Stimulus-secretion coupling of arginine-induced insulin release. Insulinotropic action of agmatine

Agmatine causes a concentration-related (0.1-1.0 mM) stimulation of insulin secretion by rat pancreatic islets exposed to D-glucose (5.6-11.1 mM) but, like L-arginine, fails to affect insulin release either in the absence of glucose or at a high concentration (16.7 mM) of the hexose. Agmatine, which accumulates in islet cells, also stimulates 45Ca net uptake by the islets. The secretory response to agmatine represents a delayed process. It is enhanced by theophylline and suppressed by antimycin A or the absence of extracellular Ca2+. The insulinotropic action of agmatine is compatible with the view that endogenously formed polyamines may play a role in the secretory response of islet cells to L-arginine.     

Interaction of the islet nitric oxide system with L-arginine-induced secretion of insulin and glucagon in mice.

1. Several recent in vitro studies have suggested that production of nitric oxide (NO) from the islet NO system may have an important regulatory influence on the secretion of insulin and glucagon. In the present paper we have investigated, mainly with an in vivo approach, the influence and specificity of the NO synthase (NOS) blocker NG-nitro-L-arginine methyl ester (L-NAME) on L-arginine-induced secretion of insulin and glucagon. 2. In freely fed mice, L-NAME pretreatment (1.2 mmol kg-1) influenced the dynamics of insulin and glucagon release following an equimolar dose of L-arginine, the specific substrate for NOS activity, in that the NOS inhibitor enhanced the insulin response but suppressed the glucagon responses. This was reflected in a large decrease in the plasma glucose levels of the L-NAME pretreated animals. 3. L-NAME pretreatment did not influence the insulin and glucagon secretory responses to the L-arginine-enantiomer D-arginine, which cannot serve as a substrate for NOS activity. 4. Replacing L-NAME pretreatment by pretreatment with D-arginine or L-arginine itself, which both carry the same cationic change and are devoid of NOS inhibitory properties, did not mimic the effects of L-NAME on L-arginine-induced hormone release. 5. Fasting the animals for 24 h totally abolished the L-NAME-induced potentiation of L-arginine stimulated insulin release suggesting that the sensitivity of the beta-cell secretory machinery to NO-production is greatly changed in the fasting state. However, the L-NAME-induced suppression of L-arginine stimulated glucagon release was unaffected by starvation. 6. In isolated islets from freely fed mice, L-arginine (5 mM) stimulated insulin release was greatly enhanced and glucagon release markedly suppressed by the presence of the NOS inhibitor L-NAME in the incubation medium. These effects were abolished in isolated islets taken from 24 h fasted mice. 7. Our present results, which showed that the NOS inhibitor L-NAME markedly enhances insulin release but suppresses glucagon release induced by L-arginine in the intact animal, give strong support to our previous hypothesis that the islet NO system is a negative modulator of insulin secretion and a positive modulator of glucagon secretion. Additionally, we observed that the importance of the beta-cell NO-production for secretory mechanisms, as evaluated by the effect of L-NAME on L-arginine-induced insulin release, was greatly changed after starvation, an effect less prominent with regard to glucagon release.     

Facilitation of insulin release by N-p-tosylglycine

N-p-tosylglycine, which inhibits transglutaminase activity in islet homogenates, was found to cause a rapid and sustained facilitation of insulin release evoked by D-glucose, L-leucine or the association of Ba2+ and theophylline in intact islets. Such a facilitating action could not be attributed to any obvious effect upon either nutrient oxidation or 45Ca net uptake and outflow. It failed to be reproduced by glycine, N alpha-p-tosyl-L-arginine methyl ester or N alpha-p-tosyl-L-lysine methyl ester. N-p-tosylglycine (5.0 mM) slightly enhanced insulin release evoked by a high concentration of glucose (16.7 mM) and failed to affect significantly the secretory response to the association of L-leucine and L-glutamine or that of D-glucose and gliclazide. N-p-tosylglycine failed to affect the incorporation of [2,5-3H]histamine in trichloroacetic acid-precipitable material in intact islets. These results suggest that N-p-tosylglycine interferes with a late event in the secretory sequence, possibly at the level of the cell boundary, rather than inhibiting the crosslinking of intracellular proteins.     

