Phase Of Glucose-induced Insulin Release Research Articles

Biophysical and Pharmacological Properties of Glucagon-Like Peptide-1 in Rats Under Isoflurane Anesthesia

Glucagon-like peptide-1 (GLP-1) increases insulin secretion and has an important role in maintaining glucose homeostasis. In this study, we evaluated the biophysical and pharmacological properties of GLP-1 by performing in vivo and in vitro experiments to determine the applicability of GLP-1 in glycemic control in rats under isoflurane anesthesia. Levels of portal GLP-1, insulin, and glucose and dipeptidyl peptidase-4 activity were measured in the basal fasting state and after gastric glucose load before, during, and after exposure to 30% O(2) in air (control) or 1.4% isoflurane in a mixture of 30% O(2) and air. The direct effects of isoflurane on GLP-1 secretion were assessed in human enteroendocrine NCI-H716 cells. Insulin release from isolated pancreatic islets was measured using a radioimmunoassay. Single pancreatic β-cell membrane potentials were recorded using whole-cell current-clamp patches perforated by β-escin. In fasting rats, inhalation of isoflurane led to a decrease in the basal levels of GLP-1 but did not affect insulin and glucose levels. Levels of GLP-1, insulin, and glucose increased after gastric administration of glucose in control rats. However, isoflurane attenuated the glucose-induced increase in GLP-1 and insulin levels and increased plasma glucose levels. In contrast, isoflurane did not affect dipeptidyl peptidase-4 activity before or after gastric glucose loading. Isoflurane (0.35 mM) inhibited GLP-1 release in NCI-H716 cells; this finding was similar to that observed in in vivo studies. In perifusion experiments, isoflurane (0.35 mM) inhibited glucose-induced insulin release, whereas exogenous GLP-1 (10 nM) enhanced insulin release. Importantly, combined administration of isoflurane and GLP-1 enhanced both phases of glucose-induced insulin release to an extent similar to that achieved with GLP-1 alone. Whole-cell patches showed that exposure to GLP-1 (10 nM) led to nearly complete restoration of glucose-stimulated depolarization that had been suppressed by isoflurane (0.35 mM). GLP-1 secretion is impaired during isoflurane anesthesia. However, our study showed that the insulinotropic action of GLP-1 was not affected by isoflurane. Furthermore, exposure to GLP-1 increased the membrane activity of pancreatic β-cells, preventing isoflurane-induced impairment of glucose-induced insulin secretion. These results support the hypothesis that GLP-1-based therapy may be a useful approach for achieving intraoperative glycemic control.

The effect of potassium, calcium and magnesium concentration on insulin and glucagon secretion of the perfused dog pancreas.

The effect of different potassium, calcium and magnesium concentrations in the perfusate on the hormone secretion of the isolated dog pancreas was investigated. A potassium concentration above 15 mMol/l shortly stimulates the insulin and glucagon secretion. Potassium ions (greater than or equal to 15 mMol/l) completely inhibit the early phase of glucose-induced insulin release. At a low Ca2+-level (0.25 mMol/l) the glucose-stimulated insulin secretion is reduced to basal values. On the other hand, the glucagon release is stimulated under these conditions. An increase of magnesium ions from 1.0 mMol/l to 2.5-7.5 mMol/l strikingly inhibits insulin and glucagon release by approximately 50%, which is compensated for insulin by increasing the Ca2+-content of the medium. Perfusates for normothermic pancreas perfusion should contain electrolyte concentrations within the physiological range.

