitro results for this series are presented herein.     

Basal metabolic rate and lipid and liver glycogen in mice infected by the nematode Nematospiroides dubius

The basal metabolic rate (BMR) of mice losing weight about two weeks after infection by Nematospiroides dubius was lower than that of uninfected mice gaining weight when fed ad libitum or losing weight on quantitatively reduced rations. There was no difference in BMR between the latter two groups.Following the injection of ¹⁴C-glucose, the high specific activity of expired CO₂ from infected and reduced ration mice was considered to be due to the utilization of energy reserves. The levels of lipid and liver glycogen were low in these two groups of mice and their specific activities, particularly in the severely affected animals, were high.It was concluded that the depressed BMR of infected mice is unrelated to anorexia, which did, however, explain the low levels of lipid and liver glycogen.     

Reciprocal regulation of glycogen phosphorylase and glycogen synthase by insulin involving phosphatidylinositol-3 kinase and protein phosphatase-1 in HepG2 cells

The effect of insulin on glycogen synthesis and key enzymes of glycogen metabolism, glycogen phosphorylase and glycogen synthase, was studied in HepG2 cells. Insulin stimulated glycogen synthesis 1.83-3.30 fold depending on insulin concentration in the medium. Insulin caused a maximum of 65% decrease in glycogen phosphorylase 'a' and 110% increase in glycogen synthase activities in 5 min. Although significant changes in enzyme activities were observed with as low as 0.5 nM insulin level, the maximum effects were observed with 100 nM insulin. There was a significant inverse correlation between activities of glycogen phosphorylase 'a' and glycogen synthase 'a' (R² = 0.66, p < 0.001). Addition of 30 mM glucose caused a decrease in phosphorylase 'a' activity in the absence of insulin and this effect was additive with insulin up to 10 nM concentration. The inactivation of phosphorylase 'a' by insulin was prevented by wortmannin and rapamycin but not by PD98059. The activation of glycogen synthase by insulin was prevented by wortmannin but not by PD98059 or rapamycin. In fact, PD98059 slightly stimulated glycogen synthase activation by insulin. Under these experimental conditions, insulin decreased glycogen synthase kinase-3 activity by 30-50% and activated more than 4-fold particulate protein phosphatase-1 activity and 1.9-fold protein kinase B activity; changes in all of these enzyme activities were abolished by wortmannin. The inactivation of GSK-3 and activation of PKB by insulin were associated with their phosphorylation and this was also reversed by wortmannin. The addition of protein phosphatase-1 inhibitors, okadaic acid and calyculin A, completely abolished the effects of insulin on both enzymes. These data suggest that stimulation of glycogen synthase by insulin in HepG2 cells is mediated through the PI-3 kinase pathway by activating PKB and PP-1G and inactivating GSK-3. On the other hand, inactivation of phosphorylase by insulin is mediated through the PI-3 kinase pathway involving a rapamycin-sensitive p70^s6k and PP-1G. These experiments demonstrate that insulin regulates glycogen phosphorylase and glycogen synthase through (i) a common signaling pathway at least up to PI-3 kinase and bifurcates downstream and (ii) that PP-1 activity is essential for the effect of insulin.