Altered anomeric specificity of glucose-induced insulin release in rabbits with duct-ligated pancreas

Ligation of the pancreatic duct in rabbits provokes a decrease in the insulin and glucagon content of the pancreas, and may lead to chronic hyperglycemia. The insulin secretory behavior of the perfused pancreas is perturbed in duct-ligated animals, and this is illustrated in several respects:

Beta-cell hypersensitivity for glucose precedes loss of glucose-induced insulin secretion in 90% pancreatectomized rats

Summary Glucose-induced insulin secretion is impaired in the presence of chronic hyperglycaemia. Insulin secretion was studied in a diabetic rat model prior to the beta cells becoming non-responsive to glucose in order to map out the sequence of changes that accompany chronic hyperglycaemia. In vitro pancreas perfusions were carried out 1 and 2 weeks after a 90% pancreatectomy; controls underwent a sham pancreatectomy. One week post 90% pancreatectomy: (i) non-fasting plasma glucose values were 2–3 mmol/l above normal, (ii) the in vitro insulin response to 16.7 mmol/l glucose was 20 ± 4% of shams, a response that was appropriate for the surgical reduction in beta-cell mass, (iii) the beta-cell sensitivity for glucose was increased as reflected by left-shifted dose-response curves for glucose-induced insulin secretion (half maximal insulin output 5.7 mmol/l glucose vs 16.5 mmol/l glucose in shams) and glucose potentiation of arginine-induced insulin secretion (half maximal insulin output 3.5 mmol/l glucose vs 14.8 mmol/l glucose in shams). This heightened beta-cell sensitivity for glucose was not a result of the hyperglycaemia, because similarly reduced half-maximal insulin responses were found after a 60% pancreatectomy, a surgical procedure in which plasma glucose values remained normal. In summary, a rise in beta-cell sensitivity for glucose precedes the loss of glucose-induced insulin secretion in diabetic rats.     

Altered anomeric specificity of glucose-induced insulin release in rabbits with duct-ligated pancreas

Ligation of the pancreatic duct in rabbits provokes a decrease in the insulin and glucagon content of the pancreas, and may lead to chronic hyperglycemia. The insulin secretory behavior of the perfused pancreas is perturbed in duct-ligated animals, and this is illustrated in several respects: 1. The steady-state insulin output evoked by L-leucine (10 mM) is higher in duct-ligated than control rabbits; 2. In the presence of the amino acid, the response to D-glucose is characterized by a delayed onset, the absence of an early secretory peak, and a sluggish return towards basal value upon removal of the hexose from the perfusate; and 3. Whereas control rabbits display a higher secretory response to alpha- than beta-D-glucose, such is no more the case in duct-ligated rabbits. The perturbation of the anomeric specificity in secretory response is most obvious in diabetic duct-ligated rabbits, in which case beta-D-glucose stimulates insulin release more efficiently than alpha-D-glucose. In both control and duct-ligated rabbits, however, the alpha-anomer is more potent than the beta-anomer in suppressing leucine-stimulated glucagon secretion. These findings are compatible with the view that chronic hyperglycemia leads to alteration in the anomeric preference of the pancreatic B-cell for alpha-D-glucose, possibly as a result of the nonenzymatic glycation of glycolytic enzymes in insulin-producing, but not glucagon-producing, islet cells.     

The effect of placental lactogen (HPL) on glucose-induced insulin release

Summary The possible effect of placental lactogen on the process of insulin secretion was investigated in rabbits by examining its action upon glucose-induced insulin secretion. Glucose injection (0.5 g/kg body weight) resulted in a prompt insulin response. When placental lactogen (5 mg/kg body weight) and glucose were injected simultaneously, no significant changes in blood glucose levels were observed as compared to those after glucose alone. Plasma insulin levels 5 min after the combined injection were higher than after glucose alone and the total insulin response appeared to be an additive effect of the two stimulating agents. Insulin secretionin vitro by pieces of pancreatic tissue from rabbits has been studied with and without the addition of glucose and HPL. The dynamics of insulin secretion were examined by sequential exposure of the same pieces of pancreatic tissue. After 60 min of preincubation, 25 μg/ml of partially purified HPL plus glucose significantly increases insulin release. It is suggested that the increase of B-cell secretion during pregnancy could be due, at least in part, to the stimulation of endogenous production of chorionic growth hormone. Traduzione a cura di G. U.     

