Effects of neuromedin B on insulin and glucagon release from the isolated perfused rat pancreas

The effect of neuromedin B (NMB) on insulin and glucagon release was studied in isolated perfused rat pancreas. Infusion of NMB (10 nM, 100 nM and 1 microM) did not affect the insulin release under the perusate conditions of 5.5 mM glucose plus 10 mM arginine and 11 mM glucose plus 10 mM arginine, although 10 nM NMB tended to slightly suppress it under the perfusate condition of 5.5 mM glucose alone. The degree of stimulation of insulin release provoked by the addition of 5.5 mM glucose to the perfusate was not affected by the presence of 10 nM NMB. The glucagon release was slightly stimulated by the infusion of 100 nM and 1 microM NMB but not by 10 nM NMB under the perfusate condition of 5.5 mM glucose plus 10 mM arginine. The effect of C-terminal decapeptide of gastrin releasing peptide (GRP-10) was also examined and similar results were obtained; 10 nM and 100 nM GRP-10 did not affect insulin release and 100 nM GRP-10 stimulated glucagon release under the perfusate condition of 5.5 mM glucose plus 10 mM arginine. The present results concerning glucagon release are consistent with the previous results obtained with isolated perfused canine and porcine pancreas. However, the results regarding insulin release are not. Species differences in insulin release are also evident with other neuropeptides such as substance P and the mechanism of such differences remains for be clarified.     

Effect of glucagon antiserum on somatostatin,glucagon and insulin release from the isolated perfused rat pancreas

In order to elucidate the effect of glucagon antiserum on the endocrine pancreas, the release of somatostatin, glucagon, and insulin from the isolated perfused rat pancreas was studied following the infusion of arginine both with and without pretreatment by glucagon antiserum. Various concentrations of arginine in the presence of 5.5 mM glucose stimulated both somatostatin and glucagon secretion. However, the responses of somatostatin and glucagon were different at different doses of arginine. The infusion of glucagon antiserum strongly stimulated basal secretion in the perfusate total glucagon (free + antibody bound glucagon) and also enhanced its response to arginine, but free glucagon was undetectable in the perfusate during the infusion. On the other hand, the glucagon antiserum had no significant effect on either insulin or somatostatin secretion. Moreover, electron microscopic study revealed degrannulation and vacuolization in the cytoplasm of the A cells after exposure to glucagon antiserum, suggesting a hypersecretion of glucagon, but no significant change was found in the B cells or the D cells. We conclude that in a single pass perfusion system glucagon antiserum does not affect somatostatin or insulin secretion, although it enhances glucagon secretion.     

Effect of exercise training on insulin and glucagon release from perfused rat pancreas

The effect of physical training on insulin and glucagon release in perfused rat pancreas was examined in the spontaneously exercised group running in a wheel cage an average of 1.4 km/day for 3 weeks and in the sedentary control group kept in the cage whose rotatory wheel was fixed on purpose. Pancreatic immunoreactive insulin (IRI) responses to glucose and arginine were reduced by 28% and 47.8% respectively in trained rats compared with untrained rats, while IRI content of the pancreas was similar in these two groups. The demonstrated decrease in insulin secretion of the beta-cell of the trained rats, in response to the glucose and arginine stimulations, may be functional in nature. On the other hand, neither pancreatic glucagon immunoreactivity (GI) response to glucose and arginine nor GI content of the pancreas was modified by exercise training. These results demonstrate that exercise training reduces IRI responses to glucose as well as to arginine stimulations, but does not modify any secretory response of pancreatic GI.     

Failure of parathormone to affect insulin and glucagon release from the perfused rat pancreas.

Parathormone (0.15 U/ml) failed to affect the rate of glucagon and insulin release by the perfused rat pancreas exposed to glucose in either low (3.3 mM) or high (8.3 mM) concentration. Parathormone also failed to interfere with the suppressive effect of glucose (16.6mM) upon glucagon release and its stimulatory action upon insulin secretion. Likewise, the biphasic release of both glucagon and insulin evoked by arginine (10.0 mM) in the presence of glucose (8.3 mM) was unaffected by parathormone. These findings suggest that the endocrine pancreas may not be a target organ for any direct and immediate action of parathormone.     

Stimulation by prostaglandin E2 of glucagon and insulin release from isolated rat pancreas.