The Cytokine Interleukin-1β Reduces the Docking and Fusion of Insulin Granules in Pancreatic β-Cells, Preferentially Decreasing the First Phase of Exocytosis

The prediabetic period in type I diabetes mellitus is characterized by the loss of first phase insulin release. This might be due to islet infiltration mediated by mononuclear cells and local release of cytokines, but the mechanisms involved are unknown. To determine the role of cytokines in insulin exocytosis, we have presently utilized total internal reflection fluorescence microscopy (TIRFM) to image and analyze the dynamic motion of single insulin secretory granules near the plasma membrane in live beta-cells exposed for 24 h to interleukin (IL)-1beta or interferon (IFN)-gamma. Immunohistochemistry observed via TIRFM showed that the number of docked insulin granules was decreased by 60% in beta-cells treated with IL-1beta, while it was not affected by exposure to IFN-gamma. This effect of IL-1beta was paralleled by a 50% reduction in the mRNA and the number of clusters of SNAP-25 in the plasma membrane. TIRF images of single insulin granule motion during a 15-min stimulation by 22 mm glucose in IL-1beta-treated beta-cells showed a marked reduction in the fusion events from previously docked granules during the first phase insulin release. Fusion from newcomers, however, was well preserved during the second phase of insulin release of IL-1beta-treated beta-cells. The present observations indicate that IL-1beta, but not IFN-gamma, has a preferential inhibitory effect on the first phase of glucose-induced insulin release, mostly via an action on previously docked granules. This suggests that beta-cell exposure to immune mediators during the course of insulitis might be responsible for the loss of first phase insulin release.

Impaired insulin secretion in isolated islets of Goto-Kakizaki rats, an animal model of non obese type 2 diabetes, is a primary event

The development of type 2 diabetes is associated with the impairment of insulin secretion. To evaluate the evolution of the secretory response, a chronological study comparing normal Wistar (W) vs Goto-Kakizaki (GK) rats, an animal model of non obese type 2 diabetes, was done. Glucose and arginine were tested in collagenase isolated islets of Langerhans with perfusion and ELISA immunoassay techniques. Fasting glycaemia and insulinemia and glucose tolerance were also evaluated. We have seen, in W rats, a mild glucose intolerance in the first two weeks of age. GK rats were always glucose intolerant with hyperglycaemia and hyperinsulinemia at fasten after one month old. Wistar islets had a characteristic biphasic response to glucose after the first two weeks of age. GK islets were always glucose irresponsive. Arginine induced an increase in insulin secretion in both animal models, independent of age, although GK rats had always a smaller response when compared to W rats. We concluded that 1) in W rats, a biphasic insulin secretion in response to glucose is observed after the first two weeks of age, simultaneously with glycaemia stabilization 2) in GK rats, both first and second phases of glucose-induced insulin release are significantly reduced and a smaller reduction in response to arginine is observed. This beta-cell disfunction is a primary event in this model of type 2 diabetes, preceding fasting hyperglycaemia and hyperinsulinemia.

Effects of S 21403 on hormone secretion from isolated rat pancreas at different glucose concentrations

We investigated the in vitro effects of therapeutical concentrations of S 21403 (a succinic acid derivative also known as KAD 1229 and mitiglinide) on insulin and glucagon secretion during a metabolic stimulus (glucose rising from 5 to 8.33 mM) or at a stable 2.22 mM glucose using the isolated perfused rat pancreas model, and we compared them with the patterns of repaglinide and glibenclamide. Control perfusions were also performed. During 8.33 mM glucose, insulin release peaked to 339.12±22.87 μU/ml in controls. S 21403 enhanced insulin release (first peak 413.02±14.90 μU/ml; P<0.03 vs. controls, P=ns vs. repaglinide, P<0.005 vs. glibenclamide). Repaglinide increased glucose-induced first peak secretion to 409.33±20.05 μU/ml within the eighth minute ( P<0.05 vs. controls, P<0.01 vs. glibenclamide). Glibenclamide did not affect the first phase of glucose-induced insulin release (peak of 338.41±29.79 μU/ml) but potentiated and delayed the second phase. No drug affected glucagon release. In conclusion, S 21403 induces a faster, more physiological pattern of insulin release than the other drugs we tested.

Contribution of neural intrapancreatic non-cholinergic non-adrenergic mechanisms to glucose-induced insulin release in the isolated rat pancreas.

