Multiple effects of leucine on glucagon, insulin, and somatostatin secretion from the perfused rat pancreas

The effects of increasing concentrations of leucine (0.2, 2.0, and 15.0 mmol/liter) on glucagon secretion from the perfused rat pancreas were examined at various glucose levels (0, 3.3, or 8.3 mmol/liter) and in the absence or presence of either arginine (5.0 mmol/liter) or glutamine (10.0 mmol/liter). At a low glucose concentration (3.3 mmol/liter), leucine caused a dose-related biphasic increase in glucagon output in the absence of arginine, but only a transient increase in the presence of the latter amino acid. These positive responses were markedly reduced and, on occasion, abolished at a high glucose concentration (8.3 mmol/liter). Moreover, at a low glucose concentration (3.3 mmol/liter) and in the presence of arginine, the highest concentration of leucine (15.0 mmol/liter) provoked a sustained and reversible inhibition of glucagon release. Likewise, leucine (15.0 mmol/liter) reversibly inhibited glucagon secretion evoked by glutamine in the absence of glucose. Thus, leucine exerted a dual effect on the secretion of glucagon, the inhibitory effect of leucine prevailing at a high concentration of the branched chain amino acid and when glucagon secretion was already stimulated by arginine or glutamine. At a physiological concentration (0.2 mmol/liter), however, leucine was a positive stimulus for glucagon release, especially in the absence of another amino acid. Concomitantly, leucine was always a positive stimulus for both insulin and somatostatin secretion. The intimate mechanisms involved in the dual effect of leucine on glucagon secretion remain to be elucidated.     

Inhibition of glucagon and insulin secretion from the perfused rat pancreas by a B-cell-selective somatostatin analog

The isolated perfused rat pancreas was used to study the effects of somatostatin and the analog des-Asn⁵-[D-Trp⁸, D-Ser¹³]-somatostatin on arginine-stimulated insulin and glucagon secretion. Even though the analog was found to inhibit glucagon secretion at concentrations of 10 and 100 ng/ml, it had a relatively more inhibitory effect on the B cell than on the A cell than did somatostatin itself. These data suggest that the A- and B-cell receptors for these two peptides differ.     

Effect of ventromedial hypothalamic lesions on the secretion of somatostatin,insulin, and glucagon by the perfused rat pancreas

Somatostatin, insulin, and glucagon secretion by the perfused pancreas were studied in adult female rats 10 days after ventromedial hypothalamic (VMH) lesions and in sham operated controls to assess the role of their hypothalamic control. Insulin secretion was significantly greater in VMH-lesioned rats both under basal conditions and after stimulation by theophylline and arginine plus theophylline. Basal glucagon secretion was greater in VMH-lesioned rats as was the glucagon response to theophylline alone and in combination with arginine. Basal somatostatin secretion was similar in VMH and control rats but somatostatin secretion induced by theophylline and by arginine plus theophylline was significantly increased in VMH-lesioned rats. Both the pancreatic content and concentration of somatostatin were increased in VMH-lesioned rats. These results indicate the presence of hyperresponsiveness of A, B, and D cells following VMH destruction and provide new evidence for a role of the hypothalamus in the regulation of pancreatic somatostatin secretion.     

Difference in calcium dependency of insulin,glucagon and somatostatin secretion in response to glibenclamide in perfused rat pancreas

Summary The extracellular calcium requirements for insulin, glucagon and somatostatin release induced by 1 g/ml of glibenclamide have been compared in the perfused, isolated rat pancreas. In the absence of glucose, the drug evoked insulin release equally well at physiological (2.6 mmol/l) and low (0.25 mmol/l) levels of total calcium. In contrast, glibenclamide evoked somatostatin release at 2.6 but not at 0.25 mmol/l of calcium. At 2.6 mmol/l of calcium, glibenclamide evoked bimodal effects (stimulation followed by inhibition) on glucagon secretion. At 0.25 mmol/l of calcium, basal secretory rates of glucagon were elevated and a small stimulatory effect of glibenclamide was seen. Addition of 0.5 mmol/l of EGTA to media with low calcium concentrations uniformly abolished the A, B and D cell secretory responses to glibenclamide. The possible modulation of calcium dependency by a non-stimulatory concentration of glucose was tested by its addition at 3.3 mmol/l to the perfusion media. Glucose enhanced glibenclamide-induced insulin secretion, both at 0.25 and 2.6 mmol/l of calcium. However, at 0.25 mmol/l of calcium, the enhancing effect of glucose was more pronounced than at 2.6 mmol/l. At 2.6 mmol/l of calcium, glucose diminished the somatostatin and abolished the glucagon response to glibenclamide. At 0.25 mmol/l of calcium, glucose did not influence somatostatin release while the presence of the sugar diminished basal and glibenclamide-induced glucagon secretion. The present data confirm the requirement of extracellular calcium for A, B and D cell secretion, demonstrating different calcium dependencies for the cell types and indicate that this dependency can, in part, be modulated by glucose.     