Effect of a new xanthine derivative on the release of insulin from rat pancreatic islets

S 9795 (1-methyl-3-isobutyl-8-[2-ethyl 1-(4-diphenylmethylpiperazinyl)]- 3,7-dihydro(1H)purine-2,6-dione) is a new xanthine derivative displaying antiasthmatic properties in animals. The drug might exert its pharmacological actions either by inhibiting phosphodiesterases, or by inhibiting cellular Ca2+ movements or antagonizing purinoreceptors. Since the process of glucose-induced insulin release is markedly influenced by xanthine derivatives, the effect of S 9795 was compared to that of two other xanthine derivatives (theophylline and 3-isobutyl-1-methylxanthine (IBMX] on glucose-induced insulin release and ionic fluxes in rat pancreatic islets. Theophylline and IBMX potentiated glucose-induced insulin release, as expected, while S 9795 inhibited the insulinotropic effect of glucose. The effect of S 9795 was observed at a concentration of 10(-5) mol/l, lower concentrations (10(-6) to 10(-9) mol/l) failing to affect glucose-induced insulin release. At 10(-5) mol/l, the drug also inhibited the secondary rise in 45Ca efflux evoked by glucose from preloaded islets and the uptake of 45Ca by incubated islets stimulated either by glucose or potassium. The drug failed to alter 86Rb fluxes in stimulated and unstimulated islets labelled with the radioisotope. These data show that S 9795 inhibits glucose-induced insulin release, possibly by blocking glucose-stimulated Ca2+ inflow into the B-cell.     

Impairment of insulin release by methylation inhibitors

The possible participation of enzymatic methylation reactions in the process of insulin release was investigated in rat pancreatic islets. The combination of 3-deazaadenosine and dl-homocysteine impaired the incorporation of ³H-methyl from l-[methyl-³H]methionine into endogenous islet proteins and phospholipids, but failed to affect turnover in the phosphatidylinositol cycle. The inhibitors of methylation decreased insulin release evoked by d-glucose or the combinations of d-glucose and gliclazide, l-leucine and l-glutamine, or Ba²⁺ and theophylline. The inhibitors of methylation did not impair either the oxidation of d-glucose or affect its capacity to decrease K⁺ conductance, stimulate Ca²⁺ inflow and provoke ⁴⁵Ca accumulation in pancreatic islets. It is proposed that, in the process of insulin secretion, a methyl acceptor protein and/or phospholipid play(s) a limited modulatory role in the coupling of cytosolic Ca²⁺ accumulation to exocytosis.     

Influence of nitric oxide synthase inhibition, nitric oxide and hydroperoxide on insulin release induced by various secretagogues.

1. Recent studies have suggested that the generation of nitric oxide (NO) and hydrogen peroxide (H2O2) by islet NO synthase and monoamine oxidase, respectively, may have a regulatory influence on insulin secretory processes. We have investigated the pattern of insulin release from isolated islets of Langerhans in the presence of various pharmacological agents known to perturb the intracellular levels of NO and the oxidation state of SH-groups. 2. The NO synthase inhibitor, NG-nitro-L-arginine methyl ester (L-NAME) dose-dependently increased L-arginine-induced insulin release. D-Arginine did not influence L-arginine-induced insulin secretion. However, D-NAME which reportedly has no inhibitory action on NO synthase, modestly increased L-arginine-induced insulin release, but was less effective than L-NAME. High concentrations (10 mM) of D-arginine as well as L-NAME and D-NAME could enhance basal insulin release. 3. The intracellular NO donor, hydroxylamine, dose-dependently inhibited insulin secretion induced by L-arginine and L-arginine+L-NAME. 4. Glucose-induced insulin release was increased by NO synthase inhibition (L-NAME) and inhibited by the intracellular NO donor, hydroxylamine. Sydnonimine-1 (SIN-1), an extracellular donor of NO and superoxide, induced a modest suppression of glucose-stimulated insulin release. SIN-1 did not influence insulin secretion induced by L-arginine or the adenylate cyclase activator, forskolin. 5. The intracellular 'hydroperoxide donor' tert-butylhydroperoxide in the concentration range of 0.03-3 mM inhibited insulin release stimulated by the nutrient secretagogues glucose and L-arginine. Low concentrations (0.03-30 microM) of tert-butylhydroperoxide, however enhanced insulin secretion induced by the phosphodiesterase inhibitor isobutylmethylxanthine (IBMX).(ABSTRACT TRUNCATED AT 250 WORDS)     