In the isolated rat pancreas the effect of intrapancreatic non-adrenergic non-cholinergic nerves was examined upon insulin, glucagon and somatostatin release during perturbations of perfusate glucose. Elevation of glucose from 1.6 to 8.3 mmol/l increased insulin and somatostatin secretion and inhibited glucagon release. The first phase of insulin secretion was significantly reduced by the neurotoxin tetrodotoxin to 55% of the controls (p < 0.05). The somatostatin response was attenuated by tetrodotoxin while the change of glucagon remained unaffected. In contrast the combined adrenergic and cholinergic blockade with atropine, phentolamine and propranolol (10(-5) mol/l) did not modify the insulin, glucagon and somatostatin response. When glucose was changed from 8.3 to 1.6 mmol/l, the reduction of insulin and somatostatin release was not modified by tetrodotoxin, but stimulation of glucagon was significantly attenuated by 60-70% (p < 0.03), which was similar to the effect of combined adrenergic and cholinergic blockade. Subsequently, the effect of neural blockade was examined during more physiological perturbations of perfusate glucose levels. When glucose was changed from 3.9 to 7.2 mmol/l, tetrodotoxin also attenuated first phase insulin response by 40% while cholinergic and adrenergic blockade had no effect. The nitric oxide synthase inhibitor NG-Nitro-L-arginine-methyl-ester (L-NAME) did not alter the glucose-induced insulin response indicating that nitric oxide is not involved in this mechanism. It is concluded that neural non-adrenergic non-cholinergic mechanisms contribute to the first, but not second phase of glucose-induced insulin release. Non-adrenergic non-cholinergic effects do not participate in regulation of glucagon and somatostatin secretion under the conditions employed.(ABSTRACT TRUNCATED AT 250 WORDS)

Gradual Delay in Glucose-Induced First Phase Insulin Secretion by the Pancreatic Islets of 7 week-, 6 month-, and 1 year-old Rats

The temporal profiles of insulin secretion by isolated pancreatic islets of male Wistar rats with various ages up to 1-year-old were studied using high glucose, potassium (K+) depolarization, and arginine as secretagogues. In the islets of 6-month-old rats, the onset and peak of glucose-induced first phase of insulin release were delayed for 1 min compared to those of 7--week-old rats. The onset and peak were further delayed for 1 min in the islets of 1-year-old rats. The onset of glucose-induced second phase of insulin secretion, and the onset and peak of K+ depolarization- and arginine-induced insulin release were not delayed in the islets of 6-month and 1-year-old rats. Glucose-stimulated increase in cytosolic free calcium ([Ca2+]i) seemed not delayed in the islets of 1-year-old rats. We conclude that the first phase of glucose-induced insulin release by the islets is selectively delayed as rat ages. It was suggested that the defect lies distal to the elevation of [Ca2+]i.

Dose-response characteristics for glucose-stimulated insulin release in man and assessment of influence of glucose on arginine-stimulated insulin release

Glucose potentiates arginine-induced insulin release. We investigated the dose-response characteristics for both phases of glucose-induced insulin release in normal man, and studied the influence of hyperglycemia on arginine-induced insulin secretion. Dose-response curves of plasma C-peptide increments achieved during 60-minute hyperglycemia clamps (7, 11, 17, 24, and 32 mmol/L) with and without a primed continuous infusion of arginine (infusion rate, 15 mg/kg/min) were analyzed with a modified Michaelis-Menten equation. The ED 50 (half-maximally stimulating blood glucose concentration) of first-phase insulin release (determined from plasma C-peptide increments at 5 minutes) was significantly lower than the ED 50 for the second phase (60 minutes; 8.4 ± 0.8 v 14.3 ± 1.3 mmol/L, respectively, P < .002). Combined glucose-arginine stimulation significantly increased insulin release. Vmax of both phases of glucose-arginine-stimulated insulin release were positively correlated ( r = .75, P < .05). The ED 50 of the influence of glucose on first-phase arginine-induced insulin release was significantly lower than the ED 50 for the second phase (9.0 ± 1.1 v 12.7 ± 1.0 mmol/L, respectively, P < .02). For each insulin secretion phase separately, the ED 50 for the influence of hyperglycemia on arginine-induced insulin release were not significantly different from the ED 50 for glucose-induced insulin secretion (without arginine). When dose-response curves of plasma insulin increments were analyzed with the same equation, the ED 50 of second-phase glucose-induced plasma insulin increments was significantly higher than the ED 50 assessed from the plasma C-peptide increments (21.6 ± 2.8 v 14.3 ± 1.3 mmol/L, respectively, P < .05) We conclude (1) that the regulation by glucose of the two phases of insulin release involves (partly) different mechanisms in vivo, (2) that this difference in regulation occurs both in glucose- and in combined glucose-arginine-stimulated insulin release; (3) that combined glucose-arginine stimulation can be used as a tool for the assessment of B-cell glucose sensitivity; (4) that analysis using plasma insulin (instead of plasma C-peptide) increments can give a falsely high assessment for the parameter of the ED 50.

