GLP-1 Concentrations Research Articles

The influence of the colon on postprandial glucagon-like peptide 1 (7-36) amide concentration in man.

Glucagon-like peptide (7-36) amide (GLP-1) is an incretin hormone of the enteroinsular axis released rapidly after meals despite the fact that GLP-1 secreting cells (L-cells) occur predominantly in the distal gut. The importance of these colonic L-cells for postprandial GLP-1 was determined in healthy control subjects and in ileostomy patients with minimal small bowel resection (<5 cm). Subjects were fed a high complex carbohydrate test meal (15.3 g starch) followed by two carbohydrate-free, high fat test meals (25 g and 48.7 g fat respectively). Circulating levels of glucose, insulin, glucagon, glucose insulinotrophic peptide (GIP) and GLP-1 were measured over a 9-h postprandial period. For both subject groups the complex carbohydrate test meal failed to elicit a rise in either GIP or GLP-1. However, both hormones were elevated after the fat load although the GLP-1 concentration was significantly reduced in the ileostomist group when compared with controls (P=0.02). Associated with this reduction in circulating GLP-1 was an elevation in glucagon concentration (P=0.012) and a secondary rise in the plasma glucose concentration (P=0.006). These results suggest that the loss of colonic endocrine tissue is an important determinant in the postprandial GLP-1 concentration. Ileostomists should not be assumed to have normal enteroinsular function as the colon appears to have an important role in postprandial metabolism.

Oral Triacylglycerols Regulate Plasma Glucagon-Like Peptide-1(7–36) and Insulin Levels in Normal and Especially in Obese Rats

In a previous study of glucose tolerance, plasma insulin levels were greatly elevated in genetically obese Wistar fatty rats but not lean rats fed a diet containing polyunsaturated fatty acids. In the present study, triacylglycerol-regulation of levels of circulating insulin and glucagon-like peptide-1 (7–36) (GLP-1) has been investigated in these rats. In the glucose tolerance test, the two plasma insulin peaks appeared in obese and lean rats intubated with glucose + corn oil, at 15- 30 min and 4 h, whereas only the first peak appeared in rats intubated with glucose alone, although the glucose response did not differ. After intubation of corn oil only, the insulin peak at 15 min was not detected but the peak at 4h was large. The two plasma GLP-1 peaks appeared 15 min and 4 h after intubation of glucose + corn oil similarly to the insulin responses, although the first peak was small and the second peak was very large. A small peak at 15 min was not significant in rats intubated glucose alone and no peak was seen at 4 h. The GLP-1 concentrations were significantly higher in the following order: portal vein > inferior vena cava > tail vein. The plasma GLP-1 increment in response to oral triacylglycerols was significantly higher in obese rats than in lean rats as was the insulin increment. Thus, oral triacylglycerols (possibly polyunsaturated) appeared to act at the gut lumen to stimulate GLP-1 secretion, which may be responsible for the second (4 h) insulin peak.

Subcutaneous glucagon-like peptide-1 improves postprandial glycaemic control over a 3-week period in patients with early Type 2 diabetes

