Insulin regulation of glycogen synthase phosphatase in primary cultures of hepatocytes

Activation of glycogen synthase in the perfused rat liver is defective in severely diabetic rats. In the present study, activation of glycogen synthase by glucose and increased incorporation of [14C]glucose into glycogen by insulin are defective in hepatocytes isolated from alloxan diabetic rats. Acute activation of glycogen synthase in hepatocytes isolated from diabetic rats was restored by treatment of the rats with insulin in vivo. Restoration of synthase activation was not achieved by incubation of hepatocytes in the presence of insulin in vitro for up to 12 h. When isolated hepatocytes from diabetic rats were placed in primary culture in a serum-free defined medium over a 3-day period, glycogen synthesis was partially restored by cortisol and triiodothyronine and dramatically increased by insulin. Concomitant with restoration of [14C]glycogen synthesis was an insulin-mediated increase in glycogen synthase I and synthase phosphatase activity. Restoration of regulation of glycogen synthesis in primary cultures of hepatocytes from diabetic rats by insulin required the presence of cortisol and triiodothyronine. Primary cultures of hepatocytes from normal rats did not require triiodothyronine for insulin to effect glycogenesis over a 3-day period. These data demonstrate that insulin acts in a chronic manner in concert with other hormones to control synthase phosphatase activity, an effect which may be influencing acute control of hepatic glycogen synthesis.     

Adaptations of glycogen metabolism in rat epididymal adipose tissue during fasting and refeeding.

It is well documented that adipose tissue glycogen content decreases during fasting and increases above control during refeeding. We now present evidence that these fluctuations result from adaptations intrinsic to adipose tissue glycogen metabolism that persist in vitro: in response to insulin (1 milliunit/ml), [3H]glucose incorporation into rat fat pad glycogen was reduced to 10% of control after a 3-day fast; incorporation increased 6-fold over fed control on the 4th day of refeeding following a 3-day fast. We have characterized this adaptation with regard to alterations in glycogen synthase and phosphorylase activity. In addition, we found that incubation of fat pads from fasted rats with insulin (1 milliunit/ml) increased glucose-6-P content, indicating that glucose transport was not the rate-limiting step for glucose incorporation into glycogen in the presence of insulin. In contrast, feeding a fat-free diet resulted in dramatic increases in glycogen content of fat pads without a concomitant increase in glucose incorporation into glycogen in response to insulin (1 milliunit/ml). Thus, fasting and refeeding appeared to alter insulin action on adipose tissue glycogen metabolism more than this dietary manipulation.     

Insulin-dependent glycogen synthesis is delayed in onset in the skeletal muscle of food-deprived aged rats

Insulin resistance with aging may be responsible for impaired glycogen synthesis in the skeletal muscle of aged rats and contribute to the well-known decreased ability to respond to stress with aging. For this reason, to assess the ability of the skeletal muscle to utilize glucose for glycogen synthesis during aging, the time course of glycogen synthesis was continuously monitored by 13C nuclear magnetic resonance for 2 h in isolated [13C] glucose-perfused gastrocnemius-plantaris muscles of 5-day food-deprived adult (6-8 months; n=10) or 5-day food-deprived aged (22 months; n=8) rats. [13C] glucose (10 mmol/L) perfusion was carried out in the presence or absence of an excess of insulin (1 micromol/L). Food deprivation only decreased glycogen level in adult rats (8.9+/-2.4 micromol/g in adults vs. 35.6+/-2.4 micromol/g in aged rats; P<.05). In the presence of an excess of insulin, muscle glycogen synthesis was stimulated in both adult and aged muscles, but the onset was delayed with aging (40 min later). In conclusion, this study highlights the important role of glycogen depletion in stimulating glycogen synthesis in muscles. Consequently, the absence of glycogen depletion in response to starvation in aged rats may be the origin of the delay in insulin-stimulated glycogen synthesis in the skeletal muscle. Glycogen synthesis clearly was not impaired with aging.     

