Fasting does not impair insulin-stimulated glucose uptake but alters intracellular glucose metabolism in conscious rats

Effects of 24-h and 48-h fasting on maximal insulin-stimulated whole-body and muscle glucose uptake, glycogen synthesis, and glycolysis were studied in conscious rats by combining the glucose clamp technique with tracer methods. Fasting decreased body weight and basal plasma glucose, plasma insulin, hepatic glucose output, and glucose clearance (P < 0.05 for all). However, maximal insulin-stimulated whole-body glucose uptake, normalized to body weight, was almost identical in fed, 24-h fasted, and 48-h fasted rats (191 +/- 8, 185 +/- 14, and 182 +/- 5 mumol.kg-1.min-1, respectively; P > 0.7). Similarly, rates of insulin-stimulated glucose uptake by four different skeletal muscles, estimated by the 2-deoxyglucose injection technique, were not different among the three groups. In contrast to glucose uptake, insulin-stimulated whole-body glycolysis was decreased significantly after fasting (36% after 48 h fasting; P < 0.05), whereas insulin-stimulated whole-body glycogen synthesis was increased (44% after 48 h fasting; P < 0.05). In fed rats, glycolysis was the major pathway for glucose metabolism during hyperinsulinemia, accounting for 60 +/- 5% of glucose uptake. This fraction was decreased significantly by fasting (P < 0.01), so that after a 48-h fast, glycolysis accounted for only 40 +/- 3% of insulin-stimulated glucose uptake and glycogen synthesis became predominant pathway, accounting for 60 +/- 3% of whole-body glucose utilization. Whole-body patterns of glucose metabolism during hyperinsulinemia were paralleled by glucose metabolism in individual muscles.(ABSTRACT TRUNCATED AT 250 WORDS)     

Circulating fatty acids are essential for efficient glucose-stimulated insulin secretion after prolonged fasting in humans

In the fasted rat, efficient glucose-stimulated insulin secretion (GSIS) is absolutely dependent on an elevated level of circulating free fatty acids (FFAs). To determine if this is also true in humans, nonobese volunteers were fasted for 24 h (n = 5) or 48 h (n = 5), after which they received an infusion of either saline or nicotinic acid (NA) to deplete their plasma FFA pool, followed by an intravenous bolus of glucose. NA treatment resulted in a fall in basal insulin concentrations of 35 and 45% and in the area under the insulin response curve (area under the curve [AUC]) to glucose of 47 and 42% in the 24- and 48-h fasted individuals, respectively. The 48-h fasted subjects underwent the same procedure with the addition of a coinfusion of Intralipid plus heparin (together with NA) to maintain a high concentration of plasma FFAs throughout the study. The basal level and AUC for insulin were now completely normalized (C-peptide profiles paralleled those for insulin). To assess the effect of an overnight fast, nonobese (n = 6) and obese (n = 6) subjects received an infusion of either saline or NA, followed by a hyperglycemic clamp (200 mg/dl). The insulin AUC in response to glucose was unaffected by lowering of the FFA level in nonobese subjects, but fell by 29% in the obese group. The data clearly demonstrate that in humans, the rise in circulating FFA levels after 24 and 48 h of food deprivation is critically important for pancreatic beta-cell function both basally and during subsequent glucose loading. They also suggest that the enhancement of GSIS by FFAs in obese individuals is more prominent than that seen in their nonobese counterparts.     

Effects of hepatic portal infusion of deionized water on metabolic and hormonal responses to exercise in rats

The present study was conducted to investigate the in vivo effects of an intrahepatic infusion of deionized water during exercise in rats. Adrenodemedullated male Sprague-Dawley rats were continuously infused for 30 min either at rest or during treadmill exercise (26 m/min, 0% grade). Rats were randomly assigned to one of three infusion conditions (52 micro ul/min) with either deionized water (PW) or saline (PS; NaCl; 0.9%) via the hepatic portal vein or deionized water through the jugular vein (JW). The exercise period caused a significant (P < 0.05) decrease in liver glycogen and relative liver water content and peripheral and portal blood glucose and insulin while increasing peripheral and portal glucagon and K+ plasma concentrations. These responses, with the exception of K+, were not influenced by the different types of infusions. The increase in K+ during exercise was significantly (P < 0.05) higher in JW rats than in the PW and PS groups. Both the infusion and exercise protocols did not significantly alter the liver weight-to-body weight ratio, plasma osmolality, free fatty acids, beta-hydroxybutyrate, Na+, Cl-, vasopressin, and catecholamine concentrations. It is concluded that an hepatic portal infusion of deionized water does not specifically alter the metabolic and hormonal responses to exercise in rats.     