To ascertain whether prostaglandins (PG) may play a role in the secretion of glucagon and in an attempt to elucidate the conflicting observations on the effects of PG on insulin release, the isolated intact rat pancreas was perfused with solutions containing 1.1 x 10(-9) to 1.8 x 10(-5)m PGE2. In the presence of 5.6 mM glucose significant increments in portal venous effluent levels of glucagon and insulin were observed in response to minimal concentrations of 2.8 X 10(-8) and 1.4 X 10(-7) PGE2, respectively; a dose-response relationship was evident for both hormones at higher concentrations of PGE2. When administered over 60 seconds, 1.4 X 10(-6)M PGE2 resulted in a significant increase in glucagon levels within 24 seconds and in insulin within 48 seconds. Ten-minute perfusions of 1.4 X 10(-6)M PGE2 elicited biphasic release of both islet hormones; Phase I glucagon release preceded that of insulin. Both phases of the biphasic glucagon and insulin release which occurred in response to 15-minute perfusions of 10 mM arginine were augmented by PGE2. These observations indicate that PGE2 can evoke glucagon and insulin release at concentrations close to those observed by others in the extracts of rat pancreas. We conclude that PG may be involved in the regulation of secretion of glucagon and insulin and may mediate and/or modify the pancreatic islet hormone response to other secretagogues.     

Effects of serotonin, of its biosynthetic precursors and of the anti-serotonin agent metergoline on the release of glucagon and insulin from rat pancreas.

We have evaluated the effect of serotonin (5-HT) and of its biosynthetic precursors 5-Hydroxytryptophan (5-HTP) and tryptophan (TRP) on the release of immunoreactive glucagon (IRG) and insulin (IRI) from isolated islets and pieces of pancrease of the rat. In isolated islets, 5-HT inhibited the IRI response to a high glucose concentration (3.0 mg/ml), without affecting the IRG response to either a low (0.5 mg/ml) or a high glucose concentration; TRP stimulated the IRG and IRI response to the low glucose concentration, while 5-HTP was ineffective. When pieces of pancreas were used, 5-HT and 5-HTP inhibited IRG response to both glucose concentrations, while IRI release was inhibited only by 5-HT. The anti-5-HT agent metergoline enhanced the release of IRG and IRI by pieces of pancreas at both glucose concentrations. The results indicate that exogenous and endogenous 5-HT inhibit basal as well as glucose-mediated IRG and IRI release; that isolated islets are less sensitive than pieces of pancreas to the inhibitory effect of 5-HT and that TRP acts as an amino acid and not as a precursor of 5-HT.     

Effects of pancreastatin on insulin, glucagon and somatostatin secretion by the perfused rat pancreas

Pancreastatin is a novel peptide, isolated from porcine pancreatic extracts, which has been shown to inhibit glucose-induced insulin release "in vitro". To achieve further insight into the influence of pancreastatin on pancreatic hormone secretion, we have studied the effects of this peptide on unstimulated insulin, glucagon and somatostatin output, as well as on the responses of these hormones to glucose and to tolbutamide in the perfused rat pancreas. Pancreastatin strongly inhibited unstimulated insulin release as well as the insulin responses to glucose and to tolbutamide. It did not significantly affect glucagon or somatostatin output under any of the above-mentioned conditions. These findings suggest that pancreastatin inhibits B-cell secretory activity directly, and not through an A-cell or D-cell paracrine effect.     

Effects of porcine diazepam-binding inhibitor on insulin and glucagon secretion in vitro from the rat endocrine pancreas.

Porcine diazepam-binding inhibitor (pDBI) is a novel peptide that has been isolated from the small bowel of the pig, and that occurs also in the islet D-cells. We have studied its effects on hormone release in vitro from the endocrine pancreas of the rat. In isolated islets, pDBI (10(-9)-10(-6)M) did not affect basal insulin release at 3.3 mM glucose, whereas stimulated release at 8.3 mM glucose was dose-dependently suppressed by 32-69% (P less than 0.01). Furthermore, insulin secretion stimulated by either 16.7 mM glucose or 1 mM IBMX (3-isobutyl-1-methylxanthine) or 1 micrograms/ml glibenclamide was suppressed by pDBI at 10(-8) M (by 28-30%, P less than 0.05) and 10(-7) M (by 43-47%, P less than 0.01). In contrast, islet insulin secretion induced by 20 mM arginine was unaffected by these concentrations of pDBI. In the perfused rat pancreas, pDBI (10(-8) M) enhanced by 30% (P less than 0.05) the first phase (0-5 min) of arginine-stimulated insulin release, whereas the second phase (5-20 min) was unchanged. Moreover, pDBI suppressed by 28% (P less than 0.05) the second phase of arginine-induced glucagon release. Arginine-induced somatostatin release was not significantly affected by the peptide. Since pDBI immunoreactivity has been localized also to islet D-cells, the present results suggest that pDBI may act as a local modulator of islet hormone release.     

