Katsuhiko Ito

Exogenous insulin dose-dependently suppresses glucopenia-induced glucagon secretion from perfused rat pancreas

To clarify the role of insulin in modulating the glucagon response to glucose concentration changes, we investigated the effects of exogenous insulin (10 mU/mL, 100 mU/mL, and 3.3 U/mL) on responses to high glucose (5.6-->16.7 mmol/L), low glucose (5.6-->1.4 mmol/L), and arginine (10 mmol/L) stimulation using the perfused rat pancreas. Although glucagon levels were slightly suppressed by all of the exogenous insulin concentrations tested for the initial few minutes at 5.6 mmol/L glucose, baseline levels were maintained thereafter. Glucagon responses to high or normal glucose concentrations were not altered, but glucopenia-induced glucagon secretion was significantly suppressed as compared with that of controls (0.77 +/- 0.14 ng/min [10 mU/mL, n = 5], 0.55 +/- 0.14 ng/min [100 mU/mL, n = 5], 0.27 +/- 0.13 ng/min [3.3 U/mL, n = 5] v 1.38 +/- 0.20 ng/min [controls, n = 9], P < 0.05, respectively). The first phase of the glucagon response to arginine was potentiated (2.03 +/- 0.24 v 1.17 +/- 0.22 ng/min, P < .05) by 10 mU/mL exogenous insulin. The second phase of the glucagon response to arginine was significantly suppressed in the presence of higher concentrations of exogenous insulin (1.16 +/- 0.23 ng/min [100 mU/mL], 0.96 +/- 0.08 ng/min [3.3 U/mL] v 1.57 +/- 0.17 ng/min, P < .05, respectively). These results suggest that glucagon secretion is modified by the combined suppressive effects of glucose and insulin, although it is mainly glucose that mediates glucagon secretion in the physiological glucose range. Glucopenia- or arginine-induced glucagon secretion is suppressed by insulin.

Effects of D-Phenylalanine- Derivative Hypoglycemic Agent A-4166 on Pancreatic α- and β-Cells: Comparative Study with Glibenclamide

We have reported that N-[(trans-4-isopropyl-cyclohexyl)-carbonyl]-D-phenylalanine (A-4166) stimulates insulin secretion in animal studies. To further elucidate the mechanisms underlying the actions of

Effects of α2- and β-adrenergic agonism on glucagon secretion from perfused pancreata of normal and streptozocin-induced diabetic rats

Insulin secretion is known to be inhibited by α2-adrenergic agonism and stimulated by β-adrenergic agonism in both experimental animals and humans. In contrast, adrenergic regulation of glucagon secretion remains controversial. This study was designed to determine the effects of α2- and β-adrenergic agonism on islet α cells, using isolated perfused pancreata of normal and streptozocin-induced diabetic (STZ-D) rats. The α2-adrenoceptor agonist clonidine at a concentration of 10−7 mol/L significantly stimulated glucagon secretion as compared with basal levels in both normal (1,286 ± 90 v 417 ± 53 ng/L, P < .01) and STZ-D rats (551 ± 86 v 130 ± 19 ng/L, P < .01). Also, the β-adrenoceptor agonist isoproterenol at a concentration of 10−7 mol/L significantly stimulated glucagon secretion as compared with basal levels in both normal (751 ± 130 v 347 ± 41 ng/L, P < .05) and STZ-D rats (182 ± 22 v 92 ± 20 ng/L, P < .01). Furthermore, these α2- and β-agonistic effects were almost completely inhibited in the presence of the α2-adrenoceptor antagonist yohimbine and the β-agonistic effects were almost propranolol at a concentration of 10−6 mol/L, respectively. Insulin secretion was markedly reduced in STZ-D rats. These results suggest that even in a severely diabetic state, not only β- but also α2-adrenergic agonism stimulates glucagon secretion from rat pancreatic α cells.

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

SummaryTo 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.

Effects of N-[(trans-4-isopropylcyclohexyl)-carbonyl]-D-phenylalanine (A-4166) on insulin and glucagon secretion in isolated perfused rat pancreas.

N-[(trans-4-isopropylcyclohexyl)-carbonyl]-D-phenylalanine (A-4166) has a structure which differs from those of other known blood glucose-lowering agents including sulfonylureas. It has been shown that oral administration of A-4166 exerts blood glucose-lowering effects in animal in vivo studies. In the present study, we investigated the effects of A-4166 on insulin and glucagon secretion at several glucose concentrations using isolated perfused rat pancreas preparations. Both 3.0 and 30 mumol/l A-4166 significantly stimulated insulin secretion as compared with basal levels at glucose concentrations of 8.0 and 11.0 mmol (p < 0.01 and p < 0.05, respectively). In contrast, glucagon secretion was not affected by administration of A-4166 up to 30 mumol/l at these glucose concentrations. At a glucose concentration of 5.6 mmol/l, neither 0.3 nor 3.0 mumol/l A-4166 produced significant changes in insulin and glucagon secretion. However, A-4166 at 30 mumol/l significantly stimulated insulin secretion and inhibited glucagon secretion as compared with basal levels (p < 0.01 and p < 0.01, respectively). We conclude that A-4166 at 3.0 and 30 mumol/l directly stimulates insulin secretion but has little effect on glucagon secretion in isolated perfused rat pancreas at glucose concentrations of 8.0 and 11.0 mmol/l. these results, taken together with previously published data, suggest that oral administration of A-4166 could be a useful strategy for stimulating endogenous insulin secretion in non-insulin-dependent diabetic patients.