Characterization of adrenocortical cell lines produced by genetically targeted tumorigenesis in transgenic mice

Microvascular hyperaemia is decreased in subjects at risk of developing non-insulin-dependent diabetes mellitus (NIDDM) who have fasting hyperglycaemia. Such microvascular abnormalities may be involved in the pathogenesis of diabetic microangiopathy. To investigate the relationship of reduced microvascular hyperaemia to metabolic and blood pressure abnormalities associated with the prediabetic state, we studied 24 subjects with fasting hyperglycaemia and 24 age- and sex-matched control subjects. The microvascular hyperaemic response to local heating of the skin on the dorsum of the foot measured by laser Doppler fluximetry was reduced in the subjects with fasting hyperglycaemia (1.18 [0.87-1.83] volts vs 1.51 [1.30-2.14] volts normal subjects; p = 0.0002) and was negatively correlated with fasting plasma insulin concentration (Rs = 0.70; p = 0.001) and positively related to insulin sensitivity determined by continuous infusion of glucose with model assessment (CIGMA) (Rs = 0.52; p = 0.01), but showed no association with fasting plasma glucose, beta-cell function 24 h ambulatory blood pressure profiles or serum lipid concentrations. These results suggests that hyperinsulinaemia, as a result of insulin resistance, may have a detrimental effect on microvascular function in the prediabetic state.     

Glucose turnover in the post-absorptive rat and the effects of halothane anaesthesia

1. Rates and rate coefficients of glucose utilization and replacement in post-absorptive rats, either conscious or under halothane anaesthesia, were determined in a thermoneutral environment by using [5-3H]- and [U-14C]glucose. Label was not injected into rats under halothane until about 0.5h after anaesthesia was initiated. 2. Comparison with the results for 24h-starved rats in the preceding paper [Heath et al. (1977) Biochem. J. 162, 643-651] showed that insulin concentrations were considerably higher but rate coefficients for glucose utilization were little altered in post-absorptive rats. Sensitivity to insulin was thus considerably increased by a 24h period of starvation in the rat. 3. Fractional recycling of glucose carbon in post-absorptive rats was under one-half of that in starved rats, reflecting the larger contribution of liver glycogenolysis to glucose production in the former. 4. In post-absorptive rats halothane decreased the mean rate of glucose utilization by about 17%. This decrease was associated with an increase in mean plasma insulin concentration, showing that halothane decreased sensitivity to insulin. 5. Recycling was slightly increased by halothane, indicating that the contribution of liver glycogen to the total glucogenic rate was decreased, probably because liver glycogen concentration were about 40% lower throughout the rate determinations in halothane. 6. Comparison of our results with earlier work shows that during and shortly after induction of halothane anaesthesia glucose turnover must have been greatly increased whereas from about 0.5h after induction it was decreased.     