1.Glucagon-like peptide-1 (7-36) amide (GLP-1) is released into the circulation after meals and is the most potent physiological insulinotropic hormone in man. GLP-1 has the advantages over other therapeutic agents for Type 2 diabetes of also suppressing glucagon secretion and delaying gastric emptying. One of the initial abnormalities of Type 2 diabetes is the loss of the first-phase insulin response, leading to postprandial hyperglycaemia.2. To investigate the therapeutic potential of GLP-1 in Type 2 diabetes, six patients were entered into a 6-week, double-blind crossover trial during which each received 3 weeks treatment with subcutaneous GLP-1 or saline, self-administered three times a day immediately before meals. A standard test meal was given at the beginning and end of each treatment period.3.GLP-1 reduced plasma glucose area under the curve (AUC) after the standard test meal by 58% (AUC, 0-240 min: GLP-1 start of treatment, 196+/-141 mmol.min-1.l-1; saline start of treatment, 469+/-124 mmol.min-1.l-1; F=16.4, P<0.05). The plasma insulin excursions were significantly higher with GLP-1 compared with saline over the initial postprandial 30 min, the time period during which the GLP-1 concentration was considerably elevated. The plasma glucagon levels were significantly lower over the 240-min postprandial period with GLP-1 treatment. The beneficial effects of GLP-1 on plasma glucose, insulin and glucagon concentrations were fully maintained for the 3-week treatment period. 4. We have demonstrated a significant improvement in postprandial glycaemic control with subcutaneous GLP-1 treatment. GLP-1 improves glycaemic control partially by restoring the first-phase insulin response and suppressing glucagon and is a potential treatment for Type 2 diabetes.

Systemic short-chain fatty acids rapidly alter gastrointestinal structure, function, and expression of early response genes.

Luminal and systemic short chain fatty acids (SCFA) stimulate mucosal proliferation but the mechanism(s) is unclear. This study examined acute effects of systemic SCFAs on gastrointestinal structure and function and signals potentially mediating SCFA-induced mucosal proliferation. Male Sprague-Dawley rats (246+/-2 g) received nutrients as either standard total parenteral nutrition (TPN) or an isoenergetic, isonitrogenous formulation containing SCFAs (TPN + SCFA). Animals were randomized to one of five treatments: standard TPN for 72 hr, TPN + SCFA for 72 hr, or standard TPN followed by TPN + SCFA for the final 6, 12, and 24 hr. SCFAs reduced (P < 0.003) ileal protein within 6 hr. Jejunal GLUT2 expression was increased (P=0.0001) in all SCFA groups and ileal GLUT2 protein in the 6-, 12-, and 24-hr SCFA groups (P < 0.05). SCFAs increased (P < 0.003) ileal proglucagon abundance following 6, 12, and 24 hr, and plasma GLP-2 concentration following 12 hr (P < 0.03). Jejunal c-myc expression was increased (P < 0.001) following 6, 12, and 24 hr of SCFAs. SCFAs increased ileal c-myc, c-jun, and c-fos expression following 24 hr (P < 0.02), 12 hr (P < 0.05) and 6, 12, and 24 hr (P=0.0001), respectively. In conclusion, systemic SCFAs increase plasma GLP-2 and ileal proglucagon mRNA, GLUT2 expression and protein, and c-myc, c-jun, and c-fos expression.

GLP-1 Tablet in Type 2 Diabetes in Fasting and Postprandial Conditions

To examine the absorption of glucagon-like peptide (GLP)-1(7-36) amide from the buccal mucosa of type 2 diabetic patients. Previously, the effects of the peptide have been studied following intravenous and subcutaneous injection. Now, a mucoadhesive, biodegradable buccal GLP-1 tablet (9 mm) containing 119 nmol has been developed as a possible alternative to injection. A total of 10 type 2 diabetic patients received a single tablet under fasting conditions and before a standard meal in this randomized placebo-controlled study. The mean peak GLP-1 concentration was 125.1 pmol/l and occurred 30 min after application. The mean placebo-adjusted area under the curve was 5,334 min pmol/l, consistent with a relative bioavailability of 6% vs. intravenous injection and 42% vs. subcutaneous injection. The half-life of total peptide activity after buccal administration was 17 min. The placebo-adjusted glucose concentrations decreased by 1.4 mmol/l in fasting experiments and by 4.2 mmol/l after a standard mixed meal. In the fasting state at 30 min, plasma insulin increased by 185% and glucagon decreased by 20%, consistent with the increase in plasma GLP-1 concentrations. The peptide exerted a significant insulinotropic effect during meals (calculated as an insulinogenic index, 0-120 min; 84.1 vs. 45.7 in placebo experiments). Potentially therapeutic plasma levels of GLP-1 were achieved after administration of a single buccal tablet in type 2 diabetic patients. The peptide had a marked glucose-lowering effect during the first 2 h. This new GLP-1 tablet may become a feasible alternative treatment for type 2 diabetic patients, although a more prolonged pharmacokinetic profile is required.