Insulin resistance in soleus muscle from obese Zucker rats. Involvement of several defective sites

1. The effect of insulin upon glucose transport and metabolism in soleus muscles of genetically obese (fa/fa) and heterozygote lean Zucker rats was investigated at 5–6 weeks and 10–11 weeks of age. Weight-standardized strips of soleus muscles were used rather than the intact muscle in order to circumvent problems of diffusion of substrates. 2. In younger obese rats (5–6 weeks), plasma concentrations of immunoreactive insulin were twice those of controls, whereas their circulating triacylglycerol concentrations were normal. Insulin effects upon 2-deoxyglucose uptake and glucose metabolism by soleus muscles of these rats were characterized by both a decreased sensitivity and a decrease in the maximal response of this tissue to the hormone. 3. In older obese rats (10–11 weeks), circulating concentrations of insulin and triacylglycerols were both abnormally elevated. A decrease of 25–35% in insulin-binding capacity to muscles of obese rats was observed. The soleus muscles from the older obese animals also displayed decreased sensitivity and maximal response to insulin. However, at a low insulin concentration (0.1m-i.u./ml), 2-deoxyglucose uptake by muscles of older obese rats was stimulated, but such a concentration was ineffective in stimulating glucose incorporation into glycogen, and glucose metabolism by glycolysis. 4. Endogenous lipid utilization by muscle was calculated from the measurements of O₂ consumption, and glucose oxidation to CO₂. The rate of utilization of fatty acids was normal in muscles of younger obese animals, but increased in those of the older obese rats. Increased basal concentrations of citrate, glucose 6-phosphate and glycogen were found in muscles of older obese rats and may reflect intracellular inhibition of glucose metabolism as a result of increased lipid utilization. 5. Thus several abnormalities are responsible for insulin resistance of muscles from obese Zucker rats among which we have observed decreased insulin binding, decreased glucose transport and increased utilization of endogenous fatty acid which could inhibit glucose utilization.     

The evolution of insulin resistance in muscle of the glucose infused rat

Glucose infusion into rats causes skeletal muscle insulin resistance that initially occurs without changes in insulin signaling. The aim of the current study was to prolong glucose infusion and evaluate other events associated with the transition to muscle insulin resistance. Hyperglycemia was produced in rats by glucose infusion for 3, 5 and 8 h. The rate of infusion required to maintain hyperglycemia was reduced at 5 and 8 h. Glucose uptake into red quadriceps (RQ) and its incorporation into glycogen decreased between 3 and 5 h, further decreasing at 8 h. The earliest observed change in RQ was decreased AMPKα2 activity associated with large increases in muscle glycogen content at 3 h. Activation of the mTOR pathway occurred at 5 h. Akt phosphorylation (Ser⁴⁷³) was decreased at 8 h compared to 3 and 5, although no decrease in phosphorylation of downstream GSK-3β (Ser⁹) and AS160 (Thr⁶⁴²) was observed. White quadriceps showed a similar but delayed pattern, with insulin resistance developing by 8 h and decreased AMPKα2 activity at 5 h. These results indicate that, in the presence of a nutrient overload, alterations in muscle insulin signaling occur, but after insulin resistance develops and appropriate changes in energy/nutrient sensing pathways occur.     

The metabolism of glucose in diaphragm muscle from normal rats, from streptozotocin-treated diabetic rats and from rats treated with anti-insulin serum

1. The metabolism of [U-¹⁴C]glucose by the isolated diaphragm muscle of normal rats, rats rendered diabetic with streptozotocin and rats with transitory insulin deficiency after an injection of anti-insulin serum was studied. 2. The incorporation of [¹⁴C]glucose into glycogen and oligosaccharides was significantly decreased in the diabetic diaphragm muscle and in the muscle from rats treated with anti-insulin serum. 3. Neither diabetes nor transitory insulin deficiency influenced the oxidation of glucose, or the formation of lactate and hexose phosphate esters from glucose. 4. Insulin fully restored the incorporation of glucose into glycogen and maltotetraose in the diabetic muscle, but the incorporation into oligosaccharides, although increased in the presence of insulin, was significantly lower than the values obtained with normal diaphragm in the presence of insulin.     

Sensitivity to insulin of glycolysis and glycogen synthesis of isolated soleus-muscle strips from sedentary, exercised and exercise-trained rats.

