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The proglucagon-derived peptide glucagon-like peptide-1 (GLP-1) is an intestinal signal peptide postprandially released from the L cells of the lower gut. Exogenously administered the synthetic hormone exerts a glucose-dependent insulinotropic effect at the pancreatic beta-cells and lowers plasma glucagon by an inhibitory effect against the alpha-cells. It delays gastric emptying by relaxation of the gastric fundus, inhibition of antral contractility, and stimulation of both the tonic and phasic motility of the pyloric sphincter. Enhancement of insulin, suppression of glucagon, and inhibition of gastric emptying are the main determinants controlling glucose homeostasis with GLP-1. Human studies employing the specific GLP-1 receptor antagonist exendin(9-39) show that endogenously released GLP-1 likewise controls fasting plasma glucagon, stimulates insulin, and influences all the motoric mechanisms known to control gastric emptying. Therefore, GLP-1 is discussed as an incretin hormone and as an enterogastrone in man. Synthetic GLP-1 also suppresses gastric acid and pancreatic enzyme secretion. The inhibitory effects on upper gastrointestinal functions are at least partly mediated by vagal-cholinergic inhibition and may involve interactions with vagal afferent pathways and/or circumventricular regions within the CNS. GLP-1 is a candidate humoral mediator of the 'ileal brake' exerting inhibition of upper gastrointestinal function preventing malabsorption and postprandial metabolic disturbances. As human studies indicate a central action of GLP-1 in reduction of food intake, it is uncertain if this is a consequence of induction of satiety or of transduction of visceral aversive stress signals.

The non-invasive detection of insulinomas remains a diagnostic problem that is not solved by means of somatostatin receptor scintigraphy. We investigated the biokinetics and specificity of uptake and degradation of the incretin hormone glucagon-like peptide-1 (GLP-1) in a rat insulinoma cell line (RINm5F) in order to ascertain whether radiolabelled GLP-1 may be suitable for specific visualisation of insulinomas in vivo. GLP-1 (7-36)amide was radioiodinated according to the iodogen method. The specificity of the uptake of [(125)I]GLP-1(7-36)amide by RINm5F cells was investigated. Degradation products of GLP-1 (7-36)amide in the cell medium were purified by HPLC. Their masses and amino acid sequences were determined by (252)Cf-plasma desorption mass spectrometry. Lysosomal degradation was inhibited and after differential centrifugation the amount of radiotracer incorporated into lysosomes was determined. Biodistribution studies were performed in a rat insulinoma model (NEDH rats and RINm5F cells) with [(123)I]GLP-1(7-36)amide and its more stable agonist [(123)I]exendin 3. The uptake of radiotracer into insulinoma cells reached a maximum within 5 min. It was inhibited by an excess of unlabelled peptide. [(125)I]GLP-1(7-36)amide accumulated in the cells if lysosomal degradation was inhibited. Degradation products of the peptide were found in the cell medium. We determined their mass and derived their amino acid sequence. Radiolabelling of exendin 3 was more difficult than that of GLP-1 because of the lack of tyrosine in its primary structure. Biodistribution studies showed rapid blood clearance and uptake of the radiotracer into the tumour and the pancreas. It was also possible to detect insulinomas in an animal model by external scintigraphy using radioiodinated GLP-1 (7-36)amide and exendin 3. GLP-1 (7-36)amide is specifically internalised into insulinoma cells by a receptor-mediated mechanism. Our results demonstrate that GLP-1 receptor-directed scintigraphy may be a new method for the detection of insulinomas in vivo. Due to the short half-life of GLP-1, its more stable analogue exendin 3 may better suit this purpose in vivo.

These in vitro studies aimed to characterize the pattern and the kinetics of endoproteolysis of the insulinotropic hormone glucagon-like peptide-1 (GLP-1) and related peptides by native ectopeptidases. Peptides were incubated with isolated rat or pig kidney brush-border microvilli membranes, which are a rich source of the ectopeptidases that are responsible for the post-secretory metabolism of peptide hormones. The proteolytic products were separated by reversed-phase HPLC column chromatography and characterised by molecular mass and primary structure. The relative importance of specific peptidases was established by measuring the effects of specific peptidase inhibitors on the kinetics of proteolysis. Dipeptidyl-peptidase-IV was found to be rate-limiting in the endoproteolysis of GLP-1. GLP-1 homologs, exendins-3 and -4, exhibited exceptional stability in the presence of isolated kidney microvilli membranes. Our finding that exendin-4 is several orders of magnitude more stable than GLP-1 and Ser-8-GLP-1 is especially noteworthy given this peptide's widely reported insulinotropic potency.

The present study was undertaken to establish in normal volunteers the alterations in beta-cell responsiveness to glucose associated with a constant infusion of glucagon-like peptide-1 (GLP-1) or a pretreatment infusion for 60 min. A high-dose graded glucose infusion protocol was used to explore the dose-response relationship between glucose and insulin secretion. Studies were performed in 10 normal volunteers, and insulin secretion rates (ISR) were calculated by deconvolution of peripheral C-peptide levels by use of a two-compartmental model that utilized mean kinetic parameters. During the saline study, from 5 to 15 mM glucose, the relationship between glucose and ISR was linear. Constant GLP-1 infusion (0.4 pmol x kg(-1) x min(-1)) shifted the dose-response curve to the left, with an increase in the slope of this curve from 5 to 9 mM glucose from 71.0 +/- 12.4 pmol x min(-1) x mM(-1) during the saline study to 241.7 +/- 36.6 pmol x min(-1) x mM(-1) during the constant GLP-1 infusion (P < 0.0001). GLP-1 consistently stimulated a >200% increase in ISR at each 1 mM glucose interval, maintaining plasma glucose at <10 mM (P < 0.0007). Pretreatment with GLP-1 for 60 min resulted in no significant priming of the beta-cell response to glucose (P = 0.2). Insulin clearance rates were similar in all three studies at corresponding insulin levels. These studies demonstrate that physiological levels of GLP-1 stimulate glucose-induced insulin secretion in a linear manner, with a consistent increase in ISR at each 1 mM glucose interval, and that they have no independent effect on insulin clearance and no priming effect on subsequent insulin secretory response to glucose.