Elevated post-prandial gastric inhibitory polypeptide concentrations in hypertriglyceridaemic subjects.

1. We investigated whether abnormalities of gastric inhibitory polypeptide (GIP) and glucagon-like peptide-1 (7-36 amide) (GLP-1) contribute to the hypertriglyceridaemia and hyperinsulinaemia in hypertriglyceridaemic subjects. Serum triglycerides and plasma glucose GIP, GLP-1 and immunoreactive insulin (IRI) concentrations were measured before and after a mixed meal in 15 hypertriglyceridaemic patients and in eight healthy normotriglyceridaemic control subjects. 2. Integrated post-prandial GIP concentrations were greater than in controls (P < 0.05) and correlated positively with both fasting and integrated post-prandial triglyceride concentrations (P < 0.05 for both). Fasting and integrated post-prandial IRI levels were higher in hypertriglyceridaemic subjects than in controls (P < 0.02 and P < 0.05 respectively) and correlated positively with fasting triglycerides (P < 0.02 and P < 0.001 respectively) and integrated post-prandial triglycerides (P < 0.005 and P < 0.05 respectively). There was no correlation between GIP concentrations and either fasting or post-prandial IRI levels. Fasting and post-prandial concentrations of GLP-1 were similar in patients and controls. 3. Hypertriglyceridaemic subjects have post-prandial hyperGIPaemia in addition to the well-documented hyperinsulinaemia. We found no association between GIP and insulin. There is, however, clear evidence for an association between post-prandial GIP concentrations and triglyceride levels. We suggest that this association may depend on changes in lipoprotein lipase activity and that there may be a feedback loop between GIP and triglyceride lipolysis.

Glucagon-Like Peptide-1 7-36 Amide and Peptide YY Have Additive Inhibitory Effect on Gastric Acid Secretion in Man

Background: Glucagon-like peptide-1 7-36 amide (GLP-1) and peptide YY (PYY) are colocalized in the L-cell of the ileal mucosa, and both peptides may function as enterogastrone hormones. However, it is not known whether they interact with regard to the effect on acid secretion. Methods: The effect of intravenous infusion of GLP-1 and PYY, either alone or in combination, on pentagastrin-induced acid secretion in eight healthy volunteers was examined. The peptides were infused at two different rates: 0.25 pmol/kg/min (low rate) and 0.5 pmol/kg/min (high rate). Results: Given alone, GLP-1 and PYY inhibited acid secretion by 26 ± 5% and 18 ± 5% (low rate) and 45 ± 8% and 38 ± 7% (high rate), respectively. Combined infusion resulted in an inhibition of 32 ± 5% (low rate) and 62 ± 7% (high rate). Both infusion rates resulted in GLP-1 and PYY plasma concentrations below or similar to postprandial levels. Conclusion: The present study suggests that the interaction between GLP-1 and PYY in man is of the additive type. The results indicate that GLP-1 and PYY have an important role in the physiologic control of gastric acid secretion.

Effects of somatostatin-28 on circulating concentrations of insulin and gut hormones in sheep.

