Glucagon-like Peptide-1 Immunoreactivity Research Articles

1753-P: Islet GLP-1R Protein Expression Is Reduced in Humans with Cystic Fibrosis–Related Diabetes

Impaired insulin secretion underlies the development of cystic fibrosis-related diabetes (CFRD). Glucagon like peptide-1 (GLP-1) is produced in islet α cells, and potentiates insulin secretion by binding GLP-1 receptors (GLP-1Rs) on β cells. In CFRD, α-cell mass is increased. Despite this, we hypothesized that levels of GLP-1 and/or its signaling components are reduced in CFRD, thereby contributing to deficits in insulin secretion. To address our hypothesis, paraffin-embedded pancreas sections from humans with CFRD and age-matched nondiabetic controls (from nPOD) underwent immunohistochemistry to quantify protein levels of active GLP-1 and GLP-1R (n=5-6/group). Spatial transcriptomics (NanoString) was also performed to assess differentially expressed α- and/or β-cell genes between CFRD and control pancreas (n=3/group; 15 CFRD + 9 control islets). GLP-1 immunoreactivity was unchanged between CFRD and control pancreas (0.33±0.15 vs 0.51±0.15, % GLP-1 area/pancreas area; p=0.4), however islet GLP-1R immunoreactivity was significantly reduced (4.35±1.21 vs 18.70±0.50, % GLP-1R area/islet area; p=0.004). Surprisingly, few α- or β-cell genes were differentially expressed between CFRD and control islets. However, a gene set test (ROAST) to assess transcripts involved in GLP-1 production/signaling revealed that those in α cells (GCG, PCSK1, PCSK2, DPP-4) significantly changed in mixed directions (up- or down-regulated; p=0.001). No such perturbation was seen in the β-cell gene set (INS, GLP1R, DPP-4, PLCG1, PIP4P2, ITPR3, ADCY1, ADCY6, PRKACA, RAPGEF4; p=0.4). In sum, our data suggest CFRD is not associated with altered intra-islet GLP-1 levels, however α-cell genes that modulate GLP-1 levels are dysregulated. Also, islet GLP-1R protein levels are reduced, though expression of genes downstream of GLP-1R is unchanged. Thus, interventions to restore protein levels of islet GLP-1R and/or promote intracellular signaling beyond GLP-1R may improve β-cell function in CFRD. Disclosure R. Vemireddy: None. J.J. Castillo: None. B.S. Fountaine: None. T.K. Bammler: None. J. MacDonald: None. R.L. Hull-Meichle: Research Support; Casma Therapeutics, Cystic Fibrosis Foundation, NIH - National Institutes of Health. Employee; Veterans Administration. Research Support; Veterans Administration. S. Gharib: None. S. Zraika: None. Funding National Institutes of Health (P30DK089507, P30DK017047)

Investigating Intestinal Glucagon After Roux-en-Y Gastric Bypass Surgery.

After Roux-en-Y gastric bypass (RYGB) surgery, postprandial plasma glucagon concentrations have been reported to increase. This occurs despite concomitant improved glucose tolerance and increased circulating plasma concentrations of insulin and the glucagon-inhibiting hormone glucagon-like peptide 1 (GLP-1). To investigate whether RYGB-induced hyperglucagonemia may be derived from the gut. Substudy of a prospective cross-sectional study at a university hospital in Copenhagen, Denmark. Morbidly obese individuals undergoing RYGB (n = 8) with or without type 2 diabetes. Three months before and after RYGB, participants underwent upper enteroscopy with retrieval of gastrointestinal mucosal biopsy specimens. Mixed-meal tests were performed 1 week and 3 months before and after RYGB. The 29-amino acid glucagon concentrations in plasma and in mucosal gastrointestinal biopsy specimens were assessed using mass spectrometry-validated immunoassays, and a new monoclonal antibody reacting with immunoreactive glucagon was used for immunohistochemistry. Postprandial plasma concentrations of glucagon after RYGB were increased. Expression of the glucagon gene in the small intestine increased after surgery. Glucagon was identified in the small-intestine biopsy specimens obtained after, but not before, RYGB. Immunohistochemically, mucosal biopsy specimens from the small intestine harbored cells costained for GLP-1 and immunoreactive glucagon. Increased concentrations of glucagon were observed in small-intestine biopsy specimens and postprandially in plasma after RYGB. The small intestine harbored cells immunohistochemically costaining for GLP-1 and glucagon-like immunoreactivity after RYGB. Glucagon derived from small-intestine enteroendocrine l cells may contribute to postprandial plasma concentrations of glucagon after RYGB.