Glucagon-like peptides (GLP) 1 and 2 are hormones derived from the post-translational processing of proglucagon in the intestinal L cells that influence intestinal motility and small bowel growth, respectively. We describe a patient with a neuroendocrine tumor of unknown primary origin with peritoneal carcinomatosis and diffuse liver metastases, who presented with constipation and nocturnal itching for over 3 years. Small bowel follow-through showed decreased small intestinal motility and marked intestinal hypertrophy. Biopsies from mesenterial lymph nodes showed, histologically, a well-differentiated neuroendocrine tumor (G1), with positive immunostaining for chromogranin A, GLP-1, GLP-2 and polypeptide YY (PYY). Jejunal biopsy demonstrated marked intestinal mucosal hypertrophy. HPLC analysis combined with RIA of tumor and serum extracts revealed that the tumor was producing and releasing fasting levels of GLP-1 of 738+/-20.7 pg/ml (normal levels (nl) <100 pg/ml), GLP-2 of 3,150+/-9 pg/ml (nl <100 pg/ml) as well as PYY 550 pg/ml (nl <100 pg/ml). Octreotide administration decreased levels of GLP-1 and GLP-2 and reduced small intestinal transit time from 150 to 50 min. However, tumor growth was not inhibited by octreotide, interferon or dacarbazine therapy and the patient died 8 months later. This is the first case report demonstrating the overproduction of GLP-1, GLP-2 and PYY from an neuroendocrine tumor, in a patient with intestinal hypertrophy and delayed intestinal transit time., (Copyright 2001 S. Karger AG, Basel)

The effects of the incretin hormone glucagon-like peptide 1 (7-36)amide (GLP-1) are mediated by the GLP-1 receptor (GLP-1R). This is expressed in a cell- and tissue-specific manner. Recently, we have cloned the 5'-flanking region of the human GLP-1R gene. The basal promoter activity is driven by the ubiquitous transcription factor Sp1. The tissue- and cell-specific expression of the gene requires several negatively acting cis-regulatory elements. We have now characterized one so far unknown distal cell-specific silencer element (DCS), repressing gene transcription of the human GLP-1R gene in fibroblasts and pancreatic A-cells, but not in pancreatic B- and D-cells. Our data suggests that the basal activity of the GLP-1R promoter is repressed in a tissue- and cell-specific manner by this novel silencer element.

Background: Glucagon-like peptide-1(7-36)amide (GLP-1) is a gut hormone released postprandially. Synthetic GLP-1 strongly inhibits gastric emptying in healthy subjects and in patients with diabetes mellitus., Aims: To investigate the effects of GLP-1 on antro-pyloro-duodenal motility in humans., Methods: Eleven healthy male volunteers were studied on two separate days. On the interdigestive study day, a basal period was followed by a 60 minute period of saline infusion and two further 60 minute periods of intravenous infusion of GLP-1 0.4 and 1.2 pmol/kg/min to achieve postprandial and supraphysiological plasma levels, respectively. On the postprandial study day, the same infusions were coadministered with intraduodenal lipid perfusion at 2.5 ml/min (2.5 kcal/min) followed by another 60 minutes of recording after cessation of GLP-1. Antro-pyloro-duodenal motility was measured by perfusion manometry., Results: GLP-1 significantly inhibited the number and amplitudes of antral and duodenal contractions in the interdigestive state and after administration of duodenal lipid. It abolished interdigestive antral wave propagation. In the interdigestive state, GLP-1 dose dependently increased pyloric tone and significantly stimulated isolated pyloric pressure waves (IPPW). Pyloric tone increased with duodenal lipid, and this was further enhanced by GLP-1. GLP-1 transiently restored the initial IPPW response to duodenal lipid which had declined with lipid perfusion. Plasma levels of pancreatic polypeptide were dose dependently diminished by GLP-1 with and without duodenal lipid., Conclusions: GLP-1 inhibited antro-duodenal contractility and stimulated the tonic and phasic motility of the pylorus. These effects probably mediate delayed gastric emptying. Inhibition of efferent vagal activity may be an important mechanism. As postprandial plasma levels of GLP-1 are sufficient to appreciably affect motility, we believe that endogenous GLP-1 is a physiological regulator of motor activity in the antro-pyloro-duodenal region.

The post-prandial release of glucagon-like peptide-1 (GLP-1) from the distal gut appears to involve a neural reflex that arises from the proximal gut. The neuropeptide calcitonin gene-related peptide (CGRP)'s potent stimulatory effect on GLP-1 release was characterized, using the isolated vascularly perfused rat ileum. CGRP, but not its homolog amylin, induced a dose-dependent and sustained release of GLP-1. This effect was greatly reduced in the presence of CGRP(8-37), was abolished by galanin, potentiated by luminal glucose and unaffected by atropine. GIP enhanced, but did not potentiate, this effect. The results reveal how CGRP is involved in the complex regulation of GLP-1 release.