The effects of intravenous somatostatin-28 (S28) infusion on glucose-stimulated and glucagon-like peptide-1(7-36)amide (GLP-1)-augmented insulin secretion were studied in sheep. S28 was infused via a jugular catheter for 15 min at a rate of 1.1 pmol/kg/min either alone or together with GLP-1 and/or glucose. S28 infusion did not significantly lower circulating basal insulin concentrations in fed sheep. Glucose-stimulated insulin secretion was significantly inhibited by S28 infusion, serum concentrations decreasing from about 200 to 150 pmol/l. GLP-1 significantly augmented glucose-stimulated insulin secretion, serum concentrations increasing from about 230 to 280 pmol/l. S28 completely counteracted this effect of GLP-1. S28 infusion also significantly decreased the circulating concentrations of glucose-dependent insulinotrophic polypeptide (GIP) and GLP-1 in fed sheep (from about 110 to 45 pmol/l for GIP and from about 25 to 15 pmol/l for GLP-1). The physiological implications of these observations are discussed with particular reference to the ruminant. It is concluded that S28 may have an important endocrine role in the control of insulin secretion and regulation of nutrient partitioning.

Point Mutations in the First and Third Intracellular Loops of the Glucagon-like Peptide-1 Receptor Alter Intracellular Signaling

The glucagon-like peptide-1 receptor (GLP-1R) is a member of the glucagon family of seven transmembrane spanning receptors. To investigate how different portions of the GLP-1R may be important in cAMP and intracellular calcium signaling, single amino acid substitutions in the receptor were made by site directed mutagenesis. Receptor binding, cAMP, and intracellular calcium measurements were made in transfected COS-7 cells. The change of amino acid H180R (His to Arg) in the first intracellular loop caused a decrease in the affinity of binding of GLP-1 from 7 nM in the wild type receptor to 150 nM and resulted in a 50% decrease in GLP-1 stimulated cAMP production. In response to 10 nM GLP-1, the receptor's ability to stimulate intracellular calcium was altered from a prolonged to a transient response of the same magnitude. Mutation in the 3rd intracellular loop at position R348G (Arg to Gly) decreased receptor affinity from 7 to 83 nM and nearly abolished cAMP production at all concentrations of GLP-1 tested. The GLP-1 stimulated rise in free intracellular calcium was also diminished and this was reversed when cells were treated with forskolin. These results also indicate that GLP-1R signaling via intracellular calcium is dependent on the receptor's ability to also generate cAMP.

Absorption of glucagon-like peptide-1 can be protracted by zinc or protamine

Pharmacokinetics of several different preparations for subcutaneous administration of GLP-1(7–36)amide (GLP-1) were studied. The difference between soluble GLP-1 at pH 4.0 and a suspension of GLP-1 crystals at pH 6.9, and how addition of zinc chloride or protamine sulphate to the GLP-1 suspension affected the absorption kinetics after s.c. administration, were investigated. Four Beagle dogs received 50 μg/kg of GLP-1 s.c. in different vehicle compositions. The vehicle contained 1 mg/ml of GLP-1, 10 mmol/1 disodium phosphate, 16 mg/ml glycerol, and in some preparations either zinc chloride or protamine sulphate. The preparations were: (1) pH 4.0, (2) pH 6.9, (3) I equiv. (relative to GLP-1) zinc chloride pH 6.9, (4) 0.5 equiv. zinc chloride pH 6.9, (5) 3 equiv. zinc chloride pH 6.9. (6) 0.5 mg/ml protamine sulphate pH 4.0, and (7) 0.5 mg/ml protamine sulphate pH 6.9. Plasma samples were analyzed employing a sandwich enzyme-linked immunosorbant assay. Addition of zinc chloride or protamine sulphate to the GLP-1 suspension decreased the absorption rate. The plasma concentration of GLP-1 was increased to a pharmacological level for at least 15 h when administered s.c. as a zinc or protamine suspension. Furthermore, the peak plasma concentration was flattened out, when the peptide was administered as a suspension containing zinc chloride or protamine sulphate, compared to a solution.