GLP-1 Localisation and Proglucagon Gene Expression in Healthy and Diabetic Mouse Ileum.

IntroductionGlucagon-like peptide-1 (GLP-1) is a polypeptide that is mainly produced by intestinal L cells and is encoded by the proglucagon gene. In this study, GLP-1 localisation was investigated in the ileum of healthy and diabetic mice by immunohistochemistry and proglucagon gene expression was assayed by reverse transcription-polymerase chain reaction.Material and MethodsThis study included 18 male Balb/c mice that were divided into diabetic, sham, and control groups. Mice in the diabetic group received 100 mg/kg of streptozotocin. Immunohistochemical expression of GLP-1 was determined using the avidin–biotin–peroxidase complex technique, and proglucagon gene expression was determined by RT-PCR.ResultsAnalysis of GLP-1 immunohistochemical localisation showed that GLP-1-immunopositive cells (L cells) were present between epithelial cells in the intestinal crypts. The intensity and localisation of GLP-1 immunoreactivity were similar among the mice in all the groups. Proglucagon gene expression levels were also statistically similar among the mice in all the groups.ConclusionNo difference was demonstrated among the mice in the diabetic, sham, or control groups with respect to proglucagon gene expression and GLP-1 localisation in the ileum, suggesting that diabetes does not affect proglucagon gene expression in the ileum.

Anti–glucagon-like peptide-1 immunoreactivity in samples of blood and ileum obtained from neonatal and adult alpacas

To compare numbers of L cells in intestinal samples and blood concentrations of glucagon-like peptide (GLP)-1 between neonatal and mature alpacas. Intestinal samples from carcasses of 4 suckling crias and 4 postweaning alpacas for immunohistochemical analysis and blood samples from 32 suckling crias and 19 healthy adult alpacas for an ELISA. Immunohistochemical staining was conducted in accordance with Oregon State University Veterinary Diagnostic Laboratory standard procedures with a rabbit polyclonal anti-GLP-1 primary antibody. Stained cells with staining results in ileal tissue were counted in 20 fields by 2 investigators, and the mean value was calculated. For quantification of GLP-1 concentrations, blood samples were collected into tubes containing a dipeptidyl peptidase-4 inhibitor. Plasma samples were tested in duplicate with a commercial GLP-1 ELISA validated for use in alpacas. Counts of stained cells (mean ± SD, 50 ± 18 cells) and plasma GLP-1 concentrations (median, 0.086 ng/mL; interquartile range, 0.061 to 0.144 ng/mL) were higher for suckling alpacas than for postsuckling alpacas (stained cells, 26 ± 4 cells; plasma GLP-1 concentration, median, 0.034 ng/mL; interquartile range, 0.015 to 0.048 ng/mL). Older alpacas had lower numbers of L cells in intestinal tissues and lower blood concentrations of GLP-1 than those in neonates. These findings suggested that there may be a decrease in the contribution of GLP-1 to insulin production in adult alpacas, compared with the contribution in neonates.

Swim training restores glucagon-like peptide-1 insulinotropic action in pancreatic islets from monosodium glutamate-obese rats

Glucagon-like peptide-1 (GLP-1) is an important modulator of insulin secretion by endocrine pancreas. In the present study, we investigated the effect of swim training on GLP-1 insulinotropic action in pancreatic islets from monosodium glutamate (MSG)-obese rats. Obesity was induced by neonatal MSG administration. MSG-obese and control (CON) exercised rats swam for 30 min (3 times week(-1) ) for 10 weeks. Pancreatic islets were isolated by colagenase technique and incubated with low (5.6 mM) or high (16.7 mM) glucose concentrations in the presence or absence of GLP-1 (10 nM). In addition, GLP-1 gene expression in ileum was quantified in fasting and glucose conditions. Exercise reduced obesity and hyperinsulinemia in MSG-obese rats. Swim training also inhibited glucose-induced insulin secretion in islets from both groups. Islets from MSG-obese rats maintained GLP-1 insulinotropic response in low glucose concentration. In contrast, in the presence of high glucose concentration, GLP-1 insulinotropic action was absent in islets from MSG-obese rats. Islets from MSG-exercised rats showed reduced GLP-1 insulinotropic action in the presence of low glucose. However, in high glucose concentration swim training restored GLP-1 insulinotropic response in islets from MSG-obese rats. In all groups, glucose intake increased GLP-1 immunoreactivity and gene expression in ileum cells in relation to fasting conditions. Swim training reduced these parameters only in ileum cells from CON-exercised rats. Neither MSG treatment nor exercise affected GLP-1 expression in the ileum. Exercise avoids insulin hypersecretion restoring GLP-1's insulinotropic action in pancreatic islets from MSG-obese rats.