Glucagon-like peptide 1 (7-36)amide (GLP-1) is an insulinotropic intestinal peptide hormone with a potential role as antidiabetogenic therapeutic agent. It mediates a potentiation of glucose-induced insulin secretion, by activation of adenylate cyclase and subsequent elevation of cytosolic free calcium, [Ca2+]cyt. We investigated the role of protein kinase A (PKA) in GLP-1 signal transduction, using isolated mouse islets as well as the differentiated beta-cell line INS-1. Two specific inhibitors of PKA, (Rp)-adenosine cyclic 3',5'-phosporothioate (Rp-cAMPS, up to 3 mM) and KT5720 (up to 10 microM), did not inhibit the GLP-1-induced [Ca2+]cyt elevation. Another PKA inhibitor, H-89, reduced the [Ca2+]cyt elevation only when applied at high concentrations (10-40 microM), higher than sufficient for PKA inhibition in many cell types. Furthermore, at these concentrations, H-89 also inhibited presumably PKA-independent processes such as glucose-induced [Ca2+]cyt elevations and intracellular calcium storage. This suggests a PKA-independent action of H-89. Similarly to H-89, the potent but unselective protein kinase inhibitor staurosporine inhibited the GLP-1-induced [Ca2+]cyt elevation only at high concentrations, at which it also inhibited glucose-induced [Ca2+]cyt elevations. The same observations as with GLP-1 were made when adenylate cyclase was stimulated with forskolin, for selective examination of signal transduction downstream of receptor and G protein. Our results suggest that the GLP-1-induced [Ca2+]cyt elevation is mediated independently of PKA and thus belongs to the yet-little-characterized ensemble of effects that are mediated by binding of cAMP to other target proteins.

Glucagon-like peptide-1-(7-36) amide (GLP-1) is an incretin hormone of the enteroinsular axis. Recent experimental evidence in animals and healthy subjects suggests that GLP-1 has a role in controlling appetite and energy intake in humans. We have therefore examined in a double-blind, placebo-controlled, crossover study in 12 patients with diabetes type 2 the effect of intravenously infused GLP-1 on appetite sensations and energy intake. On 2 days, either saline or GLP-1 (1.5 pmol. kg-1. min-1) was given throughout the experiment. Visual analog scales were used to assess appetite sensations; furthermore, food and fluid intake of a test meal were recorded, and blood was sampled for analysis of plasma glucose and hormone levels. GLP-1 infusion enhanced satiety and fullness compared with placebo (P = 0.028 for fullness and P = 0.026 for hunger feelings). Energy intake was reduced by 27% by GLP-1 (P = 0.034) compared with saline. The results demonstrate a marked effect of GLP-1 on appetite by showing enhanced satiety and reduced energy intake in patients with diabetes type 2.

Background/aims: Studies in animals suggest a physiological role for glucagon-like peptide-1-(7-36)-amide (GLP-1) in regulating satiety. The role of GLP-1 in regulating food intake in man has, however, not been investigated. Subjects-Sixteen healthy male subjects were examined in a double blind placebo controlled fashion., Methods: The effect of graded intravenous doses (0, 0.375, 0.75, and 1.5 pmol/kg/min) of synthetic human GLP-1 on food intake and feelings of hunger and satiety was tested in healthy volunteers., Results: Graded GLP-1 infusions resulted in a dose dependent reduction in food intake (maximal inhibition 35%, p<0.001 v control) and a similar reduction in calorie intake (32%; p<0.001). Fluid ingestion was also reduced by GLP-1 (18% reduction, p<0.01). No overt side effects were produced by GLP-1, but subjects experienced less hunger and early fullness in the period before a meal during GLP-1 infusion at the highest dose (p<0.05)., Conclusions: Intravenous infusions of GLP-1 decrease spontaneous food intake even at physiological plasma concentrations, implying an important role for GLP-1 in the regulation of the early satiety response in humans.

A pharmacological concentration of glucagon-like peptide-1 (GLP-1) in the insulin-deficient state clearly decreases the blood glucose level. Therefore, this study was designed to evaluate a putatively relevant effect of the gastrointestinal peptide as an adjuvant to insulin replacement therapy. GLP-1 (GLP-1(7-36) amide 10 pmol x kg(-1) x min(-1)) was infused intravenously over 8 hours in nine fasting, C-peptide-negative diabetic dogs. The animals were under normoglycemic control by glucose-controlled insulin infusion (GCII) during the night before and during GLP-1 administration. During the paired control tests, the animals received saline infusion instead of GLP-1. In addition to the insulin infusion rates required to maintain normoglycemia, hormones, metabolites, and the turnover rates for glucose (6-3H-glucose), alanine (U-14C-alanine), and urea (15N2-urea) were measured during the final 2 hours of GLP-1 administration. Circulating plasma GLP-1 levels increased from 3+/-1 to 17+/-7 pmol/L. There was no significant difference in the insulin infusion rate between the experimental and control groups (0.43+/-0.05 v. 0.40+/-0.05 mU x kg(-1) x h(-1), average over the entire interval). Glycemia was maintained at a practically identical level (4.9+/-0.3 v. 4.8+/-0.4 mmol/L). Also, the concentration of plasma insulin-which was not hyperinsulinemic--and pancreatic glucagon remained unaltered. We found no appreciable effect of GLP-1 on glucose production and metabolic clearance, alanine turnover and the formation of glucose from alanine (1.8+/-0.2 v. 1.4+/-0.2 micromol x kg(-1) x min(-1), or the urea production rate as a measure of overall amino acid catabolism (4.1+/-0.4 v. 4.1+/-0.4 micromol x kg(-1) x min(-1)). Thus, no conclusive adjuvant effect of GLP-1 was ascertained in insulin-treated diabetic dogs under normoglycemic control.