Insulin release in pancreatic islets by a glycolytic and a krebs cycle intermediate: Contrasting patterns of glyceraldehyde phosphate and succinate

Glyceraldehyde phosphate, a glycolytic intermediate, and succinic acid (as its methyl ester to make it permeable to the cell), a citric acid cycle intermediate, were the only glucose metabolites of many recently tested that stimulated insulin release. The effects of these two “new” insulin secretagogues on several pancreatic islet parameters were compared. Glyceraldehyde phosphate stimulated all of the insulin it released during the first 5 min after islets were exposed to it, and its maximum effect on calcium uptake was observed at 5 min. Monomethyl succinate stimulated insulin release mostly during the last 30 min of a 1-h incubation and its maximum effect on calcium uptake was at 60 min after it was applied to islets. Monomethyl succinate-induced insulin release, but not glyceraldehyde phosphate-induced insulin release, was inhibited by metabolic inhibitors (antimycin A, rotenone, cyanide, FCCP, fluoride, and iodoacetamide). This is consistent with the idea that monomethyl succinate is hydrolyzed to succinate which is metabolized intramitochondrially. The effects of glyceraldehyde suggest that glucose signals the first phase of insulin release by an agonist-like mechanism that originates in the cytosol and requires minimal energy. The effects of monomethyl succinate suggest that the signal for the second phase of glucose-induced insulin release originates in the mitochondrion and requires a large amount of energy.

Pertussis Toxin–Sensitive G Protein Mediation of PGE2 Inhibition of cAMP Metabolism and Phasic Glucose-Induced Insulin Secretion in HIT Cells

Although prostaglandin E2 (PGE2) is known to inhibit glucose-induced insulin secretion, it is uncertain whether PGE2 actions on the beta-cell are direct, whether they are equipotent for both phases of hormone secretion, and whether the same mechanism of action prevails throughout. Study of the HIT cell, a clonal line of pancreatic beta-cells, provides answers to these questions because perifusion with glucose and 3-isobutyl-1-methylxanthine stimulates biphasic insulin secretion. Perifusion with PGE2 decreased both the first and second phases of glucose-induced insulin release to 47 +/- 4% of controls. Pretreatment with pertussis toxin partly prevented PGE2 inhibition to 80 +/- 4% of controls for first phase and 79 +/- 4% of controls for second phase. To evaluate whether the partial prevention of PGE2 inhibition seen with pertussis toxin pretreatment was caused by Gi heterotrimer association between the preincubation period and the end of perifusion, PGE2 actions were also examined during continuous treatment with pertussis toxin. Under these conditions, PGE2 inhibition of both phases was totally prevented. However, no difference was observed in membrane protein ADP ribosylation when cells were examined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis after pretreatment or continuous treatment with pertussis toxin. Cyclic AMP (cAMP) accumulation was inhibited by PGE2 (from 3263 +/- 153 to 1549 +/- 158 fmol/10(6) cells) but less so after pretreatment with pertussis toxin (correlation between insulin release and cAMP accumulation during perifusion; n = 18, r = .85, P less than .001). Thus, PGE2 equally inhibits both phases of glucose-induced insulin secretion and cAMP generation through a pertussis toxin-sensitive G protein-mediated direct effect on the pancreatic beta-cell.