In rat alveolar macrophages lipopolysaccharides exert divergent effects on the transport of the cationic amino acids l-arginine and l-ornithine

In rat alveolar macrophages (AMphi) it was tested whether induction of iNOS by lipopolysaccharides (LPS) is accompanied by changes in L-arginine transport and whether L-ornithine, the product of arginase released from AMphi, could, via inhibition of L-arginine uptake, act as a paracrine inhibitor of NO synthesis. Rat AMphi (cultured for 20 h in the absence or presence of 1 microg/ml LPS) were incubated in Krebs-HEPES solution containing [3H]-L-arginine (0.1 microM for 2 min or 100 microM for 5 min) and the cellular radioactivity was determined as a measure of L-arginine uptake. In parallel, cells were incubated for 6 h in Krebs-HEPES solution containing 0-1 mM L-arginine and nitrite accumulation was determined. [3H]-L-arginine uptake (0.1 microM or 100 microM) occurred independently of sodium ions and was inhibited by L-ornithine (EC50: 117 and 562 microM, respectively) and with similar potencies by L-lysine. In LPS-treated AMphi the concentration inhibition curve of L-ornithine was shifted to the right by about a factor of 4, whereas that of L-lysine was only marginally shifted to the right. L-Leucine (0.1 and 1 mM) inhibited [3H]-L-arginine (0.1 microM) by 43 and 58%, respectively, and the effect of 0.1 mM L-leucine was partially sodium dependent. In LPS-treated AMphi, 0.1 mM L-leucine no longer inhibited [3H]-L-arginine and the effect of 1 mM L-leucine was attenuated. Kinetic analysis of the transport of [3H]-L-arginine and [14C]-L-ornithine revealed two components for each amino acid with Km values of 21 and 114 microM (L-arginine) and 39 and 1050 microM (L-ornithine), respectively. After LPS treatment Km2 of L-arginine transport was reduced to 63 microM and Vmax of both components was increased, whereas Km2 of L-ornithine transport was enhanced to 1392 microM and Vmax1 reduced. LPS-stimulated AMphi, incubated in amino acid-free Krebs-HEPES solution, produced about 4 nmol nitrite/10(6) cells per 6 h, and L-arginine enhanced nitrite accumulation maximally about threefold (EC50: 30 microM). L-ornithine, up to 3 mM, failed to affect significantly nitrite accumulation observed in the presence of 30 or 100 microM L-arginine. Rat AMphi express mRNA for two cationic amino acid transporters (CAT-1 and CAT-2B), and LPS markedly up-regulated mRNA for CAT-2B in parallel with mRNA for iNOS, but had no effect on that for CAT-1. In conclusion, in rat AMphi LPS up-regulates L-arginine transport and induces changes in the characteristics of the cationic amino acid transport resulting in preferential transport of L-arginine. These effects may be regarded as cellular measures to ensure a high L-arginine supply for iNOS.     