Background: This study addresses the question whether the insulinotropic gut hormone, glucagon-like peptide-1 (GLP-1), is released from the lower large bowel upon oral or rectal glucose uptake., Methods: It was evaluated whether rectum or sigmoid colon resection alters glucose homeostasis or the plasma levels of the insulinotropic gut hormone, gastric inhibitory polypeptide (GIP), or GLP-1. Six men and 3 women (age 63 +/- 8 years, BMI 25.4 +/- 4.0 kg/m2) with normal preoperative fasting glucose values were treated before and after resection of large bowel segments. Fasting oral glucose tolerance (OGT, 75 g glucose/300 ml) tests were performed both before and 10 days postoperatively. Another approach aimed to clarify whether luminal glucose stimulation in the rectum/sigmoid colon increases GLP-1 plasma levels. Ten healthy volunteers (4 males, 6 females, age 25 +/- 2 years, BMI 22.1 +/- 2.4 kg/m2) received enemas with both saline and, 7 days later, 1 g/kg body weight glucose (70% glucose solution) intrarectally., Results: Neither rectum nor sigmoid colon resection led to significant changes in the pre- and postoperative glucose responses to OGT testing, or insulin, GIP and GLP-1 release. Intrarectal glucose instillation increased blood glucose by about 10 mg/dl with parallel small elevations in immunoreactive insulin and immunoreactive C peptide. However, plasma GLP-1 levels remained unaltered., Conclusion: Our data make it unlikely that GLP-1 derived from the lower large bowel contributes significantly to maintain normal glucose tolerance.

Impaired glucose tolerance (IGT) and NIDDM are both associated with an impaired ability of the beta-cell to sense and respond to small changes in plasma glucose concentrations. The aim of this study was to establish if glucagon-like peptide 1 (GLP-1), a natural enteric peptide and potent insulin secretagogue, improves this defect. Two weight-matched groups, one with eight subjects having IGT (2-h glucose, 10.1 +/- 0.3 mmol/l) and another with seven subjects with diet-treated NIDDM (2-h glucose, 14.5 +/- 0.9 mmol/l), were studied on two occasions during a 12-h oscillatory glucose infusion, a sensitive test of the ability of the beta-cell to sense and respond to glucose. Glucose was infused with a mean rate of 4 mg x kg(-1) x min(-1), amplitude 33% above and below the mean rate, and periodicity of 144 min, with infusion of saline or GLP-1 at 0.4 pmol x kg(-1) x min(-1) for 12 h. Mean glucose levels were significantly lower in both groups during the GLP-1 infusion compared with during saline infusion: 9.2 +/- 0.4 vs. 6.4 +/- 0.1 mmol/l in the IGT subjects (P < 0.0004) and 14.6 +/- 1.0 vs. 9.3 +/- 0.7 mmol/l in NIDDM subjects (P < 0.0002). Despite this significant reduction in plasma glucose concentration, insulin secretion rates (ISRs) increased significantly in IGT subjects (513.3 +/- 77.6 vs. 583.1 +/- 100.7 pmol/min; P < 0.03), with a trend toward increasing in NIDDM subjects (561.7 +/- 122.16 vs. 642.8 +/- 128 pmol/min; P = 0.1). These results were compatible with enhanced insulin secretion in the presence of GLP-1. Spectral power was used as a measure of the ability of the beta-cell to secrete insulin in response to small changes in the plasma glucose concentration during the oscillatory infusion. Spectral power for ISR increased from 2.1 +/- 0.9 during saline infusion to 7.4 +/- 1.3 during GLP-1 infusion in IGT subjects (P < 0.004), but was unchanged in NIDDM subjects (1.0 +/- 0.4 to 1.5 +/- 0.6; P = 0.3). We concluded that low dosage GLP-1 improves the ability of the beta-cell to secrete insulin in both IGT and NIDDM subjects, but that the ability to sense and respond to subtle changes in plasma glucose is improved in IGT subjects, with only a variable response in NIDDM subjects. Beta-cell dysfunction was improved by GLP-1 infusion, suggesting that early GLP-1 therapy may preserve beta-cell function in subjects with IGT or mild NIDDM.

The gastrointestinal hormone, glucagon-like peptide-1(7-36)amide (GLP-1) is released after a meal. The potency of synthetic GLP-1 in stimulating insulin secretion and in inhibiting glucagon secretion indicates the putative physiological function of GLP-1. In vitro, the nonmammalian peptide, exendin(9-39)amide [ex(9-39)NH2], is a specific and competitive antagonist of GLP-1. This in vivo study examined the efficacy of ex(9-39)NH2 as an antagonist of exogenous GLP-1 and the physiological role of endogenous GLP-1. Six healthy volunteers underwent 10 experiments in random order. In each experiment, a 30-min period of euglycemia was followed by an intravenous infusion of glucose for 150 min that established a stable hyperglycemia of 8 mmol/liter. There was a concomitant intravenous infusion of one of the following: (1) saline, (2) GLP-1 (for 60 min at 0.3 pmol . kg-1 . min-1 that established physiological postprandial plasma levels, and for another 60 min at 0.9 pmol . kg-1 . min-1 to induce supraphysiological plasma levels), (3-5) ex(9-39)NH2 at 30, 60, or 300 pmol . kg-1 . min-1 + GLP-1, (6-8) ex(9-39)NH2 at 30, 60, or 300 pmol . kg-1 . min-1 + saline, (9 and 10) GIP (glucose-dependent insulinotropic peptide; for 60 min at 0.8 pmol . kg-1 . min-1, with saline or ex(9-39)NH2 at 300 pmol . kg-1 . min-1). Each volunteer received each of these concomitant infusions on separate days. ex(9-39)NH2 dose-dependently reduced the insulinotropic action of GLP-1 with the inhibitory effect declining with increasing doses of GLP-1. ex(9-39)NH2 at 300 pmol . kg-1 . min-1 blocked the insulinotropic effect of physiological doses of GLP-1 and completely antagonized the glucagonostatic effect at both doses of GLP-1. Given alone, this load of ex(9-39)NH2 increased plasma glucagon levels during euglycemia and hyperglycemia. It had no effect on plasma levels of insulin during euglycemia but decreased plasma insulin during hyperglycemia. ex(9-39)NH2 did not alter GIP-stimulated insulin secretion. These data indicate that in humans, ex(9-39)NH2 is a potent GLP-1 antagonist without any agonistic properties. The pancreatic A cell is under a tonic inhibitory control of GLP-1. At hyperglycemia, the B cell is under a tonic stimulatory control of GLP-1.