Comparison of the glucose dependency of glucagon-like peptide-1(7–37) and glyburide in vitro and in vivo

The purpose of the present study was to compare the glucose dependency of the insulin secretagogue activity of the sulfonylurea, glyburide, versus that of glucagon-like peptide-1(7–37) [GLP-1(7–37)] in vitro and in vivo. In freshly isolated rat islets, maximally effective concentrations of glyburide (10 μmol/L) and GLP-1(7–37) (10 μmol/L) were equally effective in stimulating insulin secretion in the presence of 15 mmol/L glucose (2,4-fold increase relative to 15 mmol/L glucose alone). At 5 mmol/L glucose, both agents increased insulin secretion, but the effect for glyburide was threefold greater than for GLP-1(7–37) (122% and 41% increase in insulin secretion, respectively). In conscious catheterized rats infused with glucose at a variable rate to clamp plasma glucose concentration at 11 mmol/L, glyburide (1 mg/kg orally) and GLP-1(7–37) (infused intravenously [IV] at 5 pmol/min/kg) produced similar increases in insulin levels (1.8-fold relative to the respective vehicle controls) that were sustained through 60 minutes of measurement. These doses of GLP-1(7–37) and glyburide were then administered to fasted and fed rats (basal plasma glucose concentration, 5.8 and 7.3 mmol/L, respectively). Relative to the vehicle control group, GLP-1(7–37) infusion produced a transitory increase (30%) in plasma insulin concentration and a modest sustained decrease (10% to 20%) in glucose in both fasted and fed rats, whereas glyburide induced a sustained 2.4- and 1.7-fold increase in plasma insulin concentration in fasted and fed rats, respectively, and a 50% decrease in plasma glucose in both fasted and fed rats. Results of these studies demonstrate the higher glucose threshold for the insulin secretagogue activity of GLP-1(7–37) relative to glyburide in vitro and in vivo.

Glucagon and Glucagon-like Peptides in Fishes

Glucagon and glucagon-like peptides (GLPs) are coencoded in the vertebrate proglucagon gene. Large differences exist between fishes and other vertebrates in gene structure, peptide expression, peptide chemistry, and function of the hormones produced. Here we review selected aspects of glucagon and glucagon-like peptides in vertebrates with special focus on the contributions made by analysis of piscine systems. Our topics range from the history of discovery to gene structure and expression, through primary structures and regulation of plasma concentrations to physiological effects and message transduction. In fishes, the pancreas synthesizes glucagon and GLP-1, while the intestine may contribute oxyntomodulin, glucagon, GLP-1, and GLP-2. The pancreatic gene is short and lacks the sequence for GLP-2. GLP-1, which is produced exclusively in its biologically active form, is a potent metabolic hormone involved in regulation of liver glycogenolysis and gluconeogenesis. The responsiveness of isolated hepatocytes to glucagon is limited to high concentrations, while physiological concentrations of GLP-1 effectively regulate hepatic metabolism. Plasma concentrations of GLP-1 are higher than those of glucagon, and liver is identified as the major site of removal of both hormones from fish plasma. Ultimately, GLP-1 and glucagon exert effects on glucose metabolism that directly and indirectly oppose several key actions of insulin. Both glucagon and GLP-1 show very weak insulinotropic activity, if any, when tested on fish pancreas. Intracellular message transduction for glucagon, especially at slightly supraphysiological concentrations, involves cAMP and protein kinase A, while pathways for GLP are largely unknown and may involve a multitude of messengers, including cAMP. In spite of fundamental differences in GLP-1 function between fishes and mammals, fish GLP-1 is as powerful an insulinotropin for mammalian B-cells as mammalian GLP-1 is a metabolic hormone if tested on piscine liver.

Plasma and intestinal concentrations of GIP and GLP-1 (7-36) amide during suckling and after weaning in pigs.