Glucagon-like peptide-1 of brainstem origin activates dorsomedial hypothalamic neurons in satiated rats

A high number of neurons express c-fos in response to unlimited food intake in fasted rats in the ventral subdivision of the hypothalamic dorsomedial nucleus (DMHv). We report here, that in same conditions, limited food consumption failed to induce Fos expression in DMHv neurons suggesting that satiation should be one of the important signals that activate these neurons. The possible origin of fibers conducting satiation signals to the DMHv could be in the lower brainstem, especially glucagon-like peptide-1 (GLP-1)-containing neurons in the nucleus of the solitary tract (NTS). We demonstrate that GLP-1-immunoreactive fibers and fiber terminals topographically overlap with activated Fos-positive neurons in the DMHv in refed rats. Using immunocytochemistry and in situ hybridization histochemistry, we demonstrated GLP-1 receptors in Fos-expressing neurons of the DMH. Unilateral transections of ascending GLP-1-containing fibers from the NTS inside the pons in refed rats (unlimited food consumption) resulted in a dramatic decrease in the density of GLP-1 fibers and in the number of Fos-immunoreactive neurons in the DMHv, but only on the side of the transection. Contralateral to the transection, neither the GLP-1 fiber density nor the number of Fos-positive cells changed significantly. Meanwhile, the density of GLP-1 immunoreactivity was markedly accumulated in transected nerve fibers caudal to the cuts, as a consequence of the interruption of the ascending GLP-1 transport route. These findings suggest that the solitary-hypothalamic projections may represent the neuronal route through GLP-1 neurons of the NTS activate DMHv neurons via GLP-1 receptors by conveying information on satiety.

Glucocorticoid regulation of preproglucagon transcription and RNA stability during stress

Stress elicits a synchronized response of the endocrine, sympathetic, and central nervous systems to preserve homeostasis and well-being. Glucagon-like peptide-1 (GLP-1), a primary posttranslational product of the preproglucagon (PPG) gene, activates both physical and psychological stress responses. The current study examined mechanisms regulating expression of PPG gene products in the hindbrain. Our results indicate that PPG mRNA decreases rapidly after exposure to acute stressors of multiple modalities. Reduced mRNA levels are accompanied by reduced GLP-1 immunoreactivity in the paraventricular nucleus of hypothalamus, suggesting release at PPG terminals. Stress-induced decrements in PPG mRNA were attenuated in adrenalectomized-corticosterone-replaced rats, suggesting that mRNA down-regulation is due at least in part to glucocorticoid secretion. In contrast, acute stress increased levels of PPG heteronuclear RNA (hnRNA) in a glucocorticoid-dependent manner, suggesting that decreases in PPG mRNA are due to increased degradation rather than reduced transcription. Glucocorticoid administration to unstressed rats is sufficient to cause decrements in PPG mRNA and increments in PPG hnRNA. These findings suggest that glucocorticoids deplete the pool of transcribed PPG mRNA and concurrently stimulate PPG gene transcription, with the latter allowing a mechanism for replenishment of PPG mRNA after stress cessation. The combination of rapid PPG mRNA depletion and initiation of PPG transcription within 30 min is consistent with a rapid action of glucocorticoids on GLP-1 bioavailability, resulting in a transient reduction in the capacity for neuropeptidergic excitation of stress responses.