The incretin effect is reduced in NIDDM, although a corresponding attenuation of incretin hormone secretion does not occur. We characterized the direct interaction of GLP-I, an important incretin hormone, and leptin on insulin secretion and signal transduction in B-cells. Leptin inhibited GLP-I stimulated insulin release from the isolated perfused rat pancreas. Both phases of the biphasic insulin secretory response were inhibited. GLP-I receptor binding and GLP-I induced cAMP generation remained unchanged. Leptin reduced the GLP-I mediated increase of cytosolic Ca2+ concentration. It had similar effects on calcium elevations induced by forskolin. The effect was more pronounced during the plateau phase than during the initial peak. These effects could help to explain leptin's inhibitory effects on insulin secretion. The inhibition of GLP-I's insulinotropic effects by leptin may be an interesting aspect in the pathophysiology of NIDDM. The existence of an "adipo-insular axis" is suggested, in which leptin represents a negative feed-back signal from the adipose tissue to the endocrine pancreas.

Glucagon-like peptide-1(7-36)amide/(7-37) (GLP-1) is an incretin hormone which plays an important role in postprandial glucose homeostasis. Since GLP-1 potentiates glucose-induced insulin secretion, stimulates insulin biosynthesis and inhibits glucagon release, it is a potential tool for the treatment of diabetes mellitus. For this, an exact understanding of the structural/functional moieties of the peptide is mandatory. The present study investigates the importance of structural features of histidine7 at the N-terminus for GLP-1 action. Based upon binding and activity data obtained from ten different GLP-1 analogues we show that not the positive charge of the free alpha-amino group but the positive charge of the imidazole side chain of histidine is crucial for GLP-1 action. The presence of a ring structure and a basic function as well as the correct positioning of both seems to be decisive.

To establish potential effects of glucagon-like peptide I (GLP-I) on blood glucose control in insulin-deficient states, GLP-I [GLP-I(7-36) amide; 10 pmol x kg(-1) x min(-1)] was infused intravenously in six fasting, canine C-peptide-negative, chronically diabetic dogs for 8 h. Blood samples were saved for the analysis of hormones, metabolites, and turnover rates of glucose (6-(3)H-glucose), alanine (U-(14)C-alanine), and urea ((15)N(2)-urea) starting 22 h after the last subcutaneous dose of exogenous insulin. Circulating plasma GLP-I levels rose under infusion from 2.9 +/- 0.8 to 41.4 +/- 10.1 pmol/l. This was efficient to significantly reduce the preexisting diabetic hyperglucagonemia. Since in the utilized model functioning pancreatic beta-cells are lacking, GLP-I had no insulinogenic effect. Compared with control experiments in the same animals receiving saline infusion, glycemia dropped from 20.8 +/- 1.9 to 16.2 +/- 1.0 mmol/l (P < 0.05). This was in parallel to the infusion of GLP-I and was most likely caused by a decrease of elevated glucose production since overall glucose turnover decreased with no alteration in glucose metabolic clearance. Alanine turnover was significantly reduced, obviously reflecting a decline in alanine production in relation to changed muscle glucose uptake under conditions of lower glycemia and overall glucose turnover. There was, however, neither an effect of GLP-I on alanine conversion into circulating glucose nor an effect on urea production rate, indicating unchanged gluconeogenesis from amino acid precursors. We conclude that the blood glucose-lowering effect of GLP-I in an animal model of insulinopenia was shown to be due to a reduction in hepatic glucose output, possibly secondary to reduction in glucagon concentrations leading to decreased glycogenolysis. Whether GLP-I might be therapeutically useful in clinical insulin-deficient diabetes needs to be verified.

So far, a wealth of data originating from in vitro or animal experiments has been collected supporting the concept that the gut hormone, glucagon-like peptide-1 (GLP-1) may serve as a model molecule for the design of a new drug for the treatment of diabetes mellitus. This is supported by observations that GLP-1 has potent insulinotropic action in patients with non-insulin-dependent diabetes mellitus (NIDDM). It enhances beta-cell sensitivity to glucose stimulated insulin secretion. GLP-1 may also have a role in the treatment of impaired glucose tolerance, where the beta-cell is already insensitive to changes in plasma glucose concentrations. It may, as has previously been shown in animal models of 'prediabetes', delay the progressive decline in glucose tolerance to NIDDM. The glucose-dependent action of this peptide is an important feature in the treatment of NIDDM as it will protect against hypoglycaemic reactions, the most serious acute side-effect of antidiabetic therapy. Glucose utilization may be enhanced which would improve metabolic control in both NIDDM and IDDM. A glucagon lowering effect will further enhance metabolic control. This article reviews current experiences of the effects of GLP-1 in human studies. It points out the outcomes and limitations of previous trials and discusses future directions for the investigation of its potential use as a new agent in diabetes treatment.