Pharmacological Characterization of the Voltage- Dependent Calcium Channel of Pancreatic B-Cell*

Pharmacological characteristics of the voltage-dependent calcium channel (VDCC) of the pancreatic B-cell were studied using omega-conotoxin (omega CgTX) and dihydropyridine (DHP) calcium channel blockers. High glucose and potassium (K+) depolarization were employed as the stimulant of insulin release. omega CgTX (greater than 50 nM), a blocker of neural, but not muscular, Ca2+ channels, partially blocked (27%) the second, but not the first, phase of glucose-induced insulin release without a significant effect on K+ depolarization-induced insulin release. The DHP Ca2+ channel blocker nifedipine inhibited both phases of glucose-induced insulin release (ED50 = 200 nM) and completely abolished both phases of response at 10 microM. In contrast, the DHP Ca2+ channel blocker only partially suppressed (75% at 10 microM) K+ depolarization-induced insulin release with an ED50 of 100 nM. We conclude that pancreatic B-cell possesses at least two classes of VDCCs; one is DHP sensitive, and the other DHP insensitive. Partial suppression of the second phase of glucose-induced insulin release by a high concentration of omega CgTX may be due to its toxic effect on the secretory machinery other than VDCC.

In vivo administration of interleudin-l inhibits glucose-stimulated insulin release

Recombinant interleukin-1β(IL-1β) was administered intraperitoneally for 3 days to normal C57BL/6ByJ (B6) mice. The islets from IL-1-treated and control animals were isolated and glucose-stimulated insulin secretion studied in the perifusion system. The total islet insulin content and the ultrastructure of the islets isolated from the animals treated with IL-1 did not differ from those seen in control animals. However, glucose-stimulated insulin release was significantly impaired after 3 days of in vivo administration of IL-1, either 3 μg/animal/day or 0.3 μg/animal/day. The administration of IL-1 inhibited an acute phase of glucose-induced insulin release, whereas neither basal insulin secretion nor insulin release from 10–30 min of perifusion with glucose was impaired. There was an only partial (27%) and non-significant restoration of the insulin secretory response to glucose stimulation 4 days after discontinuation of IL-1 treatment. We conclude that IL-1 administered in vivo is capable of adversely affecting pancreatic islet response to glucose stimulation. After 3 days of administration, these changes are confined to the process of insulin release, with the islet cell morphology and total insulin content being unaffected.

Isolation and characterization of porcine diazepam-binding inhibitor, a polypeptide not only of cerebral occurrence but also common in intestinal tissues and with effects on regulation of insulin release.

The polypeptide DBI (diazepam-binding inhibitor) has been purified from the porcine upper intestine, where it is abundant. Porcine mature DBI is composed of 86 amino acid residues and has a blocked N-terminus. The primary structure, the first DBI structure determined at the protein level, differs from those indirectly deduced for human and rat DBI at 11 and 17 positions, respectively. In total, the three mammalian DBIs differ at 22 positions but have exactly identical C-terminal 11-residue segments, highly charged and ending with C-terminal isoleucine. The porcine DBI inhibits both the early and the late phase of glucose-induced insulin release from the isolated perfused rat pancreas. Thus, the results identify by direct analysis the presence of DBI at a non-cerebral localization (gut), establish a novel structural form (porcine) and demonstrate a novel bioactivity (on insulin release). These aspects are of special interest in relation to the conserved segments, including the one at the C-terminal end, which may constitute functionally important parts of the polypeptide. It is possible that DBI belongs to a new family of gut polypeptides which inhibit glucose-mediated insulin release by hormonal and/or neurocrine mechanisms.