The stimulus-secretion coupling of glucose-induced insulin release

Summary When isolated islets of Langerhans are suddenly exposed to glucose, the entry of the hexose into islet cells first occurs at a high rate resulting in rapid equilibration of free glucose across the cell membrane; thereafter, the rate of net glucose uptake depends on its metabolism. More than 95% of the glucose taken up by the islets is converted to triosephosphate. The fractional contribution of the sorbitol and pentose-phosphate pathways to such a process does not exceed 10%. The output of lactate from the islets accounts for approximately half of the glycolytic flux. At increasing glucose concentrations up to 4.3 mM, the rate of glycolysis increases towards a first asymptotic value; at higher glucose levels (up to 27.8 mM), a sigmoidal pattern is seen tending towards a second saturation value. The total ATP content of the islets does not correlate with their insulin-secretory activity. It is suggested that, in the process of glucose-induced insulin release, glycolysis may regulate physiological processes possibly located in the micro-environment of the cell boundary.     

Effect of a phospho-oligosaccharidic putative insulin messenger on insulin release in rats

Summary The phospho-oligosaccharide extracted from rat liver and supposed to act as the insulin second messenger inhibits glucose-stimulated insulin release. In the present study, this phospho-oligosaccharide was found not to affect D-[U-¹⁴C]glucose oxidation and ⁴⁵Ca net uptake, but to inhibit insulin release evoked by either D-glucose or 2-ketoisocaproate in isolated rat islets. The relative extent of the latter inhibition was unaffected by either the concentration of D-glucose or the presence of dibutyryl-cyclic AMP, forskolin or glucagon in the incubation medium. At variance with the inhibitory effect of clonidine, that of the phospho-oligosaccharide was resistant to both blockade of ₂-adrenergic receptors or pre-treatment with the toxin of Bordetella pertussis. It is speculated, therefore, that such a phospho-oligosaccharide might interfere with a distal event in the insulin secretory sequence.     

The stimulus-secretion coupling of glucose-induced insulin release

Summary When isolated rat islets were exposed to glucose, the concentrations of NADH and NADPH, and the NADH/NAD⁺ and NADPH/NADP⁺ ratios were increased. The dose-response curve resembled that characterising the glucose-induced secondary rise in⁴⁵Ca efflux, displaying a sigmoidal pattern with a half-maximal value at glucose 7.5 mmol/l. The glucose-induced increase in NAD(P)H was detectable within 1 min of exposure to the sugar. Except for the fall in ATP concentration and ATP/ADP ratio found at very low glucose concentrations (zero to 1.7 mmol/l) no effect of glucose (2.8–27.8 mmol/l) upon the steady-state concentration of adenine nucleotides was observed. However, a stepwise increase in glucose concentration provoked a dramatic and transient fall in the ATP concentration, followed by a sustained increase in both O₂ consumption and oxidation of exogenous + endogenous nutrients. This may be essential to meet the energy requirements in the stimulated B-cell. Although no significant effect of glucose upon intracellular pH was detected by the 5,5-dimethyloxazolidine-2,4-dione method, the net release of H⁺ was markedly increased by glucose, with a hyperbolic dose-response curve (half-maximal response at glucose 2.9 mmol/l) similar to that characterising the glucose-induced initial fall in⁴⁵Ca efflux. It is proposed that the generation of both NAD(P)H and H⁺ participates in the coupling of glucose metabolism to distal events in the secretory sequence, especially the ionophoretic process of Ca²⁺ inward and outward transport, and that changes in these parameters occur in concert with an increased turn-over rate of high-energy phosphate intermediates.Recipient of a Pfizer Traval Award through the European Association for the Study of DiabetesOn leave from the Fundacion Jimenez Diaz, Madrid, Spain     

Melatonin restores endothelium-dependent relaxation in aortic rings of pancreatectomized rats