Effects of dichloroacetate on glycogen metabolism in B6C3F1 mice

Dichloroacetate (DCA) is a by-product of drinking water chlorination. Administration of DCA in drinking water results in accumulation of glycogen in the liver of B6C3F1 mice. To investigate the processes affecting liver glycogen accumulation, male B6C3F1 mice were administered DCA in drinking water at levels varying from 0.1 to 3 g/l for up to 8 weeks. Liver glycogen synthase (GS) and glycogen phosphorylase (GP) activities, liver glycogen content, serum glucose and insulin levels were analyzed. To determine whether effects were primary or attributable to increased glycogen synthesis, some mice were fasted and administered a glucose challenge (20 min before sacrifice). DCA treatments in drinking water caused glycogen accumulation in a dose-dependent manner. The DCA treatment in drinking water suppressed the activity ratio of GS measured in mice sacrificed at 9:00 AM, but not at 3:00 AM. However, net glycogen synthesis after glucose challenge was increased with DCA treatments for 1-2 weeks duration, but the effect was no longer observed at 8 weeks. Degradation of glycogen by fasting decreased progressively as the treatment period was increased, and no longer occurred at 8 weeks. A shift of the liver glycogen-iodine spectrum from DCA-treated mice was observed relative to that of control mice, suggesting a change in the physical form of glycogen. These data suggest that DCA-induced glycogen accumulation at high doses is related to decreases in the degradation rate. When DCA was administered by single intraperitoneal (i.p.) injection to na?ve mice at doses of 2-200 mg/kg at the time of glucose challenge, a biphasic response was observed. Doses of 10-25 mg/kg increased both plasma glucose and insulin concentrations. In contrast, very high i.p. doses of DCA (> 75 mg/kg) produced progressive decreases in serum glucose and glycogen deposition in the liver. Since the blood levels of DCA produced by these higher i.p. doses were significantly higher than observed with drinking water treatment, we conclude that apparent differences with data of previous investigations is related to substantial differences in systemic dose and/or dose-time relations.     

Essentiality of circulating fatty acids for glucose-stimulated insulin secretion in the fasted rat

We asked whether the well known starvation-induced impairment of glucose-stimulated insulin secretion (GSIS) seen in isolated rat pancreas preparations also applies in vivo. Accordingly, fed and 18-24-h-fasted rats were subjected to an intravenous glucose challenge followed by a hyperglycemic clamp protocol, during which the plasma-insulin concentration was measured. Surprisingly, the acute (5 min) insulin response was equally robust in the two groups. However, after infusion of the antilipolytic agent, nicotinic acid, to ensure low levels of plasma FFA before the glucose load, GSIS was essentially ablated in fasted rats, but unaffected in fed animals. Maintenance of a high plasma FFA concentration by coadministration of Intralipid plus heparin to nicotinic acid-treated rats (fed or fasted), or further elevation of the endogenous FFA level in nonnicotinic acid-treated fasted animals by infusion of etomoxir (to block hepatic fatty acid oxidation), resulted in supranormal GSIS. The in vivo findings were reproduced in studies with the perfused pancreas from fed and fasted rats in which GSIS was examined in the absence and presence of palmitate. The results establish that in the rat, the high circulating concentration of FFA that accompanies food deprivation is a sine qua non for efficient GSIS when a fast is terminated. They also serve to underscore the powerful interaction between glucose and fatty acids in normal beta cell function and raise the possibility that imbalances between the two fuels in vivo could have pathological consequences.     

Further evidence for a central role of adipose tissue in the antihyperglycemic effect of metformin

OBJECTIVE: To evaluate further the relative roles played by liver and adipose tissue in the therapeutic response to metformin in patients with type 2 diabetes. RESEARCH DESIGN AND METHODS: A total of 11 patients with diet-treated type 2 diabetes were given metformin for approximately 3 months. Measurements were made before and after treatment of 1) fasting and postprandial plasma glucose, insulin, and free fatty acid (FFA) concentrations; 2) glucose appearance (Ra) and disappearance (Rd) rates measured overnight with 3-[3H]glucose; and 3) plasma FFA concentrations during a 195-min infusion period at relatively low insulin (approximately 12-24 microU/ml) concentrations. RESULTS: Mean +/- SEM fasting plasma glucose concentration was significantly lower (175 +/- 11 vs. 224 +/- 15 mg/dl; P < 0.01) after treatment with metformin. Mean +/- SEM insulin concentrations measured from 8:00 A.M. to 5:00 P.M. did not change with treatment. However, both glucose and FFA concentrations were significantly lower (P < 0.01) when measured over the same time period, and the decreases in plasma FFA and glucose concentration were highly correlated (r = 0.81; P = 0.03). Overnight glucose turnover studies indicated that neither Ra (hepatic glucose production [HGP]) nor Rd changed significantly with treatment in association with metformin treatment. Since plasma glucose concentration was much lower after metformin treatment, the overnight glucose metabolic clearance rate (MCR) was significantly lower (P < 0.01). Finally, the ability of insulin to inhibit isoproterenol-stimulated increases in plasma FFA concentration was enhanced in metformin-treated patients (P < 0.05). CONCLUSIONS: Metformin treatment was associated with significantly lower fasting plasma glucose concentrations and lower day-long plasma glucose and FFA concentrations. Although overnight HGP was unchanged after treatment with metformin, the overnight glucose MCR was significantly increased, and the antilipolytic activity of insulin was also enhanced. Given these findings, it is suggested that at least part of the antihyperglycemic effect of metformin is due to a decrease in release of FFA from adipose tissue, leading to lower circulating FFA concentrations and an increase in glucose uptake.     