Plasma concentrations of glucose dependent insulinotropic polypeptide (GIP) and glucagon-like peptide-1(7-36)amide (GLP-1[7-36]amide) were measured after milk ingestion in 15-18 day old piglets and after weaning diet ingestion in 33 day old piglets weaned at 21 days. Intestinal concentrations of these two hormones were also measured in unsuckled piglets of less than 24 h of age, and piglets whose ages corresponded with those used for plasma measurements. Suckling piglets showed a moderate glycaemic and insulinaemic response to milk ingestion. Plasma GIP and GLP-1(7-36)amide levels were significantly elevated at 1 and 3-h post-prandially. Weaned piglets showed a much more marked glucose and insulin response to meal ingestion. Plasma GIP and GLP-1(7-36)amide levels were again significantly elevated at 1 and 3 h in these animals. The mean plasma GIP response was greater in the weaned animals compared with the suckling animals at the time points investigated. The plasma GLP-1(7-36)amide response in contrast was significantly greater at 1 h in the suckling animals. In comparison, GIP concentrations in acid ethanol extracts of the small intestine were significantly higher during suckling and GLP-1(7-36)amide concentrations significantly higher after weaning. The circulating levels of both hormones seen during suckling and after weaning were far higher than those previously reported in humans. We conclude that both milk ingestion and the weaning diet are capable of stimulating GIP and GLP-1(7-36)amide in piglets and suggest that the levels of both hormones seen in this study may be important in adipose tissue metabolism at this time.

Structural characterization by affinity cross-linking of glucagon-like peptide-1(7-36)amide receptor in rat brain.

Specific binding of glucagon-like peptide (GLP)-1(7-36)amide was detected in several rat brain areas, with the highest values being found in hypothalamic nuclei and the nucleus of the solitary tract. In hypothalamus and brainstem homogenate binding of 125I-GLP-1(7-36)amide was time, temperature, and protein content dependent and was inhibited by unlabeled proglucagon-derived peptides. The rank order of potency was GLP-1(7-36)amide >> GLP-1(1-36)amide > GLP-1(1-37) approximately equal to GLP-2 > glucagon. Scatchard analysis of the steady-state binding data was consistent with the presence of both high- and low-affinity binding sites in hypothalamus and brainstem. Brain 125I-GLP-1(7-36)amide-binding protein complexes were covalently cross-linked using disuccinimidyl suberate and analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. A single radiolabeled band of M(r) 56,000 identified in both hypothalamus and brainstem homogenates was unaffected by reducing agents. An excess of unlabeled GLP-1(7-36)amide abolished the band labeling, whereas glucagon had no effect. Other unlabeled GLPs inhibited M(r) 56,000 complex labeling with the following order of potency: GLP-1(1-36)amide > GLP-1(1-37) > GLP-2. The binding of 125I-GLP-1(7-36)amide and the intensity of the cross-linked band were similarly inhibited in a dose-response manner by increasing concentrations of unlabeled GLP-1(7-36)amide. Covalent M(r) 56,000 125I-GLP-1(7-36)amide-binding protein complexes solubilized by Triton X-100 were adsorbed onto wheat germ agglutinin.(ABSTRACT TRUNCATED AT 250 WORDS)

Secretion of glucagon-like peptide-1 and reactive hypoglycemia after partial gastrectomy.

Glucagon-like peptide-1 is a peptide hormone from the distal small intestine which stimulates insulin secretion and inhibits glucagon secretion and thereby lowers blood glucose. This hormone, therefore, could be involved in the pathogenesis of postprandial reactive hypoglycemia. We subjected 8 patients showing symptoms of early dumping after partial gastrectomy to an oral 100 g glucose load and measured blood glucose and plasma insulin, C-peptide, glucagon, enteroglucagon and GLP-1 concentrations for 3.5 h after ingestion. Ten matched controls were treated similarly. The patients had higher blood glucose concentrations for the initial 60 min, but lower values for the remaining test period with a nadir of 2.76 +/- 0.19 mmol/l at 146 +/- 17 min after glucose (controls: 3.36 +/- 0.21 mmol/l at 210 +/- 9 min). GLP-1 and enteroglucagon responses were grossly elevated in patients compared to controls and insulin and C-peptide levels were higher during the initial 60 min. Glucagon concentrations increased in patients and decreased in controls. When hypoglycemia occurred, GLP-1 levels were only moderately elevated and insulin and C-peptide levels were lower and glucagon levels higher in patients than in controls. Thus, mechanisms other than release of GLP-1 seem to be responsible for the observed changes in the concentrations of glucose and glucoregulatory hormones.