Comparative effects of linogliride fumarate and tolbutamide sodium on insulin secretion and glucose utilization in rat pancreatic islets

The effects of linogliride fumarate (0.1 mmol/liter) and tolbutamide sodium (0.5 mmol/liter) on insulin secretion and glucose utilzation were directly compared in isolated perifused rat islets. Both compounds potentiated glucose-primed (5.5 mmol/liter) insulin release. The insulin secretory response at the concentration chosen was qualitatively different for the two agents. In the presence of glucose, the insulin secretory effect of tolbutamide was biphasic, whereas linoglirde predominantly increased second-phase insulin secretion. In the absence of exogenous glucos, tolbutamide stimulated first-phase insulin release wherease linogliride was ineffective without glucose in the perifusion medium. Neither linogliride (0.1 mmol/liter) nor tolbutamide (0.5 mmol/liter) stimulated islet cell glucose usage as measured by conversion of [5?³H]-glucose to ³H₂O. The insulin secretagogue effect of linogliride was completely abolished when islet cell glucose usage was partially blocked (22% reduction) by mannoheptulose (MH) (10 mmol/liter). When glucose usage was significantly inhibited (55%) by 2-deoxyglucose (2DG) (10 mmol/liter), linogliride-stimulated insulin secretion was not significantly reduced. In contrast, tolbutamide-stimulated first-phase insulin release was not inhibited when glucose usage was reduced by either MH or 2-DG, wherease secondphase insulin release was significantly inhibited. In summary, linogliride potentiates insulin secretion in isolated islets by a glucose-dependent process, but with a kinetic profile and response to metabolic inhibitors different from that of tolbutamide.     

Similar effects of succinic acid dimethyl ester and glucose on islet calcium oscillations and insulin release

The relative contribution of glycolysis vs. oxidative metabolism to the stimulus secretion coupling mechanism of beta-cells was investigated in isolated islets. For that purpose, the secretory and intracellular calcium responses of islets to both glucose and succinic acid dimethyl ester (SAD) were compared. After 45 min of rat islet perifusion in the absence of substrates, the maximum secretory responses to glucose (20 mmol/L) and SAD (10 mmol/L) were qualitatively and quantitatively indistinguishable. Malonic acid dimethyl ester (a permeable citric acid cycle inhibitor) suppressed the insulin secretory response to both 20 mmol/L glucose and 10 mmol/L SAD (-70% on average). The inhibitor decreased within 70% the rate of 14CO2-production from 10 mmol/L [2-(14)C]pyruvate without affecting the rate of 20 mmol/L D-[5-(3)H]glucose utilization. Both, 11.1 mmol/L glucose and 10 mmol/L SAD, elevated the intracellular calcium concentration and induced a similar pattern of oscillations that were rapidly ablated by 20 mmol/L malonic acid dimethyl ester. However, the intracellular concentration of calcium declined to basal values several minutes after the introduction of the inhibitor in the presence of SAD whereas it remained elevated in the case of glucose. In conclusion: (1) An exclusive increase of mitochondrial metabolism in pancreatic islets was sufficient to mimic the effects of glucose on intracellular calcium and insulin secretion. (2) Islet glycolysis and/or the re-oxidation of cytoplasmic NADH allowed the maintenance of an elevated, though non-oscillating, intracellular calcium concentration, but a reduced response to glucose.     

NO-dependent and -independent elevation of plasma levels of insulin and glucose in rats by L-arginine.