Plasma amylin concentrations in fasted and fed rats quantified by a monoclonal immunoenzymometric assay

Amylin is a 37-amino acid peptide co-secreted from the pancreatic beta-cell with insulin in response to nutrient stimuli. Plasma amylin concentrations in the rat are reported to vary widely. We have employed a recently-developed immunoenzymometric assay to quantify plasma amylin concentrations in fasted, fed and glucose-administered rats. Fasted amylin concentrations ranged between 1.02+/-0.09 and 1.63+/-0.15pM among three different common rat strains, and increased up to 7.70+/-0.80 pM after feeding. The differences among strains and between fasted and fed rats were all significant at P<0.01 or less. Intravenous glucose administration (5.2 mmol/kg) also significantly increased plasma amylin concentrations in fasted rats from 1.5+/-0.3pM to 3.4+/-0.5pM, and in fed rats from 4.6+/-1.1 pM to 9.1+/-1.7 pM. Plasma amylin/insulin molar ratios ranged between 2.3+/-0.2% and 3.6+/-0.5% (mean 3.0%), but did not differ among strains, or between the fasted vs fed state in any strain. In conclusion, a new sensitive immunoenzymometric assay revealed fasting plasma concentrations which are lower than previously reported, and which are significantly increased by stimulation with feeding or glucose administration.     

Effects of a quick-release form of bromocriptine (Ergoset) on fasting and postprandial plasma glucose, insulin, lipid, and lipoprotein concentrations in obese nondiabetic hyperinsulinemic women

OBJECTIVE: To assess the effect on various aspects of carbohydrate and lipid metabolism of administering a quick-release formulation of bromocriptine (Ergoset) to obese, nondiabetic, hyperinsulinemic women. RESEARCH DESIGN AND METHODS: Hourly concentrations of prolactin, glucose, insulin, free fatty acid (FFA), and triglyceride were measured for 24 h before and after approximately 8 weeks of treatment with Ergoset. In addition, fasting lipid and lipoprotein concentrations and the steady-state plasma glucose (SSPG) concentration in response to a continuous infusion of somatostatin, insulin, and glucose were determined before and after Ergoset administration. RESULTS: Circulating prolactin concentrations were dramatically decreased (P < 0.001) following treatment, associated with a significant fall (P < 0.05) in 24-h-long plasma glucose, FFA, and triglyceride concentrations. Neither circulating plasma insulin concentrations nor the ability of insulin to mediate glucose disposal changed with treatment. Finally, fasting total cholesterol fell (P < 0.05) and the ratio of total to HDL cholesterol decreased (P = 0.06) in association with Ergoset treatment. CONCLUSIONS: The fact that significant metabolic improvement was seen in the obese nondiabetic hyperinsulinemic women studied suggests that Ergoset could be of therapeutic benefit in clinical conditions of hyperglycemia and/or dyslipidemia.     

Ketosis resistance in the male offspring of protein-malnourished rat dams

Plasma beta-hydroxybutyrate concentrations were measured in the offspring of rats that were fed either a control (20% protein) diet or low-protein (8% protein) diet during pregnancy and lactation. Low-protein offspring had significantly lower plasma beta-hydroxybutyrate compared with controls in the fed state (P < .04) and after fasting for 24 hours (P < .001) and 48 hours (P < .04). There were no differences in blood glucose, acetoacetate, plasma glucagon, cholesterol, or glycerol between control and low-protein offspring. However, plasma nonesterified fatty acids (NEFAs) were significantly higher in low-protein offspring in the fed state (P < .05). In contrast, plasma triglycerides and insulin were significantly lower in low-protein offspring compared with controls when fed (P < .001) and after a 24-hour fast (P < .001). These results suggest that poor maternal and early postnatal nutrition can have long-term effects on ketone body metabolism in the offspring during adulthood. This apparent ketosis resistance is similar to that observed in some forms of human diabetes.     

