Kazunori Koyama

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 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.

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.

Arachidonic acid metabolites and α2-adrenoceptor-mediated glucagon secretion in rats

Effects of phospholipase A2 inhibitor, cyclooxygenase inhibitor and lipoxygenase inhibitor on glucagon secretion induced by the alpha 2-adrenergic agonist clonidine were studied in the isolated perfused rat pancreas. The phospholipase A2 inhibitor mepacrine at 25 and 50 mumol/l significantly inhibited glucagon secretion induced by 0.1 mumol/l clonidine (P less than 0.01, respectively), whereas 5 mumol/l mepacrine did not affect clonidine-induced glucagon secretion. Also, both 100 mumol/l acetylsalicylic acid (cyclooxygenase inhibitor) and 100 mumol/l caffeic acid (lipoxygenase inhibitor) significantly inhibited clonidine-induced glucagon secretion (P less than 0.01, respectively), whereas neither 10 mumol/l acetylsalicylic acid nor 10 mumol/l caffeic acid affected clonidine-induced glucagon secretion. None of the drugs at the tested concentrations affected insulin secretion at a glucose concentration of 5.5 mmol/l. These results suggest that not only cyclooxygenase metabolites but also lipoxygenase metabolites are involved in the stimulation of glucagon secretion mediated through the alpha 2-adrenergic receptors in perfused rat pancreas.

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.

Effect of α-adrenergic blockade on blood pressure, glucose, and lipid metabolism in hypertensive patients with non-insulin-dependent diabetes mellitus

Abstract To clarify the long-term effects of α-adrenergic blockade on blood pressure, glucose, and lipid metabolism, a slective α 1 -adrenergic inhibitor (prazosin, 1.0 to 2.0 mg/day in divided doses) was administered as a single antihypertensive agent to 10 (four men and six women, aged 52 to 76 years) hypertensive patients (systolic blood pressure [SBP] ≥ 150 mm Hg or diastolic blood pressure [DBP] ≥ 90 mm Hg) with non-insulin-dependent diabetes mellitus (NIDDM) for up to 20 weeks. Blood pressure, glucose tolerance and immunoreactive insulin (IRI) response to 75 gm oral glucose load, hemoglobin A 1 (Hb A 1 ), serum lipid profile, and serum apolipoprotein were examined before and after treatment. SBP and DBP were significantly reduced at 20 weeks after treatment with the selective α 1 -adrenergic inhibitor (SBP 167 ± 6 mm Hg versus 152 ± 7 mm Hg; DBP 81 ± 3 mm Hg versus 76 ± 3 mm Hg, ( p p α 1 -inhibitor treatment compared with the baseline data before treatment; the level of Hb A 1 was not significantly changed at 4 and 20 weeks after treatment. Total cholesterol and free fatty acid (FFA) concentrations were significantly reduced at 20 weeks (total cholesterol 213 ± 13 mg/dl versus 196 ± 10 mg/dl [5.52 ± 0.34 mmol/L versus 5.08 ± 0.26 mmol/L], FFA 0.67 ± 0.11 mEq/L versus 0.48 ± 0.11 mEq/L; p p p 1 , B, C 2 , C 3 , and E was observed during treatment. These data suggest that treatment with a selective α 1 -adrenergic inhibitor elicits a favorable effect on blood pressure and lipid metabolism without causing a deterioration in blood glucose control in hypertensive NIDDM patients.

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.