Differential sensitivity to somatostatin of pancreatic polypeptide, glucagon and insulin secretion from the isolated perfused canine pancreas

This dose-response study deals with the relative inhibitory effect of somatostatin on the acetylcholine-stimulated release of pancreatic polypeptide (PP), glucagon, and insulin from the isolated canine pancrease. Somatostatin in picomolar doses potently inhibited insulin and glucagon secretion, whereas PP secretion was relatively insensitive. Also, in the absence of acetylcholine, somatostatin exerted a preferential inhibition of the release insulin and glucagon compared with PP. These findings point to a physiologically important role of somatostatin for the secretion of insulin and glucagon, but probably not for PP.     

Different effects of hypothermia on insulin and glucagon secretion from the isolated perfused rat pancreas

Summary Two series of experiments with the isolated perfused rat pancreas were performed in parallel. The conditions differed only with respect to temperature, which was 37.5 °C in one series and 28 °C in the other. The lowering of the temperature decreased insulin secretion induced by glucose as well as the insulin response to tolbutamide and acetylcholine. Unlike insulin, glucagon secretion was not significantly modified by hypothermia. Our results suggest that the mechanisms involved in glucagon and insulin secretion are different.Equipe de Recherche Associée au CNRS n 786     

Effects of exogenous insulin, glucagon, and somatostatin on islet hormone secretion in the perfused chicken pancreas

The effects of exogenous insulin were examined in the isolated perfused chicken pancreas with the duodenum excluded. At low background glucose (50 mg/dl), exogenous insulin infused at a concentration of 20,000 microU/ml elicited clear stimulation of somatostatin secretion while simultaneously inhibiting glucagon release. When the background glucose concentration was elevated to 750 mg/dl, exogenous insulin, had no effect on either somatostatin or glucagon release. When graded doses of exogenous insulin were infused into the chicken pancreas at low background glucose, low concentrations (200 microU/ml) had little effect on somatostatin or glucagon release, but higher concentrations (2000 and 20,000 microU/ml) had clear effects on both somatostatin and glucagon secretion. Glucagon infused at 100 ng/ml stimulated both insulin and somatostatin release. When somatostatin was infused at 25 ng/ml, clear inhibition of glucagon was seen with insulin inhibited to a lesser extent. This study supports the notion of a negative feedback relation between B and D-cells of the pancreatic islets and suggests a paracrine mediation.     

Effect of insulin on glucose- and arginine-stimulated somatostatin secretion from the isolated perfused rat pancreas

The effects of exogenous insulin on somatostatin secretion from the isolated perfused rat pancreas have been investigated in the presence of 5.6 mM glucose and when somatostatin secretion was stimulated by either glucose (16.7 mM) or arginine (20 mM). Insulin (15 mU/ml) significantly and rapidly suppressed glucose- and arginine-stimulated somatostatin release. However, at 5.6 mM glucose and, in the absence of other stimulators of somatostatin release, insulin had no effect on the somatostatin secretion rate.     

Stimulation by alpha-adrenergic mechanisms of the secretion of growth hormone-releasing factor (GRF) from perifused rat hypothalamus

The possible involvement of adrenergic mechanisms in regulating the secretion of growth hormone (GH)-releasing factor (GRF) from the rat hypothalamus was examined in vitro with a perifusion system. A high potassium concentration (56 mM) stimulated GRF release from the hypothalamus. The infusion of clonidine (10(-4) M), an alpha 2-adrenergic stimulant, resulted in an increase in the spontaneous release of GRF. In the presence of propranolol (10(-5) M), a beta-adrenergic blocking agent, clonidine (10(-5) and 10(-4) M) stimulated GRF release more prominently in a dose-related manner, whereas propranolol (10(-5) and 10(-4) M) by itself did not affect the spontaneous GRF release. The stimulatory effect of clonidine (10(-4) M) on GRF release in the presence of propranolol was inhibited by yohimbine (10(-4) M), an alpha 2-adrenergic blocking agent. These findings suggest that alpha 2-adrenergic mechanisms play a role in stimulating GRF release from the hypothalamus in rats.     