Glucagon-like peptide-I (GLP-I) is a potent incretin hormone and mediates its actions via the cyclic AMP (cAMP) pathway. The GLP-I receptor belongs to the family of seven-transmembrane domain receptors coupled to G proteins. We have analyzed the regulation of GLP-I receptor function and expression by its own ligand and the cAMP-dependent pathway in rat insulinoma-derived beta cells (RINm5F). The GLP-I receptor underwent rapid homologous desensitization, which occurred at the receptor level. This was characterized by a reduced binding capacity not mediated by protein kinase A (PKA). GLP-I receptor mRNA levels were down-regulated during incubation of cells by agents increasing cAMP levels including GLP-I itself. This effect was dependent upon time and concentration. Forskolin, the PKA activator 5,6-dichloro-1-beta-D-ribofuranosyl-benzimidazole-3, 5-monophosphorothiotate, and GLP-I stabilized the GLP-I receptor mRNA. All induced down-regulation of the GLP-I receptor number within 3 h, a time point at which GLP-I receptor mRNA levels were not decreased. This effect was not influenced by cycloheximide. Therefore, in addition to transcriptional effects, posttranslational mechanisms exist to regulate GLP-I receptor numbers in insulin-secreting cells.

The insulinotropic glucagon-like peptide 1 (GLP-1) originates from the lower intestines. Surprisingly, food ingestion induces a rapid increase of GLP-1 plasma levels. Therefore, a complex regulation for postprandial GLP-1 secretion must exist, which cannot be solely explained by direct contact of nutrients in the gut lumen with the GLP-1-releasing L cells. This was addressed in the present study utilizing an isolated vascularly perfused rat ileum preparation. Cholinergic (methacholine) as well as peptidergic stimulation by glucose-dependent insulin-releasing polypeptide (synonym: gastric inhibitory polypeptide) (GIP) strongly enhanced GLP-1 secretion from the rat ileum. The stimulation of GLP-1 secretion by methacholine was abolished by addition of atropine and partly reduced by galanin. Galanin dose-dependently antagonized the stimulatory effect of GIP on GLP-1 release. Atropine was without effect. Furthermore, employing double immunohistochemistry labeling techniques galanin-immunoreactive nerves were detected in the vicinity of GLP-1-immunostained cells. Our data indicate that stimulatory and inhibitory mediators regulate GLP-1 secretion and that galanin is a likely inhibitor.

The interaction of glucagon-like peptide-I (GLP-I) and galanin in clonal endocrine pancreatic cells was characterized. By Northern blot analysis the presence of GLP-I receptor mRNA was shown in B (beta TC-1 cells) and D (RIN 1048-38) cells but not in A (INR1 G9) cells, thus confirming functional data demonstrating the absence of active GLP-I receptors on glucagon-producing cells. Galanin receptors were detected on B and D cells but not on A cells. In B and D cells galanin inhibited the GLP-I stimulated adenylate cyclase activity. Treatment of insulin- and somatostatin-producing cells with GLP-I increased intracellular cAMP levels, and this was dampened by galanin, GLP-I stimulated the activity of protein kinase A in B and D cells, which was also inhibited by galanin. Galanin alone did not influence B- and D-cell function. These data show that in the endocrine pancreas B and D cells but not A cells express GLP-I and galanin receptors. The interaction of GLP-I and galanin might act in the endocrine pancreas as a physiological inhibitor of the potent incretin hormone GLP-I. Therefore, we suggest galanin is a 'decretin'.

Food ingestion induces a rapid increase in the insulinotropic glucagon-like peptide-1 (GLP-1) in plasma. Paradoxically, GLP-1 originates from the lower intestines and therefore a complex regulation of postprandial GLP-1 secretion must exist. This was addressed in the present study by utilizing an isolated vascularly perfused rat ileum preparation. Peptides and neurotransmitters thought to be candidate mediators triggering GLP-1 secretion were arterially infused and GLP-1 was measured in the venous effluent. Arterial infusion of cholinergic agonists strongly enhanced GLP-1 secretion which was counteracted by the addition of atropine. Histamine, dopamine, 5-hydoxytryptamine, gamma-aminobutyric acid, and norepinephrine had no effect. Peptides of the bombesin family were strong stimulants whereas tachykinins, enkephalins, dynorphin, TRH, calcitonin-gene-related peptide and members of the secretin family, vasoactive intestinal peptide, peptide histidine isoleucine and neuropeptide Y, were less effective. The second incretin hormone, gastric inhibitory polypeptide (GIP), was the most potent stimulant of GLP-1 secretion in our study. It enhanced GLP-1 release up to sixfold above basal during the early phase followed by a sustained secretion at 400% above basal. This stimulation remained unaffected by atropine. In conclusion, in addition to luminal stimulation of nutrients, a cholinergic impulse as well as peptidergic mediators (among them possibly GIP and GRP) may have an impact on postprandial GLP-1 secretion from the rat ileum.