Effects of total fasting or chronic food restriction on plasma endothelin levels in rats

We tested the effects of 24- and 48-h fasting and 40% calorie restriction stresses on plasma endothelin (ET)-1,2 levels in male Sprague-Dawley rats. Plasma ET-1,2 levels in pg/ml were lower in 24-h fasted rats (15.48 +/- 3.49), 48-h fasted rats (5.28 +/- 4.32), and in chronically food-deprived rats (R) (10.49 +/- 6.28) compared to ad lib-fed (AL) rats (21.23 +/- 9.38). The R rats were pair-fed 40% fewer calories than AL rats. We conclude that calorie restriction or total food deprivation stress decreases plasma ET-1,2 levels, unlike many other forms of physiological stress that have been shown to increase plasma ET-1,2 levels.     

Nutritional status modulates rat liver cytochrome P450 arachidonic acid metabolism

Alterations in nutritional status affect hepatic cytochrome P450 levels. Since cytochromes P450 participate in the metabolism of arachidonic acid, we hypothesized that changes in liver P450 arachidonic acid metabolism occur during fasting and refeeding. Male Fisher 344 rats were either fed, fasted 48 hr (F48), fasted 48 hr and then refed 6 hr (F48/R6), or fasted 48 hr and then refed 24 hr (F48/R24). F48 rats had reduced body weight, increased plasma beta-hydroxybutyrate, and reduced plasma insulin compared with the other groups. Although there was no significant change in total liver P450 content, there was a significant 20%, 48%, and 24% reduction in total hepatic microsomal arachidonic acid metabolism in F48, F48/R6, and F48/R24 rats, respectively, compared with fed rats. Epoxygenase activity decreased by 28%, 51%, and 26% in F48, F48/R6, and F48/R24 rats, respectively. In contrast, omega-1 hydroxylase activity increased by 126% in F48 rats compared with fed rats. Immunoblotting revealed that levels of CYP2C11 protein were markedly reduced, whereas levels of CYP2E1 protein were markedly increased in the F48 and F48/R6 groups. In contrast, levels of CYP1A1, CYP1A2, CYP2B1, CYP2J3, CYP4A1, and CYP4A3 were unchanged with fasting/refeeding. Northern blots revealed that levels of CYP2C11 mRNAs were decreased, whereas CYP2E1 mRNAs were increased in F48 and F48/R6 rats. Recombinant CYP2C11 metabolized arachidonic acid primarily to epoxides with preference for the 14(S),15(R)-, 11(R), 12(S)-, and 8(S),9(R)- epoxyeicosatrienoic acid enantiomers. We conclude that (1) nutritional status affects hepatic microsomal arachidonic acid metabolism, (2) reduced epoxygenase activity in F48 and F48/R6 rats is accompanied by decreased levels of CYP2C11, (3) increased omega-1 hydroxylase activity is accompanied by augmented levels of CYP2E1, and (4) the effects of fasting on CYP2C11 and CYP2E1 expression occur at the pretranslational level.     

Increased intestinal glucose absorption and postprandial hyperglycaemia at the early step of glucose intolerance in Otsuka Long-Evans Tokushima Fatty rats