Biphasic Insulin Response to High Glucose and a Role of Protons and Calcium

In order to determine the role of protons and Ca++ in the biphasic insulin response to glucose, we studied the effect of monensin, a carboxylic ionophore, on the first phase and second phase of glucose-induced insulin release and Ca++ efflux from perifused rat pancreatic islets. The agent, 1-100 nM, dose dependently inhibited the insulin release from the islets incubated for 60 min in Krebs-Ringer bicarbonate buffer containing 16.7 mM glucose. Islet 14CO2 production rates from D-(U-14C)glucose were not affected by 10 or 100 nM monensin. Perifusion of the islets prelabeled with 45Ca++ demonstrated that 100 nM monensin had only a slight inhibitory effect on the first phase insulin response to 16.7 mM glucose and no effect on 45Ca++ efflux. This agent inhibited the second phase insulin release and depressed 45Ca++ efflux. When monensin was added at the start of the second phase release, the release was inhibited. When exposed to the agent before the stimulation by glucose, the first phase insulin release was observed, albeit significantly decreased, and the start of the insulin release and 45Ca++ efflux was delayed. The agent, added 30 min after the change to high glucose, immediately inhibited the insulin release. Thus, the first phase insulin release is mediated mainly through a mechanism which is not related to protons generated from glucose metabolism. Protons may be a crucial coupling factor in the second phase insulin release.

Effect of domperidone on the release of insulin after intravenous glucose load in man.

To determine whether the blockade of the dopaminergic system is capable of modifying glucose-induced insulin release in man, the responses of insulin to an iv glucose load were measured at various domperidone infusion rates. The infusion of 5 micrograms/kg/min of domperidone increased significantly plasma insulin levels during the acute phase of glucose-induced insulin release and lowered plasma glucose values at 50 and 60 min; the k of glucose disappearance improved significantly. At lower domperidone infusion rates the acute increment of insulin after glucose load was indistinguishable from the response observed at 5 micrograms/kg/min until 0.5 microgram/kg/min, while similar responses in control and experimental tests were observed at 0.25 microgram/kg/min. A group of subjects was submitted to an arginine load in order to establish whether the effect observed with domperidone was specific for the glucose-induced insulin release; but, this time, we did not observe any significant effect during the domperidone-induced dopaminergic blockade. Furthermore, we also measured the plasma prolactin levels, to see whether the specific and well known effect of domperidone on prolactin release matches with the effect on beta-cell function. As far as prolactin is concerned, we observed a dose response effect of domperidone infusion, with a detectable elevation of prolactin at infusion rate of 0.25 microgram/kg/min. Since domperidone is a specific antagonist of dopamine D2-receptors, we propose that dopamine might exert a specific inhibiting effect on glucose-induced insulin release through this class of dopamine receptors.

In vitro effects of ethanolamine on insulin secretion

The effects of ethanolamine on insulin secretion by the perfused rat pancreas were examined. During the second phase of glucose-induced insulin secretion 5-minute perfusions of ethanolamine at final concentrations of 0.1, 1 and 10 mM inhibited insulin release in a dose-related manner. When given throughout the experiment the highest dose of ethanolamine markedly suppressed both phases of glucose-induced insulin release. The inhibitory effect of ethanolamine was blunted in the presence of phentolamine. It is concluded that ethanolamine inhibits glucose-induced insulin secretion by the perfused rat pancreas and that α-adrenergic receptors play a role in its actions on insulin output.

Effects of monensin on glucose-induced insulin release and 45Ca2+ outflow.

The β-cells in the pancreatic islets respond to glucose with a biphasic insulin release2. Protons generated from glucose metabolism have been proposed to be a coupling factor between metabolic and ionic events in the cells1,7. A rise in cytosol Ca2+ concentration is considered to be essential in stimulus-secretion coupling in the β-cells4. Thus, it seems possible that the biphasic insulin release is a reflection of a biphasic change in the Ca2+ concentration of the cytosol, the change being controlled by the rate of generation of protons. To examine the role of protons and calcium in the insulin response to glucose, we examined the effect of monensin, a carboxylic ionophore8, on the first and second phase of glucose-induced insulin release and on the 45Ca2+ efflux from perifused rat pancreatic islets.KeywordsInsulin ReleaseBiphasic ChangePerifusion SystemInhibit Insulin ReleaseFractional Outflow RateThese keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.