1. L-Arginine elevates plasma insulin in man. Recent in vitro data indicate that this is based on stimulation of endogenous nitric oxide (NO) with subsequent pancreatic release of insulin by L-arginine. L-Arginine also raises plasma glucose. 2. We studied plasma levels of insulin, glucose and NO metabolites, as well as systemic blood pressure, in anaesthetized rats during i.v. infusion of L-arginine (25-200 mg kg-1 min-1) or glucose (55 mg kg-1 min-1), before and after administration of the NO synthesis inhibitor, N omega-nitro-L-arginine methyl ester (L-NAME, 50 mg kg-1). 3. Before L-NAME, L-arginine elevated plasma insulin from about 15 to 65 ul-1 and glucose from 5.2 to 6.7 mmol l-1. These effects of L-arginine were not dose-related. 4. L-NAME alone had no effect on plasma insulin and glucose levels, but diminished the effects of a low dose (25 mg kg-1 min-1) of L-arginine on plasma insulin by about 40%, and that on plasma glucose by more than 90%. In contrast, the effects of a high dose (200 mg kg-1 min-1) of L-arginine on plasma insulin and glucose levels were not affected by L-NAME. 5. L-NAME elevated systemic blood pressure by about 35 mmHg. L-Arginine (25-100 mg kg-1 min-1) had no effect on systemic blood pressure, either before or after L-NAME. L-Arginine (200 mg kg-1 min-1) lowered systemic blood pressure, both before and after L-NAME. 6. Glucose infusion elevated plasma glucose from about 5.5 to 6.8 mmol l-1, and plasma insulin from about 18 to 26 ul-1. 7. The basal plasma levels of the NO metabolite nitrate (18 +/- 4 mumol l-1) were not affected by L-arginine (200 mg kg-1 min-1). Plasma nitrosohaemoglobin was likewise unaffected by L-arginine (200 mg kg-1 min-1). 8. We conclude that L-arginine separately elevates plasma insulin and glucose levels, both by NO-dependent and -independent mechanisms.     

DIURNAL VARIATIONS IN INSULIN SECRETION AND K+ PERMEABILITY IN ISOLATED RAT ISLETS

1. The effects of glucose on insulin secretion and 86Rb efflux from isolated rat islets were studied at six different times during a 24-h period (00.00, 04.00, 08.00, 12.00, 16.00 and 20.00 h). 2. In the absence of glucose and in the presence of substimulatory concentrations (2.8 mmol/L) of the sugar, insulin secretion did not vary with the time of day. At a glucose concentration of 5.6 mmol/L the stimulated insulin secretion was greater than basal levels only at 20.00 h. 3. At a higher sugar concentration (8.3 mmol/L) the increase in insulin secretion and the reduction in 86Rb efflux rate were more marked during the dark period. No effect of the time of day on insulin secretion was observed at glucose concentrations above 8.3 mmol/L (except in 27.7 mmol/L). 4. The time of day appears to affect insulin secretion mainly at glucose concentrations close to physiological values (5.6-8.3 mmol/L). 5. This result agrees with the ability of physiological amounts of glucose to alter the 86Rb-permeability of pancreatic B cells at the same time intervals.     

Effects of the new phosphodiesterase inhibitor griseolic acid on insulin release in rat pancreatic islets.

A new cyclic AMP phosphodiesterase inhibitor, griseolic acid (CAS 79030-08-3), in a dose-dependent manner increased insulin release and cyclic AMP level in rat pancreatic islets in the presence of 5.5 mmol/l glucose. Griseolic acid (0.26 mmol/l) or 3-isobutyl-1-methylxanthine (IBMX) (1 mmol/l) enhanced insulin release and cyclic AMP level both in the presence of 5.5 and 16.7 mmol/l glucose (no significant difference). In perifusion system, 45Ca++ efflux and insulin release showed a monophasic increase when the islets were exposed to 1.3 mmol/l griseolic acid or 1 mmol/l IBMX in the presence of 5.5 mmol/l glucose (no significant difference). In a cell-free system, griseolic acid had a stronger inhibitory effect on cyclic AMP phosphodiesterase activity than IBMX, has a stimulatory effect on insulin release through an increase of cyclic AMP by inhibiting phosphodiesterase in pancreatic islets, but it might not cross the plasma membrane easily.     

Salivary peptide P-C modulates both insulin and glucagon release from isolated perfused rat pancreas

The effect of salivary peptide P-C, saliva-derived peptide on glucose-induced insulin release was studied using perfused rat pancreas. Salivary peptide P-C (194 nM) remarkably potentiated glucose (8.3 and 16.7 mM)-induced insulin release, whereas the same concentration suppressed arginine (10 mM)-induced glucagon release. Both effects of salivary peptide P-C occurred in a concentration-dependent manner. These findings suggest that salivary peptide P-C may modulate both the levels of insulin and glucagon in vivo.     