Insulin and glucagon release from the ventral and dorsal parts of the perfused pancreas of the rat. Effects of glucose, arginine, glucagon and carbamylcholine

The ventral and the dorsal parts of the rat pancreas were perfused separately via either the superior mesenteric artery (0.6 ml/min) or the coeliac artery (1.4 ml/min). Control perfusions were performed via both arteries (2 ml/min). Expressed relative to the weight of tissue, the insulin content was comparable in the ventral and dorsal parts whereas the glucagon content was 2.5 times lower in the ventral than dorsal part. In comparison to the dorsal or total pancreas, the insulin secretory activity of the ventral pancreas was markedly decreased in response to either an elevation of the glucose concentration or the administration of carbamylcholine or arginine. The difference between the ventral and dorsal response was less marked at low glucose concentrations (3.3 or 7.0 mmol/l) and, possibly, in response to glucagon. In the case of glucagon release, a decreased response of the ventral pancreas was only observed when glucagon output was fully stimulated by the administration of arginine at a low glucose concentration. These results indicate that the B cell in the ventral pancreas responds poorly to several stimuli. There was little evidence to support the involvement of endogenous glucagon in the diminished sensitivity of the ventral B cells.     

Somatostatin, insulin and glucagon secretion by the perfused pancreas from the cysteamine-treated rat

In rats, administration of a single dose of cysteamine (300 mg/kg, intragastrically) induces a depletion of pancreatic somatostatin content (approximately 60%) without modifying pancreatic insulin or glucagon content. In perfused pancreases from cysteamine-treated rats, there was a lack of somatostatin response to glucose, arginine or tolbutamide. In the absence of stimulated somatostatin release, the secretory responses of insulin and glucagon to glucose, to arginine, and to tolbutamide were not significantly different from those observed in pancreases from control rats. Our data do not support the concept that pancreatic somatostatin plays a major role in the control of insulin and glucagon release.     

Somatostatin inhibits insulin and glucagon release by monolayer cell cultures of rat endocrine pancreas

Dihydrosomatostatin (0.001–1.0 ug/ml) inhibited both insulin and glucagon secretion by monolayer cell cultures of newborn rat pancreas. When cultures were incubated with somatostatin and then rinsed, the effect of somatostatin appeared to last longer on the pancreatic alpha cell than on the beta cell as indicated by a more prolonged inhibition of glucagon secretion than of insulin release. Submaximal inhibition of glucose-stimulated insulin release by somatostatin was partially reversed by increasing the concentration of glucose. We conclude that the effect of somatostatin appears to be mediated directly on the pancreatic endocrine cells.     

Homologous pancreastatin inhibits insulin secretion without affecting glucagon and somatostatin release in the perfused rat pancreas.

The identification of pancreastatin in pancreatic extracts prompted the investigation of its effects on islet cell function. However, in most of the investigations to date, pig pancreastatin was tested in heterologous species. Since there is great interspecies variability in the amino acid sequence of pancreastatin, we have investigated the influence of rat pancreastatin on insulin, glucagon and somatostatin secretion in a homologous animal model, namely the perfused rat pancreas. During 5.5 mM glucose infusion, pancreastatin (40 nM) inhibited insulin secretion (ca. 40%, P less than 0.025) as well as the insulin responses to 10 mM arginine (ca. 50%, P less than 0.025) and to 1 nM vasoactive intestinal polypeptide (ca. 50%; P less than 0.05). Pancreastatin failed to significantly modify glucagon or somatostatin release under any of the above experimental conditions. In addition, a lower pancreastatin concentration (15.7 nM) markedly suppressed the insulin release evoked by 11 mM glucose (ca. 85%, P less than 0.05). Our present observations reinforce the concept that pancreastatin is an effective inhibitor of insulin secretion, influencing the B-cell function directly and not through an A-cell or D-cell paracrine effect.     

Effects of salicylate on insulin and glucagon secretion by the isolated and perfused rat pancreas

The effects of sodium salicylate, a prostaglandin synthesis inhibitor, on glucose-induced secretion of insulin and glucagon by the isolated perfused rat pancreas have been studied. Sodium salicylate inhibited both basal (2.8 mM glucose) and stimulated (16.7 mM glucose) insulin release in a dose dependent manner (1, 5 and 10 mM). This inhibition is not interpretable in terms of a simple inhibition of cyclooxygenase by sodium salicylate. Basal glucagon release was not changed by 1 mM sodium salicylate but the latter partially blocked its inhibition by 16.7 mM glucose. Higher doses of sodium salicylate (5 and 10 mM) inhibited basal glucagon secretion without affecting its response to 16.7 mM glucose. These findings suggest a predominant stimulatory action of endogenous prostaglandins on glucagon release.     

Effect of the bonito insulin on the insulin release from monolayer culture of rat pancreas.