Elevated basal and post-feed glucagon-like peptide 1 (GLP-1) concentrations in the neonatal period

Glucagon-like peptide 1 (GLP-1) is an incretin hormone that stimulates glucose-induced insulin secretion, increases beta-cell proliferation, neogenesis and beta-cell mass. In adults, plasma concentrations of amidated GLP-1 are typically within the 5-10 pmol/l range in the fasting state and increases to approximately 50 pmol/l after ingestion of a mixed meal. We measured plasma glucose, insulin and amidated forms of GLP-1 prefeed and then at 20 and 60 min post-feed following ingestion of a 60-70 ml of standard milk feed in preterm (n=10, 34-37 weeks) and term newborn infants (n=12, 37-42 weeks). Reverse-phase fast protein liquid chromatography was used to characterise the molecular nature of the circulating GLP-1. Mean birth weight was 3.18 kg and mean age at sampling for GLP-1 was 7.7 days. The mean basal GLP-1 concentration was 79.1 pmol/l, which increased to 156.6 pmol/l (+/-70.9, P<0.001) and 121.5 pmol/l (+/-59.2) at 20 and 60 min respectively. Reverse-phase chromatography analysis suggested that the majority of GLP-1 immunoreactivity (>75%) represented GLP-1 (7-36) amide and (9-36) amide. Basal and post-feed amidated GLP-1 concentrations in neonates are grossly raised with the major fractions of circulating GLP-1 being (7-36) amide and (9-36) amide. Elevated GLP-1 concentrations in the newborn period may have a role in regulating maturation of enteroendocrine system and also of increasing pancreatic beta-cell mass and regeneration. The high levels of GLP-1 may be due to immaturity of the dipeptidyl peptidase IV and or lower glomerular filtration rate in the neonatal period. Further studies are required to understand the role of GLP-1 in the neonatal period.

Distribution of glucagon-like peptide-1 immunoreactivity in the hypothalamic paraventricular and supraoptic nuclei

Glucagon-like peptide-1 (GLP-1) plays a role in modulating neuroendocrine and autonomic function. The hypothalamic paraventricular nucleus (PVN) contains aggregations of GLP-1 fibers and expresses GLP-1 receptors, making it a likely site of action for GLP-1 signaling. The current study was designed to establish domains of GLP-1 action, focusing on axosomatic appositions on different neuroendocrine and autonomic cell populations in the PVN. The data indicate abundant GLP-1-immunoreactive terminal appositions on corticotropin-releasing hormone neurons in the medial parvocellular PVN. GLP-1 positive boutons can also be observed in apposition to oxytocinergic neurons and on retrogradely labeled pre-autonomic neurons projecting to the region of the nucleus of the solitary tract. In contrast, there were very few vasopressinergic neurons with GLP-1 appositions. Overall, the data indicate that the central GLP-1 system preferentially targets neurons in hypophysiotrophic zones of the PVN, consistent with excitatory actions of GLP-1 on adrenocorticotropin release. GLP-1 is also in position to influence oxytocin secretion and control outflow to brainstem cardiovascular relays.

Expression of glucagon-like peptide-1 in the taste buds of rat circumvallate papillae

Glucagon-like peptide-1 (GLP-1), a 30-amino-acid peptide hormone, is a typical peptide of the brain-gut axis and can affect the metabolism of various tissues and organs. GLP-1 is secreted by intestinal L cells in response to nutrient ingestion. Some studies have shown that taste signaling elements were co-expressed in enteroendocrine cells of the small intestine, and in particular by L-cells. The present study was performed to explore the protein and mRNA expression of GLP-1 in the taste buds of rat circumvallate papillae and to try to determine the significance of its secretion. GLP-1 immunoreactivity was observed in spindle-shaped taste bud cells, with positive cells displaying a characteristic distribution of reaction product that was confined to the cytosol. Reverse transcription polymerase chain reaction (RT-PCR) assay showed that GLP-1 mRNA was expressed in circumvallate papillae. The expression of GLP-1 suggests that it may play an important role in the taste stimulation of nutrients and gut hormone secretion.

Glucagon-like peptide-1 (GLP-1) immunoreactivity in the forebrain and pituitary of the teleost, Clarias batrachus (Linn.)

Glucagon-like peptide-1-immunoreactivity (GLP-ir) was seen in several olfactory receptor neurons in the olfactory epithelium and in the olfactory nerve fascicles that extend caudally to the bulb and innervate the glomerular layer. Thick GLP-ir fascicles were seen in the medial olfactory tracts (MOT); these probably represent the extrabulbar olfactory projections. GLP-ir fibers in the MOT penetrate into the telencephalon, swing dorsocaudally into the area ventralis telencephali/pars supracommissuralis (Vs) just above the anterior commissure. The immunoreactive fascicles continue caudally in the dorsal hypothalamus and are traceable as far as the lateral recesses. Scattered GLP-ir fibers were seen in the ventral telencephalon and preoptic area. While several immunoreactive cells were seen in the pars intermedia of the pituitary gland, few were encountered in the rostral pars distalis and proximal pars distalis. Role for the peptide in the processing of olfactory information and in the regulation of pituitary gland may be suggested.