Effect of different stimulators and a blocker of insulin release on islet Na+, K+-ATPase activity

The effect of several insulin secretagogues and a blocker upon islet Na+, K+-ATPase activity was studied using rat islet homogenates. None of the agents tested modified the enzyme activity when added directly to the enzyme assay. Activity of Na+, K+-ATPase measured in islets preincubated during 3 min with glucose 3.3, 8 or 16.6 mM, as well as with 15 mM KIC or 1.2 microM somatostatin, did not significantly change. The presence of glucagon (1.4 microM) plus theophylline (10 mM) in the preincubation medium significantly enhanced activity while tolbutamide (1.48 mM) or gliclazide (76 microM) significantly decreased such activity. These results suggest that Na+, K+-ATPase activity would not be a main common step involved in the mechanism by which glucose, KIC, glucagon + theophylline and somatostatin exert their effect on insulin secretion. Conversely, the enzyme might contribute to the stimulatory effect of gliclazide and tolbutamide on insulin release. Such effect would be secondary to the release of some cellular mediator rather than a direct action of these compounds on the enzyme. Such effect would later favor a rise in the cytosolic concentration of calcium which might trigger the release of insulin.     

二甲双胍在慢性高胰岛素处理的Hep G2细胞对胰岛素受体后信号转导的调节作用(英文)

目的:研究慢性高胰岛素对胰岛素受体后信号转导的影响及其与胰岛素抵抗的关系,并探讨二甲双胍治疗胰岛素抵抗是否通过胰岛素信号转导途径的介导。方法:人类肝癌细胞系(Hep G2)在无血清条件下首先与100nmol/L高浓度胰岛素预温育16h以产生胰岛素抵抗细胞模型,并观察不同浓度(0.01-10mmol/L)的二甲双胍对胰岛素受体后磷酯酰肌醇3激酶(PI3K)途径信号转导的影响。结果:高浓度胰岛素100nmol/L慢性处理引起了IRβ、IRS1和IRS2的酪氨酸磷酸化和蛋白表达水平的下调,p85与IRS的相互作用也有显著降低。生理治疗浓度的(0.01-0.1mmol/L)二甲双胍逆转了这种慢性高胰岛素引起的信号下调,细胞用0.1mmol/L二甲双胍预温育后,IRB、IRS1和IRS2的磷酸化反应水平分别增加了2.7倍(P<0.01),6.8倍(P<0.01)和2.3倍(P<0.01),p85与IRS1的相互作用从34％增加至86％(P<0.01),与IRS2的相互作用从30％增加至92％(P<0.01)。相反,药物浓度(1-10mmol/L)的二甲双胍进一步抑制了IRB、IRS的磷酸化及IRS与p85的相互作用。结论:高浓度胰岛素慢性处理可引起胰岛素受体后PI3K途径信号转导的下调,二甲双胍对胰岛素信号转导的作用可能是其治疗胰岛素抵抗的主要分子机制。     

Effects of isoprenaline and glucagon on insulin secretion from pancreatic islets

Summary The effects of isoprenaline and glucagon on insulin secretion from pancreatic islets were investigated. In the presence of high concentrations of isoprenaline (10–50 mol/l), glucose-induced (20 mmol/l) insulin secretion from isolated perifused mouse islets was inhibited. This inhibition was apparently mediated by ₂-adrenoceptors, as it was antagonized by rauwolscine. At low concentrations isoprenaline (0.1 or 1 mol/l) did not affect glucose-induced (2.5; 10 or 20 mmol/l) insulin secretion from perifused mouse or rat islets, even if ₂-adrenoceptors were blocked by rauwolscine. A stimulatory effect of isoprenaline on insulin secretion was also not observed in the perfused rat pancreas. However, when incubated mouse islets were exposed to glucose (10 mmol/l), insulin secretion was further enhanced by isoprenaline (0.5 mol/l). To elucidate the underlying mechanism, the effects of glucagon on insulin secretion were investigated, because glucagon is released from the pancreatic A-cells during stimulation with isoprenaline and is accumulated in the islets and the surrounding medium during incubations of pancreatic islets. Indeed, glucagon stimulated insulin secretion from perifused mouse islets in the presence of high glucose (10 or 15 mmol/l) concentrations but not of low glucose (5 mmol/l) concentrations. Thus it is concluded that direct -adrenergic stimulation of pancreatic B-cells does not occur in mouse or rat pancreatic islets. Augmentation of glucose-induced insulin secretion by isoprenaline observed in incubation systems can be explained as a result of stimulation by glucagon, which is released from pancreatic A-cells by isoprenaline.Some of the results described here were obtained during medical thesis work by S. Zielmann and g. Schütte     