Glucose-dependency of the insulin stimulatory effect of glucagon-like peptide-1 (7-36) amide on the rat pancreas.

The glucose-dependent action of GLP-1 (7-36) amide (GLP-1) on insulin secretion was studied in isolated islets and in the perfused rat pancreas. In islet experiments in the presence of non-stimulatory glucose levels (< 3 mmol/l) a GLP-1 concentration of 10 nmol/l increased insulin secretion by 83%. However, higher GLP-1 concentrations (25 and 100 nmol/l) could not further enhance this effect (85 and 83%, respectively). The onset of the stimulatory action of a supramaximal GLP-1-load (25 nmol/l) was at a glucose level of 3 mmol/l. In the perfused pancreas, 25 nmol/l GLP-1 induced a strong insulin release at 5 mmol/l glucose, but under basal glucose (2.8 mmol/l) only a slight enhancement of insulin secretion occurred during the late phase (30 to 54 min) of perfusion (P < 0.05). In conclusion, a slight but not dose-dependent stimulation of insulin secretion by supramaximal GLP-1 loads under basal glucose levels was found. The necessary GLP-1 concentrations to achieve this in vitro effect are beyond physiological or postprandial levels.

Truncated glucagon-like peptide-1 interacts with exendin receptors on dispersed acini from guinea pig pancreas. Identification of a mammalian analogue of the reptilian peptide exendin-4.

To find mammalian analogues of exendin-4, a peptide from Helodermatidae venoms that interacts with newly discovered exendin receptors on dispersed acini from guinea pig pancreas, we examined the actions of recent additions to the vasoactive intestinal peptide/secretin/glucagon family of regulatory peptides. In every respect tested, the truncated form of glucagon-like peptide-1, GLP-1(7-36)NH2, mimicked the actions of exendin-4. Like exendin-4, GLP-1(7-36)NH2 caused an increase in acinar cAMP without stimulating amylase release. GLP-1(7-36)NH2-induced increases in cAMP were inhibited progressively by increasing concentrations of the specific exendin-receptor antagonist, exendin(9-39)NH2. In dispersed acini from guinea pig and rat pancreas, concentrations of GLP-1(7-36)NH2 that stimulated increases in cAMP caused potentiation of cholecystokinin-induced amylase release. Binding of 125I-[Y39]exendin-4 or 125I-GLP-1(7-36)NH2 to dispersed acini from guinea pig pancreas was inhibited by adding increasing concentrations of unlabeled exendin-4 or GLP-1(7-36)NH2. We conclude that the mammalian peptide GLP-1(7-36)NH2 interacts with exendin receptors on dispersed acini from guinea pig pancreas. Exendin(9-39)NH2, a competitive antagonist of the actions of GLP-1(7-36)NH2 in pancreatic acini, may be a useful tool for examining the physiological actions of this peptide.

Glucose-dependency of the insulin stimulatory effect of glucagon-like peptide-1(7–36)amide on the rat pancreas

The glucose-dependent action of GLP-1 (7–36) amide (GLP-1) on insulin secretion was studied in isolated islets and in the perfused rat pancreas. In islet experiments in the presence of non-stimulatory glucose levels (<3 mmol/1) a GLP-1 concentration of 10 nmol/1 increased insulin secretion by 83%. However, higher GLP-1 concentrations (25 and 100 nmol/l) could not further enhance this effect (85 and 83%, respectively). The onset of the stimulatory action of a supramaximal GLP-1-load (25 nmol/l) was at a glucose level of 3 mmol/l. In the perfused pancreas, 25 nmol/l GLP-1 induced a strong insulin release at 5 mmol/l glucose, but under basal glucose (2.8 mmol/l) only a slight enhancement of insulin secretion occurred during the late phase (30 to 54 min) of perfusion (P<0.05). In conclusion, a slight but not dose-dependent stimulation of insulin secretion by supramaximal GLP-1 loads under basal glucose levels was found. The necessary GLP-1 concentrations to achieve this in vitro effect are beyond physiological or postprandial levels.