Glucagon-like peptide-I (GLP-I) is a potent incretin hormone that is now considered as a new therapeutic tool in the treatment of diabetes mellitus. In this study we characterized the effects of GLP-I on peptide hormone release from isolated human pancreatic islets. GLP-I stimulated insulin release in the presence of 10 mM glucose (2.8 mM glucose, 100%; 10 mM glucose, 166%; 10 mM glucose + 10 nM GLP-I, 222%) but had only a weak insulinotropic effect (128%) at 2.8 mM glucose. Glucagon release was inhibited by 10 mM glucose (2.8 mM glucose, 100%; 10 mM glucose, 72%) and by 10 nM GLP-I at 2.8 mM glucose (67%). Somatostatin secretion was increased by 10 mM glucose (2.8 mM glucose, 100%; 10 mM glucose, 166%). GLP-I stimulated somatostatin release in the presence of 2.8 mM glucose (172%). Pancreatic polypeptide (PP) secretion was enhanced by 10 mM glucose (2.8 mM glucose, 100%; 10 mM glucose, 236%). GLP-I induced PP release only in the presence of 2.8 mM glucose (184%).

The alpha-glucosidase inhibitor voglibose (AO-128) was designed to prevent rapid postprandial blood glucose rises in non-insulin-dependent diabetics. We analyzed its effect on the entero-insular axis in 72 healthy volunteers in a double-blind study design before, after the 1st dose, and on the 7th day of a 7-day treatment protocol (3 daily loads). Six parallel groups of 12 volunteers received voglibose (0.5, 1.0, 2.0, or 5.0 mg) or placebo (two groups). Blood was drawn at regular intervals up to 180 min after a standardized breakfast to analyze the levels of glucose, insulin, C peptide, gastric inhibitory polypeptide, and glucagon-like peptide 1 (GLP-1). As expected, after ingestion of voglibose, slight to moderate gastro-intestinal discomfort but no severe side-effects were reported. In a dose-dependent manner, voglibose significantly reduced postprandial increases of blood glucose, insulin, and C peptide. At the lower loads (0.5 and 1 mg voglibose three times daily), these effects were more pronounced after 7 days. The postprandial increase of gastric inhibitory polypeptide was already reduced after the first load of 2 and 5 mg voglibose. In comparison to the placebo group, this inhibition became also significant for the lower loads after 7 days. Interestingly, GLP-1, originating from the lower intestines, was increasingly released under voglibose treatment. The first administration of 1 mg voglibose enhanced GLP-1 secretion > 80% above controls. Treatment with 1 mg voglibose three times daily over 7 days revealed a maximal mobilizing effect on endogenous GLP-1 (> 90% above controls) which was not further increased by 2- or 5-mg loads. We conclude that voglibose treatment effectively inhibits intestinal disaccharidases and thereby mobilizes the endogenous pool of insulinotropic GLP-1.

The nutrient-dependent glucagon-like peptide-1 (7-36) amide (GLP-1) release was studied in comparison to the glucose-dependent insulin-releasing polypeptide (GIP) response in 10 healthy volunteers each undergoing various protocols. Plasma samples were saved up to 120 min after challenges by oral, intravenous or intraduodenal administration of nutrients. Basal plasma-GLP-1 concentrations ranged between 0.4 and 1.4 pM, maximal postprandial GLP-1 levels peaked between 10 and 12 pM. Intravenous glucose (25 g i.v.) did not change basal GLP-1 levels. Oral administration of glucose (50 g) induced a biphasic GLP-1 release peaking at 30-60 min and a biphasic GIP release peaking at 5 and 45 min. This increase paralleled the secretion of insulin. Oral galactose (100 g) and amino acids (25 g) also induced a rapid plasma GLP-1 response. After fat (67 g corn oil) a strong and long-lasting (> 120 min) increase of GLP-1 plasma levels occurred. When a mixed liquid meal was given (6 g soybean oil, 5 g casein, 13 g glucose) immunoreactive (IR)-GLP-1 rapidly increased and peaked after 5 min with declining levels after 30 min. In response to an intraduodenal infusion of a small glucose load (5.34 g within 120 min) a rapid, short-lasting GLP-1 response occurred whereas plasma GIP and insulin levels remained unaltered. Luminal perfusion of an isolated vascularly perfused rat ileum with a polydiet induced a rapid rise of portally released IR-GLP-1 which was followed by a sustained release. Glucose evoked sodium-dependently a sharp increase of IR-GLP-1 levels followed by a plateau release. The intraluminal infusion of a mixture of amino acids or fat was without any effect on IR-GLP-1. We hypothesize that in contrast to GIP the GLP-1 release from L cells is triggered by nervous reflexes, by putative humoral factor(s) being released from the upper small intestine in addition to nutrient stimuli acting at the luminal surface of the gut.

The preproglucagon gene encodes, in addition to glucagon, two smaller peptides with structural similarity: glucagon-like peptides 1 and 2. Glucagon-like peptide 1 (GLP-1) 7-36 amide is the most powerful incretin candidate. In the present study, GLP-1 immunoreactivity was investigated in tissue specimens of various types of gastroenteropancreatic tumors, and the serum-levels of GLP-1 were assayed. Immunohistochemical staining of 88 tumors revealed GLP-1 immunoreactivity in 17 neoplasias (19.3%), viz., in 7 out of 33 non-functioning tumors, 4 out of 20 gastrinomas, 4 out of 13 insulinomas, 1 out of 3 vasoactive-intestinal-polypeptide (VIP)omas and 1 adrenocorticotropic-hormone (ACTH)-producing tumor. In these tumors, GLP-1-immunoreactive cells were distributed either diffusely, arranged in clusters, or as single cells. All GLP-1-positive tumors were immunoreactive for glucagon or glicentin, 10 tumors were immunoreactive for pancreatic polypeptide, and 8 tumors for insulin. Ultrastructural analysis of 8 GLP-1-positive tumors, with the immunogold technique, demonstrated GLP-1 immunoreactivity mainly in cells resembling the A-cells of the pancreas or the L-cells of the gut. Of the 17 GLP-1-immunoreactive tumors, 15 were primarily located in the pancreas. Additionally, 2 non-functioning tumors of the rectum were GLP-1 immunoreactive. Five tumors were GLP-1 immunoreactive from 9 patients with multiple endocrine neoplasia I syndrome. Patients with GLP-1-immunoreactive tumors were characterized by a significantly lower rate of distant metastases (P < 0.01) and a higher rate of curative resections (P < 0.05).(ABSTRACT TRUNCATED AT 250 WORDS)

