EFFECT OF FASTING ON 32P TRANSLOCATIONS IN PRE-LABELLED PANCREATIC ISLETS

ABSTRACT The rapid, short-lived efflux of inorganic 32P-orthophosphate that occurs when pre-labelled pancreatic islets are exposed to nutrient insulin secretagogues (the "phosphate flush") has been proposed to reflect some early step in β-cell secretory activation. In the present study, glucose-initiated phosphate efflux was studied during fasting. Pancreatic islets were isolated from fed and 48-h fasted rats by collagenase digestion. After pre-labelling with 32P-orthophosphate and basal perifusion with 0.5 mg/ml glucose, tissue analyses disclosed similar stores of radioactivity in the two groups of islets. Stimulatory perifusion with glucose at this time failed to promote insulin release from islets which had been secured from fasted donors although the "phosphate flush" was preserved. However, the characteristics of phosphate efflux were altered. Maximal glucose-induced phosphate release was greater with islets from fasted animals whereas phosphate release in response to low level stimulation with glucose was diminished. Accordingly, the dose-response curve for glucose-initiated phosphate efflux in islets from fasted rats was displaced to the right and compatible with a decreased sensitivity to glucose at the activation site for the "phosphate flush." Thus, while glucose is unable to enhance insulin release in vitro after fasting, glucose still elicits increased phosphate efflux. However, the phenomenon appears to be attended by an impaired responsiveness to activation by glucose, supporting the contention that some early step in the sequence of stimulus secretion coupling in the β-cell may be obtunded after food deprivation.

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Sustained exposure of toxically damaged mouse pancreatic islets to high glucose does not increase β-cell dysfunction

Journal of Endocrinology ◽

10.1677/joe.0.1230047 ◽

1989 ◽

Vol 123 (1) ◽

pp. 47-51 ◽

Cited By ~ 8

Author(s):

D. L. Eizirik ◽

S. Sandler

Keyword(s):

Pancreatic Islets ◽

Insulin Release ◽

High Glucose ◽

Β Cells ◽

Β Cell ◽

Glucose Stimulation ◽

Cell Dysfunction ◽

Mouse Pancreatic Islets ◽

Insulin Secretory Response

ABSTRACT The aim of this study was to clarify whether prolonged in-vitro exposure of either normal or damaged β cells to a high glucose environment can be toxic to these cells. For this purpose NMRI mice were injected intravenously with a diabetogenic dose of streptozotocin (SZ; 160 mg/kg) or vehicle alone (controls). Their islets were isolated 15 min after the injection and subsequently maintained in culture for 21 days in the presence of 11·1 or 28 mmol glucose/l. After this period, during acute glucose stimulation, the control islets showed a marked increase in their insulin release in response to a high glucose stimulus. In the SZ-exposed islets there was a decrease in DNA and insulin contents, and a deficient insulin secretory response to glucose. However, in the SZ-damaged islets as well as in the control islets, culture with 28 mmol glucose/l compared with 11·1 mmol glucose/l did not impair islet retrieval after culture, islet DNA content or glucose-induced insulin release. Thus, the degree of damage was similar in the SZ-treated islets cultured at the two concentrations of glucose. These results suggest that glucose is not toxic to normal or damaged mouse pancreatic islets over a prolonged period in tissue culture. Journal of Endocrinology (1989) 123, 47–51

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A role for polyamines in stimulus-secretion coupling in the pancreatic β-cell

Bioscience Reports ◽

10.1007/bf01138169 ◽

1984 ◽

Vol 4 (10) ◽

pp. 869-877 ◽

Cited By ~ 16

Author(s):

P. J. Bungay ◽

J. M. Potter ◽

M. Griffin

Keyword(s):

Pancreatic Islets ◽

Insulin Release ◽

Intracellular Concentration ◽

Secretory Process ◽

Pancreatic Β Cell ◽

Β Cell ◽

Rapid Synthesis ◽

Stimulus Secretion Coupling ◽

Significant Change

Measurement of the content of polyamines in pancreatic islets indicated that no significant change in their concentration took place during glucose-stimulated insulin release. The finding, together with the absence of any effect of α-difluoromethylornithine on glucosestimulated insulin release suggested that rapid synthesis of polyamines is not involved in stimulus-secretion coupling in the β-cell. The concentration of polyamines found in islets were high enough for them to act as substrates for the Ca2+-dependent islet transglutaminase during insulin release. This was further demonstrated by the ability of islet transglutaminase to incorporate [14C]putrescine into proteins from islet homogenates and by the demonstration of an increase in the covalent incorporation of [14C]putrescine into the proteins of intact islets following their challenge with glucose. Unlike monoamine substrates of transglutaminase, putrescine failed to effectively inhibit insulin release when its intracellular concentration was increased. A role for polyamines in the secretory process through their incorporation into islet proteins is suggested.