In rats turned hyperglycemic by a subtotal pancreatectomy, a decreased relaxation response of aortic rings to acetylcholine (ACh) was found; this effect was amplified by preincubation in a high glucose medium (44 mmol/L). The relaxation response to ACh did not occur in endothelium-denuded rings or after the aortic rings were exposed to l-nitro-arginine methyl ester [L-NAME, a nitric oxide (NO) synthase inhibitor]. Incubation with the NO donor sodium nitroprusside (SNP) restored the impaired relaxation response seen in endothelium-denuded or L-NAME-treated aortic rings. Pancreatectomy decreased the vasorelaxation of aortic rings caused by SNP. Only in pancreatectomized rats, incubation in a high glucose medium impaired the relaxation effect of SNP. To assess whether melatonin preincubation reversed the impaired relaxation response to ACh (intact endothelium aortic rings) or to SNP (endothelium-denuded or L-NAME-treated rings) in hyperglycemic rats, cumulative dose-response curves were performed in the presence of 10(-5) mol/L melatonin. Melatonin preincubation did not modify ACh-induced relaxation of aortic rings in a normal glucose concentration but was highly effective in preventing the impairment of relaxation caused by a high glucose solution. Melatonin was also effective in restoring the impaired SNP-induced vasorelaxation seen in endothelium-denuded or L-NAME-treated aortic rings from hyperglycemic rats. The results further support the improvement by melatonin of the endothelial-mediated relaxation in blood vessels of diabetic rats.     

The stimulus-secretion coupling of glucose-induced insulin release

Summary Above a threshold of 3.0–4.2 mmol/l, D-glucose provoked a transient increase in ³²P fractional outflow rate from rat pancreatic islets prelabelled with ³²P-orthophosphate. Nutrients which stimulate insulin release in the absence of glucose, -ketoisocaproate and L-leucine, also provoked a phosphate flush. No flush occurred in islets exposed to non-insulinotropic nutrients (L-glutamine and L-lactate) or non-nutrient secretagogues (arginine, tolbutamide, theophylline). A late increase in ³²P fractional outflow rate was observed in Ca²⁺ deprived islets stimulated with BaCl₂ and theophylline. The occurrence of a phosphate flush did not appear to be attributable to changes in insulin release, cyclic AMP content, membrane polarisation, K⁺ conductance, or reduced pyridine nucleotide content. The ³²P response to glucose was slightly decreased in the absence of extracellular Ca²⁺ or HCO₃ ^-, markedly impaired in the absence of K⁺, and virtually abolished in the presence of menadione (10 mol/l). It is proposed that the occurrence of a phosphate flush is linked to the metabolism of nutrient secretagogues, possibly via an increase in O₂ uptake and the production rate of NAD(P)H and ATP.     

Nicotinamide partially reverses the interleukin-1 beta inhibition of glucose-induced insulin release in pancreatic islets.

Interleukin-1 beta (IL-1 beta) is known to inhibit glucose-induced insulin release by pancreatic islets. We studied the effect of nicotinamide, an inhibitor of poly[adenosine diphosphate (ADP)-ribose] synthetase and a free-radical scavenger, on this IL-1 beta-induced inhibition using rat pancreatic islets. In static experiments, groups of five islets were incubated for 24 hours in culture medium CMRL-1066, with or without 50 U/mL IL-1 beta, in the presence or absence of nicotinamide (dose range, 0 to 50 mmol/L), and then exposed for 1 hour to either 1.4 or 19.4 mmol/L glucose, 10 mmol/L arginine, or 10 mumols/L glyburide. Basal insulin secretion was 183 +/- 32 pg/islet/h (mean +/- SE, n = 7) and 176 +/- 39 (n = 7) in control islets and in islets exposed to 50 U/mL IL-1 beta, respectively. Glucose-stimulated insulin secretion was significantly reduced (185 +/- 41) in IL-1 beta-exposed islets in comparison to control islets (2,037 +/- 363). In parallel, arginine-stimulated insulin release was inhibited by IL-1 beta exposure (166 +/- 31 pg/islet/h, mean +/- SE, n = 3) in comparison to control islets (1,679 +/- 307). In contrast, IL-1 beta exposure did not significantly reduce glyburide-induced insulin secretion (1,516 +/- 231 and 1,236 +/- 214 in control and IL-1 beta-exposed islets, respectively; mean +/- SE, n = 3). When islets were simultaneously exposed to IL-1 beta and increasing concentrations of nicotinamide, a dose-dependent recovery of glucose-induced insulin secretion was observed, with the maximum effect at 25 mmol/L nicotinamide (1,007 +/- 123, P less than .001).(ABSTRACT TRUNCATED AT 250 WORDS)     