Pancreatic polypeptide,glucagon and insulin secretion from the isolated perfused canine pancreas

Summary The release of pancreatic polypeptide (PP) by gut hormones, acetyl choline and adrenaline was investigated in an isolated perfused pancreas preparation. PP was potently released by 1 nmol/l caerulein (186±12%, p<0.001) and gastric inhibitory peptide (GIP) (211±31%, p<0.005) as well as by 1 [mol/l acetyl choline (1097±59%, p<0.001). A significant two-fold release of PP was also evoked by 1 nmol/l vasoactive intestinal peptide (VIP) (129±38%, p<0.02 and gastrin (108±25% p<0.01). Insulin release, induced by high glucose concentration was enhanced by both GIP (210 ±38%, p<(0.01) and VIP (48±5%, p<0.001). In addition GIP enhanced the release of glucagon by 179±18% (p<0.001) at 1.4 mmol/l glucose and by 127±24% (p<0.005) at 8.3 mmol/l glucose. Thus no simple inter-relationship appears to exist between the control of the three circulating islet hormones.     

The effects of growth hormone-releasing factor (GRF) and dopamine on growth hormone (GH) secretion in acromegaly

GRF (1-40), dopamine (DA), DA plus GRF and placebo were administered to 6 acromegalic patients. The GRF administration induced a highly variable GH release (GH delta % 167.3 +/- 21.4; mean +/- SE). GRF did not provoke any change in PRL serum levels. During simultaneous GRF and DA administration GH release was found to be reduced (GH delta % 80.2 +/- 17.8) compared to that observed for a corresponding period of time after GRF alone (p less than 0.05). Our data underscore that in acromegaly the DA tonus inhibits GH secretion after GRF by acting directly at the pituitary level.     

The interaction of Vasoactive Intestinal Polypeptide (VIP), glucose and arginine on the secretion of insulin,glucagon and somatostatin in the perfused rat pancreas

Summary Vasoactive Intestinal Polypeptide (VIP) increased the release of insulin, glucagon and somatostatin from the perfused rat pancreas. The amount of these hormones released was dependent upon the prevailing glucose concentration. VIP stimulated glucagon release in the absence of glucose, while insulin and somatostatin release were increased by VIP only in the presence of glucose concentrations of 4.4 mmol/l and above. Glucagon secretion stimulated by arginine in the presence of 4.4 mmol/l glucose was potentiated by VIP. In contrast, VIP did not induce any further increase in the secretion of insulin and somatostatin over that stimulated by arginine. At higher concentrations of glucose (6.7, 16.7, and 33.3 mmol/l) VIP continued to stimulate insulin and somatostatin release, this effect being synergistic on early-phase insulin release. The effects of VIP on islet cells thus depend on the levels of modulating nutrients.     

Kisspeptin-13 inhibits insulin secretion without affecting glucagon or somatostatin release: study in the perfused rat pancreas

Kisspeptins are a family of peptides encoded by the KISS1 gene, which binds to G-protein-coupled receptor (GPR54), an orphan GPR54 related to galanin receptors. Endogenous forms composed of 54, 14, and 13 amino acids have been identified. Kisspeptin and GPR54 mRNAs have been detected in pancreatic B and A cells. Furthermore, kisspeptin-54 has been shown to slightly stimulate the last phase of glucose-induced insulin secretion in mouse and human islets and to inhibit insulin release in MIN6 cells. We have investigated the effect of kisspeptin-13 on insulin, glucagon, and somatostatin secretion. The study was performed in the perfused rat pancreas. Glucose, arginine, carbachol, and exendin-4 were used as secretagogues. Hormones were measured by RIA. Kisspeptin-13 reduced glucose-induced insulin secretion in a dose-dependent manner (IC(50)=1.2 nM) and inhibited the insulin responses to both carbachol and exendin-4. Kisspeptin-13 blocked arginine-induced insulin secretion without affecting the glucagon or somatostatin responses to this amino acid, thus indicating that kisspeptin-13 influences B cells directly, rather than through an A- or D-cell paracrine effect. The reduction of the insulin response to exendin-4 induced by kisspeptin-13 was also observed in pertussis toxin-treated rats, thus suggesting an inhibition independent of G(i) proteins. In view of the potent insulinostatic effect of kisspeptin-13, it is tempting to speculate that kisspeptins may be implicated in the regulation of B-cell secretion.     

Effects of gamma-aminobutyric acid on insulin, glucagon, and somatostatin release from isolated perfused dog pancreas

The effects of gamma-aminobutyric acid (GABA) on the secretion of insulin, glucagon, and somatostatin were studied in the isolated dog pancreas. Insulin secretion was inhibited in a dose-related fashion for 10 min or more by as little as 1 microM GABA. A prompt but small and transitory rise in somatostatin secretion, lasting only 1 min, occurred at GABA concentrations of 10 and 100 microM, levels that exert inhibitory effects on nervous tissue. Bicuculline, a GABA antagonist, inhibited insulin secretion and did not antagonize GABA-mediated insulin inhibition. The results suggest that GABA in concentrations that are known to exist in islet tissue can influence the secretion of islet hormones.     