Effects of lipoxygenase and cyclooxygenase inhibitors on glucose-stimulated insulin secretion from the isolated perfused rat pancreas

Some of the metabolites of arachidonic acid formed in the lipoxygenase and cyclooxygenase pathways stimulate insulin release. We studied the relative importance of each of these pathways in the modulation of glucose-induced insulin release by using inhibitors of arachidonate metabolism. Perfusion of the isolated rat pancreas with two chemically different inhibitors of cyclooxygenase, flurbiprofen and sodium salicylate, markedly inhibited prostaglandin E2 release, but had little effect on glucose-induced insulin release or on potentiation of insulin release caused by prior exposure to glucose. On the other hand, nordihydroguaiaretic acid (NDGA), a lipoxygenase inhibitor, not only inhibited both phases of glucose-induced insulin release but also abolished the potentiation effect. These effects of NDGA prevailed, when it was administered together with flurbiprofen, which caused profound inhibition of prostaglandin E2 release. We conclude that 1) lipoxygenase pathways play a dominant role in glucose-stimulated insulin release, and 2) endogenous lipoxygenase metabolites influence the potentiating effect of glucose on the release of insulin in response to a subsequent stimulation.

Multiple effects of growth hormone on insulin release from isolated pancreatic islets

The direct effects of growth hormone (GH) on the endocrine pancreas were studied in isolated islets of rats. To study GH-induced insulin release, islets were incubated in RPMI 1640 medium containing 1 or 10 μg/ml of bovine GH for 120 minutes. Islets incubated in the absence of GH served as controls. During the incubation, GH significantly increased the insulin concentration in the medium. To study the effect of GH on subsequent glucose-induced insulin release, islets were preincubated with GH; then the islets were transferred to perifusion chambers and after a 30-minute stabilization period with 2.8 mM glucose, the islets were stimulated by addition of 16.7 mM glucose for 60 minutes. These perifusions were performed in the absence of GH. Glucose-induced insulin release from control and GH-pretreated islets peaked at five minutes during the first phase (0–8 minutes) and plateaued at 25 minutes during the second phase (9–60 minutes). Preincubation with GH (1 μg/ml) did not change baseline or first phase release, but significantly suppressed the second phase of insulin release. When islets were preincubated with 10 μg/ml of GH, both phases of glucose-induced insulin release were suppressed; the total amount of insulin released by GH-pretreated islets was suppressed by 36.6% during the subsequent glucose stimulation period. These data indicate that GH stimulates insulin release by itself (GH-induced insulin release) but inhibits subsequent glucose-induced insulin release in a dose-dependent manner.

Potentiation of the insulin-releasing capacity of tolbutamide by thiols: Studies on the isolated perfused pancreas

It has been suggested that the islet thiol redox status plays a role in the regulation of beta-cell sensitivity in response to insulin secretagogues. Employing the isolated perfused rat pancreas, the effect of reduced glutathione (1 mM) and L-cysteine (5 mM) on insulin release induced by tolbutamide (0.2 mg/ml), glucose (5.6 and 11.1 mM) and tolbutamide (0.1 mg/ml) in the presence of 5.6 mM glucose was studied. In the absence of glucose or in the presence of 5.6 mM of glucose neither glutathione nor L-cysteine stimulated the release of insulin. Reduced glutathione potentiated the secretion induced by glucose (11.1 mM) during the first and the second phase. L-Cysteine potentiated only the first phase of glucose-induced insulin release, whereas the second phase was depressed. Both of the tested thiols potentiated the insulin secretory action of either tolbutamide (0.2 mg/ml) alone or tolbutamide (01. mg/ml) in the presence of glucose (5.6 mM). The data suggest that supplementation of thiols to the pancreatic beta-cells perse cannot initiate the insulin secretory process. It is also suggested that GSH and L-cysteine increase the sensitivity of beta-cells to the stimulatory action of tolbutamide and/or glucose.