The half-maximal stimulation of the rates of glycolysis and glycogen synthesis in soleus-muscle strips from sedentary animals occurred at a concentration of insulin of about 100 microunits/ml. In soleus-muscle strips from exercise-trained rats (5 weeks of treadmill training), half-maximal stimulation of the rate of glycolysis occurred at about 10 microunits of insulin/ml, whereas that for glycogen synthesis occurred between 10 and 100 microunits of insulin/ml. The sensitivity of glycolysis to insulin after exercise training is similar to that of adipose tissue from sedentary animals. This finding suggests that, in sedentary animals, the effects of normal changes in insulin concentration may affect muscle primarily indirectly via the anti-lipolytic effect on adipose tissue, whereas after training insulin may effect the rate of glycolysis in muscle directly. A single period of exercise did not change the sensitivity of glycolysis in soleus muscle to insulin, nor probably that of glycogen synthesis. It is suggested that the improvement in insulin sensitivity of glycolysis in muscle caused by exercise-training could account, in part, for the well-established improvement in glucose tolerance and insulin sensitivity observed in man and rats after exercise-training.     

Regulation of glycogen metabolism in primary cultures of rat hepatocytes. Restoration of acute effects of insulin and glucose in cells from diabetic rats

Defects in the deposition of glycogen and the regulation of glycogen synthesis in the livers of severely insulin-deficient rats can be reversed, in vivo, within hours of insulin administration. Using primary cultures of hepatocytes isolated from normal and diabetic rats in a serum-free chemically defined medium, the present study addresses the chronic action of insulin to facilitate the direct effects of insulin and glucose on the short term regulation of the enzymes controlling glycogen metabolism. Primary cultures were maintained in the presence of insulin, triiodothyronine, and cortisol for 1-3 days. On day 1 in alloxan diabetic cultures, 10(-7) M insulin did not acutely activate glycogen synthase over a period of 15 min or 1 h, whereas insulin acutely activated synthase in cultures of normal hepatocytes. By day 3 in hepatocytes isolated from alloxan diabetic rats, insulin effected an approximate 30% increase in per cent synthase I within 15 min as was also the case for normal cells. The acute effect of insulin on synthase activation was independent of changes in phosphorylase alpha. Whereas glycogen synthase phosphatase activity could not be shown to be acutely affected by insulin, the total activity in diabetic cells was restored to normal control values over the 3-day culture period. The acute effect of 30 mM glucose to activate glycogen synthase in cultured hepatocytes from normal rats after 1 day of culture was missing in hepatocytes isolated from either alloxan or spontaneously diabetic (BB/W) rats. After 3 days in culture, glucose produced a 50% increase in glycogen synthase activity during a 10-min period under the same conditions. These studies clearly demonstrate that insulin acts in a chronic manner in concert with thyroid hormones and steroids to facilitate acute regulation of hepatic glycogen synthesis by both insulin and glucose.     

Effects of eicosapentaenoic acid (EPA) treatment on insulin sensitivity in an animal model of diabetes: improvement of the inflammatory status

In addition to decreased insulin sensitivity, diabetes is a pathological condition associated with increased inflammation. The ω-3 fatty acids have been proposed as anti-inflammatory agents. Thus, the major goal of this study was to analyze the effects of fatty acid supplementation on both insulin sensitivity and inflammatory status in an animal model of type 2 diabetes. Diabetic rats (Goto-Kakizaki model) were treated with eicosapentaenoic acid (EPA) or linoleic acid at 0.5 g/kg body weigh (bw) dose. In vivo incorporation of (14)C-triolein into adipose tissue was improved by the ω-3 administration. In vitro incubations of adipose tissue slices from EPA-treated rats showed an increase in (14)C-palmitate incorporation into the lipid fraction. These observations were linked with a decreased rate of fatty acid oxidation. EPA treatment resulted in a decreased fatty acid oxidation in incubated strips from extensor digitorum longus (EDL) muscles. The changes in lipid utilization were associated with a decrease in insulin plasma concentration, suggesting an improvement in insulin sensitivity. These changes in lipid metabolism were associated with an activation of AMP-activated protein kinase (AMPK) in white adipose tissue. In addition, EPA treatment resulted in a decreased content of peroxisome proliferator-activated receptor-α (PPARα) and PPARδ and in increased GLUT4 expression in skeletal muscle. Moreover, EPA increased 2-deoxy-D-[(14)C]glucose (2-DOG) uptake in C2C12 myotubes, suggesting an improvement in glucose metabolism. Concerning the inflammatory status, EPA treatment resulted in a decreased gene expression for both tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6) both in skeletal muscle and adipose tissue. The data suggest that EPA treatment to diabetic rats clearly improves lipid metabolism although the evidences on insulin sensitization are less clear.     