Otsuka Long-Evans Tokushima Fatty (OLETF) rats are reported to be obese Type II (non-insulin-dependent) diabetic rats with insulin resistance and impaired insulin secretion. To investigate the contribution of intestinal glucose absorption to postprandial hyperglycaemia, we determined the plasma xylose concentrations after an 0.8 g/kg oral xylose load which was used as a test of small intestinal glucose absorption in 6-week-old OLETF rats and weight-matched Long-Evans Tokushima Otsuka (LETO) rats. An oral glucose tolerance test showed that OLETF rats developed hyperglycaemia at 60 and 90 min after the glucose load, though the fasting plasma glucose concentration, insulin concentration and insulin-induced in vivo glucose utilization rate were similar. Consistently, in an oral D-xylose loading test, the peak concentration of plasma xylose in OLETF rats was increased by 58.7% compared with that of LETO rats (p < 0.005). The disappearance rate of plasma xylose concentrations after intravenous xylose loading did not differ between the two strains. Co-treatment with 0.4 g/kg phlorizin, a specific inhibitor of sodium-dependent glucose transporter 1 (SGLT1), abolished both plasma glucose and xylose concentrations after the loads. Morphological studies showed that both the small intestinal wet weight and surface area were 30% larger in the OLETF rats than in the LETO rats. Furthermore, the SGLT1 mRNA content of OLETF rats also increased compared with LETO rats. These results suggest that an increased SGLT1 expression concomitant with intestinal hypertrophy in OLETF rats is partly associated with postprandial hyperglycaemia before the onset of insulin resistance and hyperinsulinaemia.     

Neonatal de-afferentation of capsaicin-sensitive sensory nerves increases in vivo insulin sensitivity in conscious adult rats

Sensory neuropeptides, released from the peripheral nervous system, might modulate glucose homeostasis by antagonizing insulin action. The effects of de-afferentation of functional small diameter unmyelinated C-fibres (sensory nerves) on in vivo insulin-mediated intracellular glucose metabolism were investigated by using euglycaemic insulin (6 and 18 mU/kg x min) clamps with [3-(3)H]-glucose infusion in 24 adult rats, treated neonatally with either capsaicin (CAP) (50 mg/kg) or vehicle (CON). Following the clamp, skeletal muscle groups, liver and adipose tissue were freeze-clamped. At plasma insulin levels of approximately 90 mU/l, CAP-rats showed a 21% increase in whole body glucose uptake compared with CON (24.4 +/- 1.6 vs 20.1 +/- 0.8 mg/kg min, p < 0.02), which was paralleled by a 20% increase in whole body glycolysis (12.6 +/- 0.8 vs 10.5 +/- 0.5 mg/ kg.min p < 0.05) (concentration of 3H2O in plasma). Whole body skeletal muscle glycogenesis was increased by 80% in CAP-rats (5.7 +/- 0.7 vs 3.1 +/- 0.7 mg/kg x min, p < 0.05) with increased muscle glycogen synthase activity. Whole body (muscle, liver and adipose tissue combined) de novo lipogenesis also was increased in CAP-rats compared with CON (0.69 +/- 0.10 vs 0.44 +/- 0.06 mg/kg x min, p < 0.05) (incorporation of [3-(3)H]-glucose counts into glycogen or fat). Hepatic glucose production was lower in CAP-rats compared with CON (0.6 +/- 0.6 vs 2.1 +/- 0.7 mg/kg x min, p < 0.05). Plasma glucagon, corticosterone, epinephrine and norepinephrine levels were reduced in CAP-rats: 43 +/- 2 compared with 70 +/- 6 pg/ml, 855 +/- 55 compared with 1131 +/- 138 nmol/l, 513 +/- 136 compared with 1048 +/- 164 pmol/l and 928 +/- 142 compared with 1472 +/- 331 pmol/l, respectively, p < 0.05. At plasma insulin levels of approximately 400 mU/l, CAP-rats showed no differences in peripheral and hepatic insulin action compared with CON. We conclude that the removal of endogenous sensory neuropeptides, by de-afferentation of capsaicin-sensitive sensory nerves, increases in vivo insulin sensitivity, but not responsiveness: 1) primarily through an increased sensitivity of skeletal muscle glycogen synthesis to insulin; 2) through a reduction in the levels of counter-regulatory hormones, thereby creating a milieu which favours overall in vivo insulin sensitivity with respect to glucose uptake, glucose production, glycolysis, glycogenesis and lipogenesis.     