Effects of K+ and arginine on insulin, glucagon, and somatostatin release from the in vitro perfused rat pancreas

Rat pancreases were perfused in vitro for 5-min periods with K+ alone (8, 10, and 12 mM) or in the presence of arginine (10 mM). Alone, K+ caused bursts of insulin, glucagon, and somatostatin (SRIF) release; with arginine, it caused a burst of insulin and sustained SRIF release, but caused sustained suppression of glucagon. This suppression correlated better with SRIF than insulin release. Therefore, if a paracrine effect is responsible for the inhibition of glucagon secretion under these circumstances, SRIF is a more likely candidate than insulin.     

Effects of endotoxin and interleukin-1 on glucagon and insulin secretion from the perfused rat pancreas.

This study evaluated the in vivo effects of endotoxin and interleukin-1 (IL-1) on the simultaneous secretion of glucagon and insulin. The hypothesis that endotoxin, or IL-1 as a mediator, induces hyperglucagonemia secondary to pancreatic hypersecretion of glucagon was examined. Hormone secretion was measured using the in vitro perfused rat pancreas preparation. In response to an arginine stimulus, glucagon secretion was neither stimulated nor inhibited significantly by endotoxin or IL-1. Insulin secretion was significantly potentiated with both endotoxin and IL-1. In response to a low-glucose stimulus, glucagon secretion was significantly inhibited by endotoxin treatment, while insulin secretion was increased by endotoxin or IL-1. These results indicate that neither endotoxin nor IL-1 treatment resulted in glucagon hypersecretion, although either of these agents could induce insulin hypersecretion. Thus, the mechanism of endotoxin-induced hyperglucagonemia cannot be explained by a hypersecretory state of glucagon secretion. The parallel respective effects of endotoxin and IL-1 on glucagon and insulin secretion are consistent with the concept that IL-1 mediates some of the effects of endotoxin on the endocrine pancreas.     

Effect of glucose/sulfonylurea interaction on release of insulin, glucagon, and somatostatin from isolated perfused rat pancreas.

The effect of a sulfonylurea, glibenclamide, on the release of insulin, glucagon, and somatostatin was studied in the isolated perfused rat pancreas. At glucose concentrations of 1.1 mM or less, the drug stimulated somatostatin release, whereas glucagon release, after 2-3 min of increase, was markedly inhibited. Insulin release was moderately stimulated, and maximal release occurred relatively late. A moderate glucose load (6.7 mM) inhibited glibenclamide-induced release of somatostatin, whereas the two in combination exerted an additive action on insulin release. Greater glucose loads, which by themselves would stimulate somatostatin release, only marginally suppressed glibenclamide-induced somatostatin release. The insulinogenic effect of these glucose levels was not modified by glibenclamide. Glibenclamide may thus stimulate both the alpha and beta as well as delta cells of the pancreas, depending on glucose concentration. We suggest a paracrine (local) interaction of somatostatin with the alpha and beta cells, which has an important role in the kinetics of insulin and glucagon release induced by sulfonylureas.     

Neurotransmitters partially restore glucose sensitivity of insulin and glucagon secretion from perfused streptozotocin-induced diabetic rat pancreas

Summary To elucidate the mechanisms of insensitivity of hormone secretion to glucose in streptozotocin-induced diabetic rat islets, we investigated the effects of acetylcholine (ACh) and norepinephrine on insulin and glucagon secretion in response to changes in glucose concentration, using perfused pancreas preparations. Basal insulin secretion at a blood glucose level of 5.6 mmol/l was significantly higher and basal glucagon secretion significantly lower in streptozotocin-induced diabetic rats than in controls, and neither high (16.7 mmol/l) nor low (1.4 mmol/l) blood glucose concentrations influenced insulin or glucagon secretion. Addition of 10^–6 mol/l ACh to the perfusate increased glucose-stimulated insulin secretion. Also, 10^–6 mol/l ACh, 10^–7 mol/l norepinephrine, as well as a combination of both, induced marked glucagon secretion, this was suppressed by high blood glucose level. Although simultaneous addition of 10^–6 mol/l ACh and 10^–7 mol/l norepinephrine induced only a slight increase in glucagon secretion in response to glucopenia, there was a significant increase in glucagon secretion in conjunction with an ambient decrease in insulin. Histopathological examination revealed a marked decline in acetylcholinesterase and monoamine-oxidase activities in the islets of streptozotocin-induced diabetic rats. We speculate that reduction of the potentiating effects of ACh and norepinephrine lessens glucose sensitivity of islet beta and alpha cells in this rat model of diabetes.Abbreviations STZ Streptozotocin - STZD streptozotocin-induce diabetic - ACh acetylcholine - AChE acetylcholinesterase - NE norepinephrine - MAO monoamine-oxidase