Comparative Study on the Distribution of Glucagon-like Peptide-1 (GLP-1)-immunoreactive Cells in the Intestine of Chicken and Ostrich.

The distribution of glucagon-like peptide-1 (GLP-1)-immunoreactive cells was investigated in the intestine of the chicken and ostrich. GLP-1-immunoreactive endocrine cells were distributed only in the jejunum and ileum of both avian species. In chicken, these cells had a pyramidal or spindle-like shape with a cytoplasmic process and were observed at the lower part of villi and in intestinal crypts. In ostriches, endocrine cells immunoreactive with GLP-1 antiserum showed a flask- or spindle-like shape with a longer cytoplasmic process and were found in the middle and lower parts of intestinal villi. The cytoplasmic process reached to the intestinal lumen in both avian species. The frequencies of occurrence of GLP-1-immunoreactive cells in the jejunum and ileum were 19.71±8.78 and 35.82±13.87, respectively (cell numbers per mucosal area : cells/mm2, mean±SD), in the chicken, and 7.72±5.50 and 6.72±3.24 in the ostrich. The difference in the frequency of occurrence of cells showing GLP-1 immunoreactivity was significant between jejunum and ileum in the chicken, but not in the ostrich. These findings suggest that GLP-1 might be one of the major hormones in the avian intestine. There is, however, a species difference in the distributional pattern of cells containing GLP-1.

Elevated plasma glucagon-like peptide 1 and 2 concentrations in ileum resected short bowel patients with a preserved colon

BACKGROUNDThe glucagon-like peptides (GLP) 1 and 2 are secreted postprandially from L cells located mainly in the ileum. Both hormones prolong intestinal transit and GLP-2 is intestinotrophic in rodents. Patients...

Regulation of pancreatic PC1 and PC2 associated with increased glucagon-like peptide 1 in diabetic rats.

The pancreatic processing enzymes, PC1 and PC2, convert proinsulin to insulin and convert proglucagon to glucagon and glucagon-like peptide 1 (GLP-1). We examined the effect of streptozotocin (STZ) treatment on the regulation of these enzymes and the production of insulin, glucagon, and GLP-1 in the rat. Pancreatic PC1 and PC2 mRNA increased >2-fold and >4-fold, respectively, in rats receiving intraperitoneal STZ (50 mg/kg) daily for 5 days. Immunocytochemistry revealed that, although pancreatic islet cells in the STZ-treated rats were sparse and atrophic PC1, PC2, glucagon, and GLP-1 immunoreactivity increased dramatically in the remaining islet cells. Heightened PC1 and PC2 expression was seen in cells expressing glucagon but not in insulin-expressing cells. Furthermore, in STZ-treated rats, bioactive GLP-1(7-36 amide) accumulated in pancreatic extracts and serum 3- and 2.5-fold, respectively, over control animals. This treatment also caused a 2-fold increase in the ratio of amidated forms of GLP-1 immunoreactivity to total glucagon immunoreactivity in the pancreas but did not affect the ratio of proinsulin to insulin. We conclude that hyperglycemic rats have an increased expression of prohormone converting enzymes in islet alpha cells, leading to an increase in amidated GLP-1, which can then exert an insulinotropic effect on the remaining beta cells.

Identification of amidated forms of GLP-1 in rat tissues using a highly sensitive radioimmunoassay