Different effects of IGF-I on insulin-stimulated glucose uptake in adipose tissue and skeletal muscle

The effect of insulin-like growth factor I (IGF-I) on insulin-stimulated glucose uptake was studied in adipose and muscle tissues of hypophysectomized female rats. IGF-I was given as a subcutaneous infusion via osmotic minipumps for 6 or 20 days. All hypophysectomized rats received L-thyroxine and cortisol replacement therapy. IGF-I treatment increased body weight gain but had no effect on serum glucose or free fatty acid levels. Serum insulin and C-peptide concentrations decreased. Basal and insulin-stimulated glucose incorporation into lipids was reduced in adipose tissue segments and isolated adipocytes from the IGF-I-treated rats. In contrast, insulin treatment of hypophysectomized rats for 7 days increased basal and insulin-stimulated glucose incorporation into lipids in isolated adipocytes. Pretreatment of isolated adipocytes in vitro with IGF-I increased basal and insulin-stimulated glucose incorporation into lipids. These results indicate that the effect of IGF-I on lipogenesis in adipose tissue is not direct but via decreased serum insulin levels, which reduce the capacity of adipocytes to metabolize glucose. Isoproterenol-stimulated lipolysis, but not basal lipolysis, was enhanced in adipocytes from IGF-I-treated animals. In the soleus muscle, the glycogen content and insulin-stimulated glucose incorporation into glycogen were increased in IGF-I-treated rats. In summary, IGF-I has opposite effects on glucose uptake in adipose tissue and skeletal muscle, findings which at least partly explain previous reports of reduced body fat mass, increased body cell mass, and increased insulin responsiveness after IGF-I treatment.     

Sensitivity of the soleus muscle to insulin in resting and exercising rats with experimental hypo- and hyper-thyroidism.

1. The effects of hypothyroidism (caused by surgical thyroidectomy followed by treatment for 1 month with propylthiouracil) and of hyperthyroidism [induced by subcutaneous administration of L-tri-iodothyronine (T3)] on glucose tolerance and skeletal-muscle sensitivity to insulin were examined in rats. Glucose tolerance was estimated during 2 h after subcutaneous glucose injection (1 g/kg body wt.). The sensitivity of the soleus muscle to insulin was studied in vitro in sedentary and acutely exercised animals. 2. Glucose tolerance was impaired in both hypothyroid and hyperthyroid rats in comparison with euthyroid controls. 3. In the soleus muscle, responsiveness of the rate of lactate formation to insulin was abolished in hypothyroid rats, whereas the sensitivity of the rate of glycogen synthesis to insulin was unchanged. In hyperthyroid animals, opposite changes were found, i.e. responsiveness of the rate of glycogen synthesis was inhibited and the sensitivity of the rate of lactate production did not differ from that in control sedentary rats. 4. A single bout of exercise for 30 min potentiated the stimulatory effect of insulin on lactate formation in hyperthyroid rats and on glycogen synthesis in hypothyroid animals. 5. The data suggest that thyroid hormones exert an interactive effect with insulin in skeletal muscle. This is likely to be at the post-receptor level, inhibiting the effect of insulin on glycogen synthesis and stimulating oxidative glucose utilization.     

Effect of trihydroxyoctadecadiene acids from Bryonia alba L. on the activity of glycogen metabolism enzymes in alloxan diabetes

It was shown previously in experiments on white rats with alloxan diabetes that trihydroxyoctadecadiene acids from Bryonia alba L. have a hypoglycemic action. The present paper is concerned with the effects of the above-indicated compounds on the activity of glycogen phosphorylase (a- and b-forms), phosphoprotein phosphatase and hexokinase in liver and muscle tissues of white rats with alloxan diabetes. One of the possible mechanisms of the hypoglycemic action of trihydroxyoctadecadiene acids is discussed.     