Indomethacin treatment causes loss of insulin action in rats: involvement of prostaglandins in the mechanisms of insulin action

Glucose tolerance tests in rats showed that after indomethacin treatment plasma insulin levels rose five-fold higher than in untreated controls. Accordingly, the pancreatic islets of indomethacin-treated rats secreted insulin at a threefold higher rate. Glucose tolerance tests additionally showed that indomethacin treatment led to a retarded disposal of the elevated blood glucose. Both effects appear to be caused by an attenuation of the hormone responsiveness for insulin and noradrenaline (-adrenoceptor action) by indomethacin. The following observations support this view: insulin and adrenaline (-adrenoceptor action) lost their ability to lower cyclic adenosine monophosphate (AMP) levels in hepatocytes; the glycogen content of liver and skeletal muscle was reduced by 95% and 65%, respectively; in adipocytes the stimulation of glucose transport by insulin was reduced by 60%. These effects of indomethacin can be reversed by the addition of exogenous prostaglandin E (PGE), as elevated cyclic AMP systhesis was again sensitive to -adrenergic inhibition in the liver: These results indicate a relationship between prostaglandins and insulin action. These effects of indomethacin could result from reduced synthesis of cyclic PIP (prostaglandylinositol cyclic phosphate), a proposed second messenger for insulin and -adrenoceptor action, whose synthesis was decreased by indomethacin treatment and increased by the addition of exogenous PGE. Stimulation of glucose transport by cyclic PIP was unaffected by indomethacin treatment, in contrast to the stimulation by insulin. Inhibition of PGE and cyclic PIP synthesis resulted in a metabolic state comparable to insulin resistance in non-insulin-dependent diabetes mellitus.Presented in part at the following meetings: Mechanisms in the Action of Insulin, Oslo (Norway) 1987; 50th Annual Meeting of the American Diabetes Association, Atlanta (USA) 1990 [1]; 34. Symposium Deutsche Gesellschaft für Endokrinologie, Hannover (Germany) 1990 [2]; 28. Jahrestagung der Deutschen Diabetes-Gesellschaft, Ulm (Germany) 1993 [3]     

Hyperglycemia and inhibition of glucose-induced insulin release in 6-aminonicotinamide treated rats

The effects of 6-aminonicotinamide (6-AN) on blood glucose and insulin release were studied in rats. 6-AN at 4mg/100g body weight slowly raised blood glucose concentrations to a significantly higher level than the control values 6 hours after an intraperitoneal injection. At this time severe glucose intolerance and low IRI response were noticed during an intravenous glucose tolerance test. Adrenalectomized rats replaced by hydrocortisone also presented hyperglycemia, glucose intolerance and low IRI response during IVGTT under treatment with 6-AN but to a lesser extent than in the intact rats. In in vitro experiments, decreased insulin release from the perfused pancreata of rats pretreated with 6-AN was found both in the 1st and 2nd phases of response to glucose stimulation. These data indicate that 6-AN-induced hyperglycemia is attributed to the inhibition of insulin release in adrenalectomized rats except for the hypothetical effect of 6-AN which diminishes an action of insulin on cellular glucose transport.     