The development of a sensitive radioimmunoassay (RIA) for C-terminally amidated forms of glucagon-like peptide-1 (GLP-1) is described. Rabbits immunized with GLP-1(7–36)amide conjugated to bovine serum albumin with glutaraldehyde produced antisera containing high-affinity antibodies directed against an epitope that included the free amidated C-terminus of the peptide. These antisera could be used in a sensitive RIA (detection limit 0.1 fmol/tube) that measured GLP-1(7–36)amide and GLP-1(1–36)amide equally. Total concentrations of amidated GLP-1 immunoreactivity in extracts of rat hypothalamus, pancreas and intestine were determined by RIA, and resolved into GLP-1(7–36)amide, GLP-1(1–36)amide and unidentified cross-reacting substances by HPLC. Whereas only GLP-1(7–36)amide could be identified in the hypothalamus, in amounts that represented 55–94% of total glucagon-like immunoreactivity (GLI), the pancreas produced chiefly GLP-1(1–36)amide, representing 0.8–3.4% of total GLI, and only trace or undetectable amounts of GLP-1(7–36)amide (0–0.36% of total GLI). This argues against any role of intrapancreatic GLP-1(7–36)amide in the secretion of insulin. In the terminal ileum total amidated GLP-1 immunoreactivity represented 27–73% of total GLI, and in five of six specimens only GLP-1(7–36)amide could be identified on HPLC, in amounts representing 13–17% of total GLI. Only one specimen of terminal ileum contained HPLC-identified GLP-1(1–36)amide (13% of total GLI) in addition to GLP-1(7–36)amide (31% of total GLI). Acid–ethanol extraction of peptide-free rat plasma with added GLP-1(7–36)amide gave recoveries of 91±SEM 2% in the range 20–200 pmol/l. Basal plasma amidated GLP-1 in six unanaesthetized rats was 4.1±1.1 pmol/l and rose to a maximum of 15.4±3.0 pmol/l 10 min after intragastric glucose 1 g/kg, illustrating the modest level of plasma responses of amidated forms of GLP-1.

Reciprocal cellular distribution of glucagon-like peptide-1 (GLP-1) immunoreactivity and GLP-1 receptor mRNA in pancreatic islets of rat.

The respective cellular distribution of glucagon-like peptide-1 (GLP-1) immunoreactivity and mRNA expression of the GLP-1 receptor was compared in rat pancreas by means of immunohistochemistry and in situ hybridization. GLP-1 immunoreactivity was present in the marginal zone of rat pancreatic islets. In contrast, GLP-1 receptor mRNA signals were confined to the central part of pancreatic islets. Neither GLP-1 immunoreactivity nor GLP-1 receptor mRNA signals were detected in the exocrine pancreatic acinar cells or duct cells. The differential distribution of GLP-1 immunoreactivity and GLP-1 receptor mRNA signals indicates that the GLP-1 amino acid sequence is present in the alpha-cell zone of pancreatic islets, whereas the GLP-1 receptor is expressed mainly by beta cells. Thus, our data by in situ hybridization demonstrates the significant expression of GLP-1 receptors on beta cells but makes a significant expression on alpha cells rather unlikely.

Luminal glucagon-like peptide-1(7-36) amide-releasing factors in the isolated vascularly perfused rat colon.

Glucagon-like peptide-1 (GLP-1) is released from endocrine cells of the distal part of the gut after ingestion of a meal. GLP-1 secretion is, in part, under the control of hormonal and/or neural mechanisms. However, stimulation of the colonic L cells may also occur directly by the luminal contents. This was examined in the present study, using an isolated vascularly perfused rat colon. GLP-1 immunoreactivity was measured in the portal effluent after luminal infusion of a variety of compounds which are found in colonic contents (nutrients, fibers, bile acids, short-chain fatty acids (SCFAs)). Oleic acid (100 mM) or a mixture of amino acids (total concentration 250 mM), or starch (0.5%, w/v) did not increase GLP-1 secretion over basal value. A pharmacological concentration of glucose (250 mM) elicited a marked release of GLP-1 which was maximal at the end of infusion (400% of basal), while 5 mM glucose was without effect on secretion. Pectin evoked a dose-dependent release of GLP-1 over the range 0.1-0.5% (w/v) with a maximal response at 360% of basal when 0.5% pectin was infused. Cellulose or gum arabic (0.5%) did not modify GLP-1 secretion. The SCFAs acetate, propionate or butyrate (5, 20 and 100 mM) did not induce a significant release of GLP-1. Among the four bile acids tested, namely taurocholate, cholate, deoxycholate and hyodeoxycholate, the last one was the most potent at eliciting a GLP-1 response with a maximal release at 300% and 400% of the basal value when 2 and 20 mM bile acid were administered respectively.(ABSTRACT TRUNCATED AT 250 WORDS)

Regulation of glucagon-like peptide-1-(7-36) amide secretion by intestinal neurotransmitters and hormones in the isolated vascularly perfused rat colon