Lack of Effect of Synthetic Human Gastric Inhibitory Polypeptide and Glucagon-LikePeptide 1 [7-36 Amide] Infused at Near-Physiological Concentrations on Pentagastrin-Stimulated Gastric Acid Secretion in Normal Human Subjects

Gastric inhibitory polypeptide (GIP) and glucagon-like peptide 1 [7-36 amide] (GLP-1) are glucose-dependent insulinotropic gut hormones. Under experimental conditions, both have been shown to reduce stimulated gastric acid secretion. To study their individual and combined effects on pentagastrin-stimulated (0.1 micrograms/kg/h from -90 to 120 min) gastric volume, acid and chloride output, on separate occasions, synthetic human GIP (1 pmol/kg/min) and/or GLP-1 [7-36 amide] (0.3 pmol/kg/min) or placebo (0.9% NaCl with 1% albumin) were infused intravenously (from -30 to 120 min) in 9 healthy volunteers. At 0 min, a glucose infusion was started that mimicked the glycemic profile after an oral glucose load of 50 g/400 ml and allowed for the glucose-dependent insulinotropic action of GIP and GLP-1 [7-36 amide]. Pentagastrin stimulated acid output significantly, but neither GIP nor GLP-1 [7-36 amide] either alone or in combination, reduced pentagastrin-stimulated gastric acid secretion. The circulating concentrations of GIP and GLP-1 [7-36 amide] obtained at steady state during exogenous administration of synthetic peptides were similar to or higher than those reached after oral glucose (endogenous secretion). In conclusion, (penta)gastrin-stimulated gastric acid secretion is not inhibited by physiological circulating concentrations of GIP or GLP-1 [7-36 amide]. Therefore, the insulinotropic action of these intestinal hormones is physiologically more important than their possible role as enterogastrone.

Internalization of glucagon-like peptide-1(7–36)amide in rat insulinoma cells

Glucagon-like peptide-1(7-36)amide [GLP-1(7-36)amide] is supposed to be an important physiologic incretin. Recently, high affinity receptors for GLP-1(7-36)amide have been demonstrated on rat insulinoma-derived RINm5F cells. The present study examined the internalization and degradation of the GLP-1-receptor complex. Internalization of the peptide was time- and temperature-dependent. At 37 degrees C binding and internalization was rapid. At 60 min 35% of 125I-labeled GLP-1(7-36)amide was internalized. Incubation in the presence of increasing concentrations of non-labeled GLP-1(7-36)amide resulted in a decrease of internalization of 125I-labeled peptide indicating that this process is saturable. Incubation in the presence of 0.2 mM chloroquine, an inhibitor of intracellular hormone degradation, resulted in intracellular accumulation of 125I-GLP-1(7-36)amide. HPLC-supported analysis of cell content after internalization of 125I-GLP-1(7-36)amide during a 60-min incubation period at 37 degrees C revealed an elution profile showing two maxima of radioactivity: one represented intact labeled GLP-1(7-36)amide, the other an intracellular degradation product of the peptide. Chloroquine caused a 5-fold increase of the peak representing intact 125I-GLP-1(7-36)amide thus demonstrating inhibition of degradation of labelled peptide. Furthermore, a 4-fold increase of the other peak occurred possibly mirroring a delay of release of degradation products by chloroquine. It was excluded that chloroquine is able to interfere with GLP-1(7-36)amide-binding to its receptor.