Non-insulin and non-exercise related increase of glucose utilization in rats and mice

The effects of high-energy phosphate contents in muscles on glucose tolerance and glucose uptake into tissues were studied in rats and mice. Enhanced glucose tolerance associated with depleted high-energy phosphates and elevated glycogen content in muscles and liver was observed in animals fed creatine analogue beta-guanidinopropionic acid (beta-GPA). Distribution of infused 2-[1-14C]deoxy-D-glucose in tissues especially in the soleus muscle, kidney, and brain was greater in mice fed beta-GPA than controls. The glucose uptake was decreased when the contents of ATP and glycogen were normalized following creatine supplementation. Plasma insulin in animals at rest was lower and its concentration after intraperitoneal glucose infusion tended to be less in animals fed beta-GPA than controls (p > 0.05), although the pattern of insulin response to glucose loading was similar to the control. The daily voluntary activity in beta-GPA fed mice was also less than controls. These results suggest that improved glucose tolerance is not related to elevated insulin concentration and/or decreased glycogen following exercise. Such improvement may be due to an increased mitochondrial energy metabolism caused by depletion of high-energy phosphates.     

Unchanged gene expression of glycogen synthase in muscle from patients with NIDDM following sulphonylurea-induced improvement of glycaemic control

We have previously shown that the mRNA expression of muscle glycogen synthase is decreased in non-insulin-dependent diabetic (NIDDM) patients; the objective of the present protocol was to examine whether the gene expression of muscle glycogen synthase in NIDDM is affected by chronic sulphonylurea treatment. Ten obese patients with NIDDM were studied before and after 8 weeks of treatment with a weight-maintaining diet in combination with the sulphonylurea gliclazide. Gliclazide treatment was associated with significant reductions in HbA1C (p=0.001) and fasting plasma glucose (p=0.005) as well as enhanced beta-cell responses to an oral glucose load. During euglycaemic, hyperinsulinaemic clamp (2 mU x kg-1 x min-1) in combination with indirect calorimetry, a 35% (p=0.005) increase in whole-body insulin-stimulated glucose disposal rate, predominantly due to an increased non-oxidative glucose metabolism (p=0.02) was demonstrated in teh gliclazide-treated patients when compared to pre-treatment values. In biopsies obtained from vastus lateralis muscle during insulin infusion, the half-maximal activation of glycogen synthase was achieved at a significantly lower concentration of the allosteric activator glucose 6-phosphate (p=0.01). However, despite significant increases in both insulin-stimulated non-oxidative glucose metabolism and muscle glycogen synthase activation in gliclazide-treated patients no changes were found in levels of glycogen synthase mRNA or immunoreactive protein in muscle. In conclusion, improved blood glucose control in gliclazide-treated obese NIDDM patients has no impact on the gene expression of muscle glycogen synthase.     

Effect of 14 days'' subcutaneous administration of the human amylin analogue, pramlintide (AC137), on an intravenous insulin challenge and response to a standard liquid meal in patients with IDDM

Individuals with insulin-dependent diabetes mellitus (IDDM or type 1 diabetes) are deficient in both insulin and amylin, peptides secreted by the beta cell. We have investigated the effects of amylin replacement therapy employing the human amylin analogue, pramlintide (25, 28, 29-pro-human amylin, previously referred to as AC137), upon the responses to a standardized insulin infusion (40 mU. kg-1. h-1) for 100 min and a liquid Sustacal meal (360 kcal) in 84 healthy IDDM patients. Following baseline evaluations, patients were randomly assigned to receive subcutaneous injections of placebo, 30, 100 or 300 micrograms pramlintide 30 min before meals for 14 days. There was no meaningful difference between adverse events reported by the 30-micrograms pramlintide and the placebo groups, but ten subjects withdrew due to nausea, eight of these in the 300-micrograms dose group. Peak plasma pramlintide concentrations for the 30-micrograms group were 21 +/- 3 and 29 +/- 5 pmol/l on Days 1 and 14, respectively. These values are similar to postprandial plasma amylin concentrations in normal volunteers. The plasma glucose, free insulin, glucagon, epinephrine and norepinephrine concentrations during the insulin infusion test before and after therapy were identical in each of the group. Prior to pramlintide therapy, Sustacal ingestion produced a 4.0-4.8 mmol/l rise in plasma glucose concentrations in each of the groups. Pramlintide therapy reduced postprandial hyperglycaemia as reflected by the 3-h incremental AUCglucose (AUCglucose above or below fasting glucose concentration) Day 1 vs Day 14: 30 micrograms, 322 +/- 92 vs -38 +/- 161 mmol/l.min, p = 0.010; 100 micrograms, 317 +/- 92 vs -39 +/- 76 mmol/l.min, p = 0.001; and 300 micrograms, 268 +/- 96 vs -245 +/- 189 mmol/l.min, p = 0.077. Thus, pramlintide therapy with these regimens did not appear to impair either in vivo insulin action or the counter-regulatory response to hypoglycaemia but did show a clear effect of blunting postprandial hyperglycaemia following a standardized meal.     