A novel insulin sensitizer (S15511) enhances insulin-stimulated glucose uptake in rat skeletal muscles.

Type 2 diabetes is preceded by the presence of skeletal muscle insulin resistance, and drugs that increase insulin sensitivity in skeletal muscle prevent the disease. S15511 is an original compound with demonstrated effects on insulin sensitivity in animal models of insulin resistance. However, the mechanisms behind the insulin-sensitizing effect of S15511 are unknown. The aim of our study was to explore whether S15511 improves insulin sensitivity in skeletal muscles. Insulin sensitivity was assessed in skeletal muscles from S15511-treated rats by measuring intracellular insulin-signaling activity and insulin-stimulated glucose transport in isolated muscles. In addition, GLUT4 expression and glycogen levels were assessed after treatment. S15511 treatment was associated with an increase in insulin-stimulated glucose transport in type IIb fibers, while type I fibers were unaffected. The enhanced glucose transport was mirrored by a fiber type-specific increase in GLUT4 expression, while no improvement in insulin-signaling activity was observed. S15511 is a novel insulin sensitizer that is capable of improving glucose homeostasis in nondiabetic rats. The compound enhances skeletal muscle insulin sensitivity and specifically targets type IIb muscle fibers by increasing GLUT4 expression. Together these data show S15511 to be a potentially promising new drug in the treatment and prevention of type 2 diabetes.     

Glycogen metabolism in white and red muscle or normal and diabetic rats. The glycogen concentration and glycogen synthesis from glucose.

The amount of glycogen and its synthesis from glucose was studied in white muscle (extensor digitorum longus -- EDL) and red muscle (soleus -- SOL) of normal rats and rats with alloxan diabetes by the anthrone method. The amount of glycogen was higher in the white muscle of normal rats, both after a 24 hours' fast (0.37+/-0.02 mg/g as against 0.29+/-0.01 mg/g in the SOL) and with feeding ad libitium (0.72+/-0.05 mg/g as against 0.58+/-0.03 mg/g in the SOL). After a 24 hours' fast, the glycogen content of both muscles was non-significantly higher in alloxan-diabetic rats than in normal animals, whereas in diabetic animals fed ad libitum it was significantly lower than in normal rats fed in the same manner (0.54+/-0.07 mg/g in the EDL and 0.33+/-0.03 mg/g in the SOL). The difference between the glycogen content of the white and red muscle of diabetic rats was also in favour of the white muscle. Muscle glycogenesis from intragastrically administered glucose was higher in the red muscle in all the experimental groups. In normal fed ad libitum the glycogen content of the EDL did not change after glucose administration, but in the SOL it rose from 0.58+/-0.03 to 0.83+/-0.05 mg/g. In fasting (24 hours) normal rats it rose sharply in both muscles, from 0.037+/-0.02 to 0.57+/-0.03 mg/g in the EDL and from 0.29+/-0.01 to 0.87+/-0.06 mg/g in the SOL. In fasting (24 hours) diabetic animals, the glycogen content rose after glucose in the SOL only, from 0.36+/-0.01 to 0.66+/-0.06 mg/g. The differences found in glycogen synthesis in the white and red muscle of normal and diabetic rats are discussed mainly from the aspect of the existence of a relationship between the glycogen concentration and glycogen synthetase activity.     

Effects of hyperthyroidism on the sensitivity of glycolysis and glycogen synthesis to insulin in the soleus muscle of the rat.