Glucagon-like peptide-1 (GLP-1) is promptly released from endocrine cells of the distal part of the gut after oral ingestion of a meal. To test the possibility that hormones produced by the proximal small intestine or transmitters of the enteric nervous system may be involved in the early phase of meal-induced GLP-1 secretion, various intestinal regulatory peptides and neurotransmitters of the gut were administered intraarterially in the isolated vascularly perfused rat colon preparation. The release of GLP-1 in the portal effluent was measured by a specific RIA. Intraarterial infusion of glucose-dependent insulinotropic peptide (GIP) over the concentration range 0.25-1 nM evoked a dose-dependent release of GLP-1, with a maximal response of 350% of the basal value. Tetrodotoxin did not modify the GIP-induced release of GLP-1. Secretin or cholecystokinin did not stimulate the secretion of GLP-1. Bombesin (10(-9)-10(-7) M) provoked a dose-dependent release of GLP-1, consisting of an early peak, followed by a sustained response. Calcitonin gene-related peptide (5 x 10(-8) M) induced a dramatic rise of GLP-1 immunoreactivity in the portal effluent (peak at 800% of the basal value 10 min after the start of infusion). Similarly, the beta-adrenergic agonist isoproterenol at concentrations of 10(-7) and 10(-6) M provoked a pronounced release of GLP-1 (peak at 500% of the basal value with 10(-6) M isoproterenol). Finally, the muscarinic cholinergic agonist bethanechol at a concentration of 10(-4) M evoked a gradual increase in GLP-1 immunoreactivity, which reached a maximal value (900% over basal) at the end of the 30-min infusion period. The lowest concentration of bethanechol used in the present study (10(-5) M) did not increase portal GLP-1 immunoreactivity over the basal value. Tetrodotoxin did not modify the bethanechol-, isoproterenol-, calcitonin gene-related peptide-, or bombesin-induced GLP-1 release. In conclusion, the present study conducted with the isolated vascularly perfused rat colon shows that there are interactions between the two most potent incretins, GIP and GLP-1, probably through an enteroendocrine pathway. Additionally, several transmitters of the gut are potent stimulants of GLP-1 release and, therefore, represent potential tools in the treatment of the noninsulin-dependent diabetes mellitus.

Regulation of glucagon-like peptide-1-(7-36) amide secretion by intestinal neurotransmitters and hormones in the isolated vascularly perfused rat colon.

Glucagon-like peptide-1 (GLP-1) is promptly released from endocrine cells of the distal part of the gut after oral ingestion of a meal. To test the possibility that hormones produced by the proximal small intestine or transmitters of the enteric nervous system may be involved in the early phase of meal-induced GLP-1 secretion, various intestinal regulatory peptides and neurotransmitters of the gut were administered intraarterially in the isolated vascularly perfused rat colon preparation. The release of GLP-1 in the portal effluent was measured by a specific RIA. Intraarterial infusion of glucose-dependent insulinotropic peptide (GIP) over the concentration range 0.25-1 nM evoked a dose-dependent release of GLP-1, with a maximal response of 350% of the basal value. Tetrodotoxin did not modify the GIP-induced release of GLP-1. Secretin or cholecystokinin did not stimulate the secretion of GLP-1. Bombesin (10(-9)-10(-7) M) provoked a dose-dependent release of GLP-1, consisting of an early peak, followed by a sustained response. Calcitonin gene-related peptide (5 x 10(-8) M) induced a dramatic rise of GLP-1 immunoreactivity in the portal effluent (peak at 800% of the basal value 10 min after the start of infusion). Similarly, the beta-adrenergic agonist isoproterenol at concentrations of 10(-7) and 10(-6) M provoked a pronounced release of GLP-1 (peak at 500% of the basal value with 10(-6) M isoproterenol). Finally, the muscarinic cholinergic agonist bethanechol at a concentration of 10(-4) M evoked a gradual increase in GLP-1 immunoreactivity, which reached a maximal value (900% over basal) at the end of the 30-min infusion period. The lowest concentration of bethanechol used in the present study (10(-5) M) did not increase portal GLP-1 immunoreactivity over the basal value. Tetrodotoxin did not modify the bethanechol-, isoproterenol-, calcitonin gene-related peptide-, or bombesin-induced GLP-1 release. In conclusion, the present study conducted with the isolated vascularly perfused rat colon shows that there are interactions between the two most potent incretins, GIP and GLP-1, probably through an enteroendocrine pathway. Additionally, several transmitters of the gut are potent stimulants of GLP-1 release and, therefore, represent potential tools in the treatment of the noninsulin-dependent diabetes mellitus.

Glucagon-like peptide 1 immunoreactivity in gastroentero-pancreatic endocrine tumors: a light- and electron-microscopic study

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)