Adrenoceptor antagonists, but not guanethidine, reduce glucopenia-induced glucagon secretion from perfused rat pancreas

This study was designed to investigate (1) whether norepinephrine is released in response to glucopenia in vitro, thereby stimulating glucagon secretion and, (2) the modulating effects of norepinephrine on insulin and glucagon secretion, using isolated perfused rat pancreas preparations. Simultaneous addition of the adrenergic receptor antagonists yohimbine, prazosin and propranolol, each at a concentration of 10-(5) mol/l, significantly potentiated glucose-stimulated insulin secretion (6.23 +/- 0.76 vs. 2.11 +/- 0.72 (control) nmol/min, P < 0.01), and suppressed glucopenia-induced glucagon secretion (0.59 +/- 0.10 vs. 1.34 + 0.18 (control) ng/min, P < 0.05). Also, 10-(5) mol/l yohimbine alone significantly potentiated glucose-stimulated insulin secretion (4.86 +/- 0.50 nmol/min, P < 0.05). The norepinephrine release inhibitor, guanethidine, significantly inhibited tyramine-induced secretion of both norepinephrine (7.86 +/- 0.77 vs. 49.7 +/- 2.3 nmol/min, P < 0.01) and glucagon (0.31 +/- 0.08 vs. 1.21 +/- 0.15 ng/min, P < 0.01), but exerted no effects on glucopenia-induced secretion of either norepinephrine or glucagon. We conclude that these results further support the concept that the neurotransmitter norepinephrine is released in response to glucopenia in vitro, and modulates insulin and glucagon secretion. Our data do not, however, provide evidence indicating that glucopenia-induced glucagon secretion is mainly mediated by activation of sympathetic nerve terminals around the alpha-cells in the isolated perfused rat pancreas.

Effects of diazoxide on α- and β-cell function in isolated perfused rat pancreas

Abstract To elucidate the effects of diazoxide on insulin and glucagon secretion at normal, high and low glucose concentrations and 10 mmol/l arginine, we performed pancreatic perfusion experiments. The insulin secretion rate in response to 16.7 mmol/l glucose was dose-dependently suppressed by concomitant infusion of diazoxide (100 and 300 μmol/l). Both the first and second phases of glucose-stimulated insulin secretion were significantly reduced in the presence of diazoxide as compared with controls. Basal glucagon secretion rate at 5.6 mmol/l glucose was significantly reduced by the administration of both 100 and 300 μmol/l diazoxide. Furthermore, the glucagon secretion rate at a high glucose concentration (16.7 mmol/l) was significantly lower with 300 μmol/l diazoxide than in the control. The glucagon secretion rate with glucopenia (1.4 mmol/l) was also significantly lower with 100 and 300 μmol/l diazoxide than in the control. The insulin secretion rate in response to 10 mmol/l arginine was also dose-dependently suppressed by concomitant infusion of diazoxide. The glucagon secretion rate in response to 10 mmol/l arginine was, however, significantly higher with 100 μmol/l diazoxide while not being significantly different with 300 μmol/l diazoxide. These findings suggest that some mechanism(s) which can be inhibited by diazoxide is involved in glucagon, as well as insulin, secretion in isolated perfused rat pancreas.

Defective Insulin and Glucagon Secretion in Isolated Perfused Pancreata of Diabetic WBN/Kob Rats

To elucidate the pathophysiology of diabetes mellitus in male WBN/Kob rats, we performed pancreatic perfusion experiments and histopathological studies. Intraperitoneal glucose tolerance tests showed a diabetic pattern in 12-month-old WBN/Kob rats. In perfused pancreata of WBN/Kob rats, both the first and the second phases of insulin secretion in response to a 16.7 mM glucose challenge were markedly reduced compared with those in age-matched Wistar rats (p < 0.01, respectively). Furthermore, the insulin secretion rate in response to glucopenia (1.4 mM) was significantly higher (p < 0.05) and the decrement in insulin secretion was significantly lower (p < 0.05) in WBN/Kob rats. The decrement in glucagon secretion with 16.7 mM glucose was significantly blunted (p < 0.001), and the glucagon secretion rate in response to glucopenia was also significantly lower in WBN/Kob rats than in controls (p < 0.01). Although insulin secretion in response to 10 mM arginine was also moderately reduced in WBN/Kob rats (p < 0.05), the glucagon secretion rates in response to 10 mM arginine were similar in the two groups. Histopathological examination revealed widespread disappearance of acinar cells and islets, inflammatory changes, and marked fibrosis in the pancreata of WBN/Kob rats. Irnrnunohistochemical studies showed decreased numbers of B cells in the islets of WBN/Kob rats. These findings suggest that this WBN/Kob rat strain is a useful model for studying not only pathogenesis, but also pathophysiology, i.e., defective hormonal secretion, in some types of human diabetes mellitus.