1. The effects of hyperthyroidism on the sensitivity and responsiveness of glycolysis and glycogen synthesis to insulin were investigated in the isolated incubated soleus muscle of the rat. 2. Hyperthyroidism, which was induced by administration of tri-iodothyronine (T3) to rats for 2, 5 or 10 days, increased fasting plasma concentrations of glucose, insulin and free fatty acids. 3. Administration of T3 for 2 or 5 days increased the rates of glycolysis at all insulin concentrations studied: this was due to increased rates of both glucose phosphorylation and glycogen breakdown, but there was no effect of T3 on the sensitivity of glycolysis to insulin. However, administration of T3 for 10 days increased the sensitivity of the rate of glycolysis to insulin. 4. The concentration of adenosine in the gastrocnemius muscles of the rats was not different from controls after 5 days, but it was markedly decreased after 10 days of T3 administration. If these changes are indicative of changes in the soleus muscle, the increased sensitivity of glycolysis to insulin found after 10 days' T3 administration could be due to the decrease in the concentration of adenosine. 5. Administration of T3 decreased the sensitivity of glycogen synthesis to insulin and the glycogen content of the soleus muscles. This may explain the decreased rates of non-oxidative glucose disposal found in spontaneous and experimental hyperthyroidism in man. 6. The rates of glucose oxidation did not change after 2 days, but they were increased after 5 and 10 days of T3 administration.     

In vitro toxicity of alloxan for guinea pig B cells: comparison with rat B cells

Previous studies have shown that guinea pigs are resistant to the in vivo diabetogenic action of alloxan and that this resistance may be accompanied by a regeneration of B cells in the initial days following administration of the drug. In the studies reported here, we used the measurement of insulin and glucagon released over a 7-day culture period as indices of islet cell viability and examined effects of in vitro exposure to alloxan upon subsequent release of insulin and glucagon from guinea pig (alloxan-resistant) and rat (alloxan-sensitive) islet cell cultures. An alloxan dose-dependent decrease in subsequent insulin release was found. However, whereas the lowest concentration of the drug (1 mM) produced a significant depression in insulin release in rat islet cultures, with maximal depression occurring after exposure to 5 mM alloxan, insulin release from guinea pig cultures was not significantly depressed by 1 or 2 mM alloxan, and 5 mM alloxan treatment produced a submaximal depression. Furthermore, insulin release from guinea pig but not rat cultures increased transiently at between 6 and 18 hr during the first day following exposure to all doses of alloxan. Treatment with high doses of the drug (40 mM or greater) caused the same maximal chronic depression of insulin release for both species. In contrast, glucagon release from cultures of both species was not affected significantly following alloxan treatment. Thus, guinea pig B cells are more resistant than those of the rat to the action of alloxan, but this resistance can be overcome by employing high doses of the drug. Other factions unidentified by the present studies may also be involved in the failure of guinea pigs to develop diabetes following in vivo treatment with alloxan.     

Studies on gluconeogenesis and ketogenesis in isolated hepatocytes from alloxan diabetic rats.

Gluconeogenesis and ketogenesis were studied in isolated hepatocytes obtained from normal and alloxan diabetic rats. Insulin treatment maintained near-normal blood glucose levels and caused an increase in glycogen deposition. The third day after insulin withdrawal the rats displayed a diabetic syndrome marked by progressive hyperglycemia and glycogen depletion. Net glucose production in liver cells isolated from alloxan diabetic rats progressively increased with time up to 72 hr after the last in vivo insulin injection. Maximal glucose production was observed at 72 hr with 10 mM alanine, lactate, pyruvate, or fructose. Glucose production decreased at 96 hr. The same pattern was observed with the incorporation of labeled bicarbonate into glucose. Ketogenesis in liver cells and hepatic lipid content also peaked at 72 hr.     

Glycogen overload by postexercise insulin administration abolished the exercise-induced increase in GLUT4 protein

Summary To elucidate the role of muscle glycogen storage on regulation of GLUT4 protein expression and whole-body glucose tolerance, muscle glycogen level was manipulated by exercise and insulin administration. Sixty Sprague-Dawley rats were evenly separated into three groups: control (CON), immediately after exercise (EX0), and 16 h after exercise (EX16). Rats from each group were further divided into two groups: saline- and insulin-injected. The 2-day exercise protocol consisted of 2 bouts of 3-h swimming with 45-min rest for each day, which effectively depleted glycogen in both red gastrocnemius (RG) and plantaris muscles. EX0 rats were sacrificed immediately after the last bout of exercise on second day. CON and EX16 rats were intubated with 1 g/kg glucose solution following exercise and recovery for 16 h before muscle tissue collection. Insulin (0.5 μU/kg) or saline was injected daily at the time when glucose was intubated. Insulin injection elevated muscle glycogen levels substantially in both muscles above saline-injected group at CON and EX16. With previous day insulin injection, EX0 preserved greater amount of postexercise glycogen above their saline-injected control. In the saline-injected rats, EX16 significantly increased GLUT4 protein level above CON, concurrent with muscle glycogen supercompensation. Insulin injection for EX16 rats significantly enhanced muscle glycogen level above their saline-injected control, but the increases in muscle GLUT4 protein and whole-body glucose tolerance were attenuated. In conclusion, the new finding of the study was that glycogen overload by postexercise insulin administration significantly abolished the exercise-induced increases in GLUT4 protein and glucose tolerance.     