依折麦布对糖尿病前期不稳定性心绞痛患者胰岛素释放的影响

目的观察依折麦布对糖尿病前期不稳定性心绞痛患者胰岛素释放的影响。方法选取糖尿病前期不稳定性心绞痛患者222例,随机分为对照组和依折麦布组,对照组给予阿托伐他汀治疗;依折麦布组给予阿托伐他汀联合依折麦布治疗,分别治疗7天、1月、3月,比较两组葡萄糖耐量试验(OGTT)和胰岛素释放试验(IRT)结果。结果与对照组比较,依折麦布组7天、1月、3月OGTT 0.5 h、1 h、2 h血糖显著降低(P0.05、0.01),0.5 h、1 h、2 h、3 h血糖曲线下面积显著减少(P0.05、0.01、0.001);IRT 0.5 h、1 h胰岛素明显升高(P0.05、0.01),2 h胰岛素降低(P0.05),0.5 h、1 h、2 h胰岛素曲线下面积增加(P0.01、0.001、0.05)。不同时间点血糖曲线下面积和胰岛素曲线下面积存在负相关(r=-0.387,P0.01);不同时间段血糖减低量与胰岛素增加量存在正相关(r=0.473,P0.001)。与对照组比较,依折麦布组稳态模型胰岛素抵抗指数于1月、3月明显降低(P0.05、0.01);胰岛素作用指数于7天、1月、3月明显升高(P0.05、0.01);稳态模型-β于3月明显升高(P0.05)。结论依折麦布改善糖尿病前期不稳定性心绞痛患者胰岛素分泌,提高胰岛素敏感性,改善糖耐量。     

Fentanyl inhibits glucose-stimulated insulin release from β-cells in rat pancreatic islets

AIM： TO explore the effects of fentanyl on insulin release from freshly isolated rat pancreatic islets in static culture. METHODS： Islets were isolated from the pancreas of mature Sprague Dawley rats by common bile duct intraductal collagenase V digestion and were purified by discontinuous Ficoll density gradient centrifugation. The islets were divided into four groups according to the fentanyl concentration： control group （0 ng/mL）, group I （0.3 ng/mL）, group I （3.0 ng/mL）, and group III （30 ng/mL）. In each group, the islets were co-cultured for 48 h with drugs under static conditions with fentanyl alone, fentanyl ＋ 0.1 μg/mL naloxone or fentanyl ＋ 1.0 μg/mL naloxone. Cell viability was assessed by the MTT assay. Insulin release in response to low and high concentrations （2.8 mmol/L and 16.7 mmol/L, respectively） of glucose was investigated and electron microscopy morphological assessment was performed. RESULTS： Low- and high-glucose-stimulated insulin release in the control group was significantly higher than in groups I and II （62.33 ± 9.67 μIU vs 47.75 ± 8.47 μIU, 39.67 ± 6.18 μIU and 125.5 ± 22.04 μIU vs 96.17 ± 14.17 μIU, 75.17 ± 13.57 μIU, respectively, P 〈 0.01） and was lowest in group III （P 〈 0.01）. After adding 1 μg/mL naloxone, insulin release in groups II and II was not different from the control group. Electron microscopy studies showed that the islets were damaged by 30 ng/ml fentanyl. CONCLUSION： Fentanyl inhibited glucose-stimulated insulin release from rat islets, which could be prevented by naloxone. Higher concentrations of fentanyl significantly damaged β-cells of rat islets.     

Effect of cholera toxin on insulin release in monolayer cultures of the endocrine pancreas

Summary The effect of cholera toxin on insulin release by monolayer cultures of endocrine pancreas has been studied. The toxin has a marked stimulatory effect upon insulin release at concentrations as low as 10^–10M. The toxin had a small effect at low glucose concentrations, but was strongly stimulatory at high glucose concentrations and in the presence of arginine. Its effect could be detected within 30 min of application and only two minutes' exposure to the toxin was required for it to subsequently stimulate release. In comparative studies on insulin release the toxin was equal to, or slightly more potent than, 1.5 M glucagon and significantly more potent than cyclic AMP (10 mM) or theophylline (5 mM).     

周围血单个核细胞抑制胰岛素释放的实验研究

将新发病的ＩＤＤＭ病人周围血单个核细胞（ＰＢＭＣ）同大鼠胰岛共同培养２０个小时后用Ｌ－精氨酸刺激，收集并测定基础和刺激后培养上清中胰岛素的含量，结果显示：ＩＤＤＭ病人ＰＢＭＣ作用下的基础胰岛素释放（１１７．９±１４．０±μＵ·１０ｉｓｌｅｔｓ－１／２０ｈ）（ｎ＝１１）和刺激后胰岛素释放（１４７．５±３２．３μＵ·１０ｉｓｌｅｔｓ－１／３ｈ）（ｎ＝１１），显著低于正常人ＰＢＭＣ作用下的基础胰岛素释放（１８４．８±２９．５μＵ）（ｎ＝１０，Ｐ＜０．０１）和刺激后胰岛素释放（１９５．０±２７．４μＵ）（ｎ＝１０，Ｐ＜０．０１）。结果表明新发病的ＩＤＤＭ病人ＰＢＭＣ能够抑制大鼠胰岛基础和刺激后胰岛素的释放。     