The effect of bonito insulin on insulin release was examined in the monolayer culture of rat pancreatic beta-cells. The beta-cells were preincubated for 5 to 20 hr with or without a small dose (100 microunits/ml) of bonito insulin in the medium containing 100 mg% glucose. And then, they were incubated in 300 mg% glucose alone or together with bonito insulin for 5 hr. There was no significant difference between the IRI release from these beta-cells with or without bonito insulin. The concentration of bonito insulin was augmented from 100 microunits/ml to 500, 1,000 and 2,000 microunits/ml. A significant inhibitory effect on the glucose-induced insulin release was observed only after the preincubation for 20 hr with 2,000 microunits/ml of bonito insulin.     

Stimulation by prostaglandin E2 of glucagon and insulin release from isolated rat pancreas

To ascertain whether prostaglandins (PG) may play a role in the secretion of glucagon and in an attempt to elucidate the conflicting observations on the effects of PG on insulin release, the isolated intact rat pancreas was perfused with solutions containing 1.1 × 10⁻⁹ to 1.8 × 10⁻⁵M PGE₂. In the presence of 5.6 mM glucose significant increments in portal venous effluent levels of glucagon and insulin were observed in response to minimal concentrations of 2.8 × 10⁻⁸ and 1.4 × 10⁻⁷M PGE₂, respectively; a dose-response relationship was evident for both hormones at higher concentrations of PGE₂. When administered over 60 seconds, 1.4 × 10⁻⁶M PGE₂ resulted in a significant increase in glucagon levels within 24 seconds and in insulin within 48 seconds. Ten-minute perfusions of 1.4 × 10⁻⁶M PGE₂ elicited biphasic release of both islet hormones; Phase I glucagon release preceded that of insulin. Both phases of the biphasic glucagon and insulin release which occurred in response to 15-minute perfusions of 10 mM arginine were augmented by PGE₂. These observations indicate that PGE₂ can evoke glucagon and insulin release at concentrations close to those observed by others in the extracts of rat pancreas. We conclude that PG may be involved in the regulation of secretion of glucagon and insulin and may mediate and/or modify the pancreatic islet hormone response to other secretagogues.     

Stimulation by prostaglandin E2 of glucagon and insulin release from isolated rat pancreas

To ascertain whether prostaglandins (PG) may play a role in the secretion of glucagon and in an attempt to elucidate the conflicting observations on the effects of PG on insulin release, the isolated intact rat pancreas was perfused with solutions containing 1.1 × 10⁻⁹ to 1.8 × 10⁻⁵M PGE₂. In the presence of 5.6 mM glucose significant increments in portal venous effluent levels of glucagon and insulin were observed in response to minimal concentrations of 2.8 × 10⁻⁸ and 1.4 × 10⁻⁷M PGE₂, respectively; a dose-response relationship was evident for both hormones at higher concentrations of PGE₂. When administered over 60 seconds, 1.4^−10−6M PGE₂ resulted in a significant increase in glucagon levels within 24 seconds and in insulin within 48 seconds. Ten-minute perfusions of 1.4 × 10⁻⁶M PGE₂ elicited biphasic release of both islet hormones; Phase I glucagon release preceded that of insulin. Both phases of the biphasic glucagon and insulin release which occurred in response to 15-minute perfusions of 10 mM arginine were augmented by PGE₂. These observations indicate that PGE₂ can evoke glucagon and insulin release at concentrations close to those observed by others in the extracts of rat pancreas. We conclude that PG may be involved in the regulation of secretion of glucagon and insulin and may mediate and/or modify the pancreatic islet hormone response to other secretagogues.     

Adrenergic modulation of insulin and glucagon secretion from the isolated perfused rat pancreas

In order to observe the effect of the adrenergic system on pancreatic glucagon secretion in the isolated perfused rat pancreas, phenylephrine, an alpha-adrenergic agonist, and isoproterenol, a beta-adrenergic agonist, were added to the perfused solution. 1.2 microM phenylephrine suppressed glucagon secretion at 2.8 mM glucose, and it also decreased insulin secretion at 11.1 mM glucose. 240 nM isoproterenol enhanced glucagon secretion not only at 2.8 mM glucose, but also at 11.1 mM glucose, as well as insulin secretion at 11.1 mM. In order to study the role of intra-islet noradrenalin, phentolamine, an alpha-adrenergic antagonist, and propranolol, a beta-adrenergic antagonist, were infused with the perfused solution. 10 and 100 microM phentolamine caused an increase in insulin secretion, and 25 microM propranolol decreased insulin secretion, while they did not cause any change in glucagon secretion. From these results, it can be concluded that alpha-stimulation suppresses not only insulin but also glucagon secretion, while beta-stimulation stimulates glucagon secretion, as well as insulin secretion. Intra-islet catecholamine may have some effect on the B cell, whereas it seems to have no influence on the A cell.