The influence of insulin and of contraction on glucose metabolism in the perfused diaphragm muscle from normal and streptozotocin-treated rats

1. The metabolism of [U-(14)C]glucose in perfused resting and contracting diaphragm muscle from normal rats and rats made diabetic with streptozotocin was studied in the presence and absence of insulin. 2. The incorporation of [U-(14)C]-glucose into glycogen and oligosaccharides was stimulated by insulin under all experimental conditions studied. 3. In the normal perfused resting diaphragm muscle the incorporation of radioactivity from [(14)C]glucose into lactate and CO(2) was not affected by insulin. 4. Periodic contractions, induced by electrical stimulation of the perfused diaphragm muscle in the absence of insulin, caused an increased incorporation of (14)C into glycogen and hexose phosphate esters, whereas incorporation of (14)C into lactate was greatly decreased. Production of (14)CO(2) in the contracting muscle was not significantly different from that in resting muscle. Addition of insulin to the perfusion liquid caused a further increase in formation of [(14)C]-glycogen in contracting muscle to values reached in the resting muscle in the presence of insulin. Formation of [(14)C]lactate was also stimulated by insulin, to values close to those found in the resting muscle in the presence of insulin. 5. In the diabetic resting muscle the rate of glucose metabolism was very low in the absence of insulin. Insulin increased formation of [(14)C]glycogen to the value found in normal muscle in the absence of insulin. Production of (14)CO(2) and formation of [(14)C]hexose phosphate remained unchanged. 6. In the diabetic contracting muscle production of (14)CO(2) was increased to values approaching those found in normal contracting muscle. Formation of [(14)C]lactate and [(14)C]glycogen was also increased by contraction, to normal values. Only traces of [(14)C]hexose phosphate were detectable. Addition of insulin to the perfusion medium stimulated formation of [(14)C]glycogen, to values found in normal contracting muscle. Production of [(14)C]hexose phosphate was stimulated by insulin, to approximately the values found in the normal contracting muscle. Production of (14)CO(2) and [(14)C]lactate, however, was not significantly affected by insulin. 7. These results indicate that the defects of glucose metabolism observed in perfused resting diabetic diaphragm muscle can be partially corrected by contraction, and in the presence of insulin the contracting diabetic muscle has a completely normal pattern of glycogen synthesis and lactate production, but CO(2) production remains impaired.     

Effects of aging on the responsiveness and sensitivity of glucose metabolism to insulin in the incubated soleus muscle isolated from Sprague-Dawley and Wistar rats.

1. The effects of aging on the sensitivity and responsiveness of glucose transport, lactate formation and glycogen synthesis to insulin were studied in the incubated stripped soleus muscle isolated from aging Sprague-Dawley and Wistar rats. 2. As Sprague-Dawley rats aged from 5 to 13 weeks, there were marked increases in the concentrations of insulin that were required for half-maximal stimulation (i.e. EC50 value, which is a measure of sensitivity) of glucose transport, lactate formation and glycogen synthesis. 3. In marked contrast, there were no alterations in sensitivities of any of these processes to insulin in soleus muscle prepared from Wistar rats aged between 6 and 12 weeks. 4. However, in soleus muscles from 85-week-old Wistar rats the rates of glycogen synthesis in response to basal, sub-maximal and maximal concentrations of insulin were markedly decreased. The insulin EC50 value of glycogen synthesis was increased 4-fold, but was unchanged for lactate formation. 5. The insulin-stimulated rates of glucose transport in soleus muscles from 5- or 85-week-old Wistar rats were not significantly different.