Effects of cationic modification on 45Ca uptake and insulin release by transplantable rat insulinoma cells maintained in tissue culture

1. Acute effects of cations on 45Ca uptake and insulin release by transplantable rat insulinoma cells were examined after 2-3 days culture in RPMI-1640 containing 11.1 mM glucose. 2. At 2.6 mM Ca2+, rat insulinoma cells (greater than 95% viability) released 78-158 ng insulin/10(6) cells during 60 min incubation with uptake at 2.19-3.24 nmol 45Ca/10(6) cells. 3. Addition of 2 mM La3+, Co2+, Mn2+, Zn2+ or Ba2+ did not affect 45Ca uptake. Insulin release was also unaffected by these cations with the exception of 87% inhibition in the presence of La3+ or Zn2+. 4. Omission of 5.9 mM K+, 1.2 mM Mg2+, 115 mM Na+ or H+ (pH 8.5) did not affect 45Ca uptake or insulin release, irrespective of osmotic compensation using choline chloride or sucrose. Rat insulinoma cells were similarly unresponsive to addition of 30.9 mM K+, 12 mM Mg2+ or H+ (pH 6.3). 5. Omission of 2.6 mM Ca2+ (with or without addition of 1 mM EGTA) or addition of 20.5 mM Ca2+ did not affect insulin release. 6. The results indicate that rat insulinoma cells are little affected by cationic modifications which have profound effects on Ca2+ handling and insulin release by pancreatic beta-cells. Dysregulation of insulin release by insulinoma cells is associated with marked irregularities in the control of transmembrane Ca2+ fluxes and sensitivity to extracellular Ca2+.     

Effect of a non-sulphonylurea hypoglycaemic agent, KAD-1229 on hormone secretion in the isolated perfused pancreas of the rat.

1. We examined the cooperative effect of a newly synthesized oral hypoglycaemic agent, KAD-1229 with glucose on insulin, glucagon and somatostatin secretion in the isolated perfused pancreas of the rat. 2. KAD-1229 stimulated concentration-dependently the first phase of insulin secretion without the second phase in the presence of 2.8 mM glucose, while it stimulated both the first and the second phase of insulin release in the presence of 5.6 mM glucose. It was confirmed that the first phase of insulin release is depolarization-induced release with no other additional signal transduction. 3. KAD-1229 also enhanced insulin release evoked by 16.7 mM glucose, a concentration known to inhibit the ATP-sensitive K+ current completely. 4. A low concentration (2.8 mM) of glucose stimulated somatostatin release transiently, while a higher concentration (16.7 mM) of glucose exerted a sustained stimulation. KAD-1229 stimulated somatostatin secretion in a concentration-dependent manner irrespective of glucose concentrations. 5. When glucagon release was stimulated with 2.8 mM glucose, KAD-1229 inhibited this hypoglycaemia-induced glucagon secretion. 6. When pancreata from rats pretreated with streptozotocin (STZ) 60 mg kg-1 were perfused, the basal secretion of glucagon was markedly elevated, and the glucagon response to the low glucose was abolished. Further, the insulin and somatostatin responses to KAD-1229 were largely attenuated. KAD-1229 showed transient enhancement followed by inhibition of the glucagon release from the STZ-pretreated rat pancreas. 7. We conclude that KAD-1229 stimulates insulin and somatostatin release, while it inhibits glucagon release following transient stimulation.