Characterization of apolipoprotein E7 (Glu(244)-->Lys, Glu(245)--->Lys), a mutant apolipoprotein E associated with hyperlipidemia and atherosclerosis

To examine the mechanism by which free fatty acids (FFA) induce insulin resistance in human skeletal muscle, glycogen, glucose-6-phosphate, and intracellular glucose concentrations were measured using carbon-13 and phosphorous-31 nuclear magnetic resonance spectroscopy in seven healthy subjects before and after a hyperinsulinemic-euglycemic clamp following a five-hour infusion of either lipid/heparin or glycerol/heparin. IRS-1-associated phosphatidylinositol 3-kinase (PI 3-kinase) activity was also measured in muscle biopsy samples obtained from seven additional subjects before and after an identical protocol. Rates of insulin stimulated whole-body glucose uptake. Glucose oxidation and muscle glycogen synthesis were 50%-60% lower following the lipid infusion compared with the glycerol infusion and were associated with a approximately 90% decrease in the increment in intramuscular glucose-6-phosphate concentration, implying diminished glucose transport or phosphorylation activity. To distinguish between these two possibilities, intracellular glucose concentration was measured and found to be significantly lower in the lipid infusion studies, implying that glucose transport is the rate-controlling step. Insulin stimulation, during the glycerol infusion, resulted in a fourfold increase in PI 3-kinase activity over basal that was abolished during the lipid infusion. Taken together, these data suggest that increased concentrations of plasma FFA induce insulin resistance in humans through inhibition of glucose transport activity; this may be a consequence of decreased IRS-1-associated PI 3-kinase activity.     

Evaluation of BTS 67 582, a novel antidiabetic agent, in normal and diabetic rats

1. The effect of BTS 67 582, a novel antidiabetic agent, has been evaluated on plasma glucose and plasma insulin in normal and streptozotocin-induced diabetic rats. 2. BTS 67 582 (3 to 300 mg kg-1, p.o.) caused a dose- and time-dependent reduction in plasma glucose and an increase in plasma insulin in both fasted and glucose-loaded normal rats. The ED50 for the glucose lowering effect of BTS 67 582 in fasted rats was 37.6, 18.4 and 18.5 mg kg-1 at 1, 2 and 4 h after administration respectively. 3. In streptozotocin-induced (50 mg kg-1, i.v.) diabetic rats, BTS 67 582 (37-147 mg kg-1, p.o.) caused significant reductions of plasma glucose following a glucose load, whereas glibenclamide (100 mg kg-1, p.o.) was ineffective. BTS 67 582 significantly increased plasma insulin compared to controls whereas glibenclamide did not. 4. BTS 67 582 did not displace [3H]-glibenclamide from its binding sites in rat brain, guinea-pig ventricle or the HIT-T15 insulinoma beta-cell line. BTS 67 582 does not therefore appear to modulate its action via an effect on the 'sulphonylurea' receptor. 5. In fasted rats, the glucose lowering effect of BTS 67 582 (100 mg kg-1 p.o.) and glibenclamide (1 mg kg-1, p.o.) were antagonized by diazoxide (30 mg kg-1, i.p.). In addition BTS 67 582, like glibenclamide, caused a dose-dependent rightward shift of cromakalim-induced relaxation of noradrenaline precontracted rat aortic strips, suggesting the involvement of KATP channels. 6. In summary, BTS 67 582 produces a blood glucose-lowering effect in normal and streptozotocin-induced diabetic rats associated with increased insulin concentrations. This effect appears to be due to a blockade of ATP-sensitive potassium channel activity via a different binding site to that of glibenclamide.