Effect of in vivo vanadate treatment on insulin receptor tyrosine kinase activity in partially pancreatectomized diabetic rats

Liver-purified insulin receptor tyrosine kinase (IRTK) activity was examined in partially pancreatectomized rats following normalization of blood glucose concentration by either phlorizin or vanadate treatment. Chronic moderate hyperglycemia did not modify the IRTK activity, despite the presence of in vivo and in vitro insulin resistance. Oral vanadate administration for 3 weeks normalized glucose tolerance and caused a 2.5-fold increase in basal IRTK activity. In contrast, correction of hyperglycemia with phlorizin, an inhibitor of renal glucose reabsorption, did not change the IRTK activity, although glucose tolerance was returned to normal. The vanadate-induced effect on basal IRTK was due to an increase in Vmax of the enzyme; the Km remained unchanged. The insulin-stimulated IRTK activity was not affected by either vanadate or phlorizin treatment. These results suggest that: 1) partial (90%) pancreatectomy in rats causes insulin resistance in the absence of in vitro alterations in IRTK and 2) correction of chronic hyperglycemia with vanadate, but not with phlorizin, is associated with an increased basal activation of the protein tyrosine kinase in liver insulin receptors.     

Effects of glimepiride on insulin and glucagon release from isolated rat pancreas at different glucose concentrations

The effects of glimepiride, the newest sulphonylureic compound, on pancreatic insulin and glucagon secretion were studied using the classical, isolated, perfused rat pancreas model. The influence of four different environmental glucose conditions (during a glycaemic stimulus with glucose increasing from 5 to 8.33 mM and at stable 0, 5 and 2.22 mM glucose levels) on the effects of glimepiride was also assessed. At a pharmacological concentration glimepiride strongly stimulated beta-cell activity, producing a characteristic biphasic insulin release with a sharp first-phase secretory peak, followed by a prolonged and sustained second phase. Environmental glucose concentrations markedly influenced the extent, but not the pattern of glimepiride-induced insulin secretion, as hormone release dropped significantly when the glucose level was reduced. Glimepiride failed to influence alpha-cell activity at any of the environmental glycaemic levels.     

Hyperresponse in calcium-induced insulin release from electrically permeabilized pancreatic islets of diabetic GK rats and its defective augmentation by glucose

Summary In spontaneously diabetic GK rats, insulin secretion from pancreatic beta cells in response to glucose is selectively impaired, probably due to deficient intracellular metabolism of glucose and impaired closure of K_ATP channels during glucose stimulation. By using electrically permeabilized islets of GK rats, we explored the functional modulations in exocytotic steps distal to the rise in [Ca^{2 +} ]i in the diabetic condition. At 30 nmol/l Ca^{2 +} (basal conditions) insulin release was similar between GK and non-diabetic control Wistar rats. In response to 3.0 μmol/l Ca^{2 +} (maximum stimulatory conditions), insulin release was significantly augmented in permeabilized GK islets (p < 0.01). Raising glucose concentrations from 2.8 to 16.7 mmol/l further augmented insulin release induced by 3.0 μmol/l Ca^{2 +} from permeabilized control islets(p < 0.001), but had no effect on that from permeabilized GK islets. The stimulatory effect of glucose on insulin release from permeabilized control islets was partly inhibited by 2,4-dinitrophenol, an inhibitor of mitochondrial oxidative phosphorylation (p < 0.01). The hyperresponse to Ca^{2 +} in GK islets may play a physiologically compensatory role on the putative functional impairment both in [Ca^{2 +} ]i rise and energy state in response to glucose in diabetic β cells, and may explain the relative preservation of insulin release induced by non-glucose depolarizing stimuli, such as arginine, from pancreatic islets in non-insulin-dependent diabetes mellitus. [Diabetologia (1995) 38: 772–778] Received: 17 September 1994 and in revised form: 29 December 1994