Glucagon-like peptide-I (GLP-I) is a potent incretin hormone and is considered as a new therapeutic tool in the treatment of diabetes mellitus. This study was designed to precisely characterize the binding behavior and activation of the recombinant GLP-I receptor against naturally occurring ligands of the glucagon/VIP/secretin peptide hormone family. CHO-cells were stably transfected with a plasmid containing a cDNA encoding for the rat GLP-I receptor. Northern blot analysis with this cDNA showed a single band of 2.7 kb in CHO cells, while in RINm5F cells, three bands of 2.7, 3.4, and 3.6 kb were specifically labelled. In receptor-binding studies 125I-GLP-I was displaced by GLP-I and weakly by PHI and oxyntomodulin but not by helodermin, helospectin I, helospectin II, secretin, VIP, and PACAP-38. Intracellular cAMP generation was stimulated by GLP-I, PHI, and oxyntomodulin. Helodermin, helospectin I, helospectin II, secretin, VIP, and PACAP-38 were not able to displace 125I-GLP-I from its receptor or to stimulate intracellular cAMP production. This data shows that the GLP-I receptor is characterized by a high ligand specificity.

Recent data revealed the existence of specific receptors for glucagon-like peptide-1(7-36)amide (GLP-1) on rat lung membranes. Utilizing slide-mount autoradiography of fresh frozen lung tissue sections, we have localized binding sites for GLP-1 on mucous glands in the trachea and on vascular smooth muscle of the pulmonary artery. When tracheas were incubated in a modified Ussing chamber, the addition of GLP-1 to the submucosal side increased 35S-sulfate-labeled macromolecule secretion (191 +/- 12% above basal, P < 0.005). The optimal secretory response elicited by GLP-1 was approximately 23% of the maximal secretory response after a maximal acetylcholine stimulation. Other proglucagon-derived peptides such as glucagon, oxyntomodulin, and GLP-2 had no effect. In isolated rings of arteries, GLP-1 (10(-8) to 10(-5) M) induced a dose-dependent and time-reversible relaxation of preconstricted arteries. In a preparation with denuded epithelium, GLP-1 lost its effect. In conclusion, GLP-1 might represent another neuropeptide that acts as neurotransmitter of the peptidergic, nonadrenergic-noncholinergic nervous system that innervates the airways.

Normal transcription and postranslational processing of the preprotachykinin (PPT)-I gene and regulated release of substance P and neurokinin A by the rat pancreatic endocrine cell-line, RINm5F, has been demonstrated, using radioimmunoassays (RIAs), reversed-phase (rp)HPLC and Northern blot analysis. This is the first stable cell-line found to express the PPT-I gene and provides an opportunity for investigating PPT-I gene expression and tachykinin biosynthesis. RIN5mF cells are a model for the pancreatic beta-cell, which is not known to exhibit PPT-I gene expression which may, therefore, be a feature of the transformed state of these cells. These data may imply that the tachykinins are important in pancreatic islet embryogenesis.

Glucagon-like peptide-1 (GLP-1) is the main product of the intestinal processing of proglucagon. It is released from the intestinal K-cells into the circulation in response to the oral ingestion of food. At the pancreatic beta cell GLP-1 is a potent insulin secretagogue in the presence of elevated glucose levels, defining glucagon-like peptide-1 as a new incretin. Its action is mediated by specific receptors coupled to the adenylate cyclase system by a stimulatory G-protein. Finally, glucagon-like peptide-1 stimulates proinsulin gene expression and it is thus involved at several levels in the regulation of insulin synthesis and secretion.

A highly specific monoclonal antibody directed against the C-terminal part of glucagon-like peptide-1 (GLP-1) was raised to immunohistochemically evaluate the distribution of GLP-1 containing cells in the entire gastrointestinal tract including pancreas of rat, pig and man. In the pancreas GLP-1-immunoreactive cells were found variously shaped and predominantly located in the periphery of the islets. Ultrastructurally, GLP-1 was co-localized with glucagon in the alpha-granula of A-cells and was mainly restricted to the electrondense core. In the intestine open type cells reaching the lumen via a slender apical process were stained with the GLP-1 antibody. They occurred in all parts of the crypts but predominantly in the basal portion. The density of GLP-1 immunoreactive cells varied between species in a characteristic order: rat greater than pig greater than man. In pig and human gut a large number of cells occurred in the distal jejunum and ileum. A continuous increase of cell densities was found from the proximal to the distal colon resulting in highest numbers in the rectum. In rats the highest cell density occurred in the ileum. Again, a continuous increase of GLP-1-positive cell numbers was evident from the proximal to the distal portion of small and large bowel. GLP-1 was partly co-localized with PYY. The GLP-1 positive cells appeared electronmicroscopically as L-cells with the typical large granula. This morphological data indicates that GLP-1-releasing cells in the small intestine are appropriately positioned in the distal part to sense and respond to the presence of nutrients that have escaped the absorptive surface of the upper small intestine.