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Direct effect of parathyroid hormone on insulin secretion from pancreatic islets

AJP Endocrinology and Metabolism ◽

10.1152/ajpendo.1990.258.6.e975 ◽

1990 ◽

Vol 258 (6) ◽

pp. E975-E984 ◽

Cited By ~ 5

Author(s):

G. Z. Fadda ◽

M. Akmal ◽

L. G. Lipson ◽

S. G. Massry

Keyword(s):

Insulin Secretion ◽

Parathyroid Hormone ◽

Pancreatic Islets ◽

Insulin Release ◽

Direct Effect ◽

Indirect Evidence ◽

Cytosolic Calcium ◽

Dependent Manner ◽

Stimulation Of

Indirect evidence indicates that parathyroid hormone (PTH) interacts with pancreatic islets and modulates their insulin secretion. This property of PTH has been implicated in the genesis of impaired insulin release in chronic renal failure. We examined the direct effect of PTH-(1-84) and PTH-(1-34) on insulin release using in vitro static incubation and dynamic perifusion of pancreatic islets from normal rats. Both moieties of the hormone stimulated in a dose-dependent manner glucose-induced insulin release but higher doses inhibited glucose-induced insulin release. This action of PTH was modulated by the calcium concentration in the media. The stimulatory effect of PTH was abolished by its inactivation and blocked by its antagonist [Tyr-34]bPTH-(7-34)NH2. PTH also augmented phorbol ester (TPA)-induced insulin release, stimulated adenosine 3',5'-cyclic monophosphate (cAMP) generation by pancreatic islets, and significantly increased (+50 +/- 2.7%, P less than 0.01) their cytosolic calcium. Verapamil inhibited the stimulatory effect of PTH on insulin release. The data show that 1) pancreatic islets are a PTH target and may have PTH receptors, 2) stimulation of glucose-induced insulin release by PTH is mediated by a rise in cytosolic calcium, 3) stimulation of cAMP production by PTH and a potential indirect activation of protein kinase C by PTH may also contribute to the stimulatory effect on glucose-induced insulin release, and 4) this action of PTH requires calcium in incubation or perifusion media.

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Nitric oxide donor SIN-1 inhibits insulin release

AJP Cell Physiology ◽

10.1152/ajpcell.1996.271.4.c1098 ◽

1996 ◽

Vol 271 (4) ◽

pp. C1098-C1102 ◽

Cited By ~ 29

Author(s):

A. Sjoholm

Keyword(s):

Nitric Oxide ◽

Diabetes Mellitus ◽

Protein Kinase C ◽

Insulin Secretion ◽

Protein Kinase ◽

Beta Cell ◽

Pancreatic Islets ◽

Insulin Release ◽

Kinase C ◽

Stimulus Secretion Coupling

Preceding the onset of insulin-dependent diabetes mellitus, pancreatic islets are infiltrated by macrophages secreting interleukin-1 beta, which exerts cytotoxic and inhibitory actions on islet beta-cell insulin secretion through induction of nitric oxide (NO) synthesis. The influence of the NO donor 3-morpholinosydnonimine (SIN-1) on insulin secretion from isolated pancreatic islets in response to various secretagogues was investigated. Stimulation of insulin release evoked by glucose, phospholipase C activation with carbachol, and protein kinase C activation with phorbol ester were obtained by SIN-1, whereas the response to adenylyl cyclase activation or K(+)-induced depolarization was not affected. It is concluded that enzymes involved in glucose catabolism, phospholipase C or protein kinase C, may be targeted by NO. Reversal of SIN-1 inhibition of glucose-stimulated insulin release by dithiothreitol suggests that NO may inhibit insulin secretion partly by S-nitrosylation of thiol residues in key proteins in the stimulus-secretion coupling. These adverse effects of NO on the beta-cell stimulus-secretion coupling may be of importance for the development of the impaired insulin secretion characterizing diabetes mellitus.

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Effect of perchlorate on calcium uptake and insulin secretion in mouse pancreatic islets

Biochemical Journal ◽

10.1042/bj2480109 ◽

1987 ◽

Vol 248 (1) ◽

pp. 109-115 ◽

Cited By ~ 5

Author(s):

J Sehlin

Keyword(s):

Insulin Secretion ◽

Beta Cell ◽

Pancreatic Islets ◽

Insulin Release ◽

Glucose Oxidation ◽

Calcium Uptake ◽

Maximum Effect ◽

Mouse Pancreatic Islets ◽

Stimulus Secretion Coupling ◽

Ca2 Channels

Microdissected beta-cell-rich pancreatic islets of non-inbred ob/ob mice were used in studies of how perchlorate (CIO4-) affects stimulus-secretion coupling in beta-cells. CIO4- at 16 mM potentiated D-glucose-induced insulin release, without inducing secretion at non-stimulatory glucose concentrations. The potentiation mainly applied to the first phase of stimulated insulin release. In the presence of 20 mM-glucose, the half-maximum effect of CIO4- was reached at 5.5 mM and maximum effect at 12 mM of the anion. The potentiation was reversible and inhibitable by D-mannoheptulose (20 mM) or Ca2+ deficiency. CIO4- at 1-8 mM did not affect glucose oxidation. The effects on secretion were paralleled by a potentiation of glucose-induced 45Ca2+ influx during 3 min. K+-induced insulin secretion and 45Ca2+ uptake were potentiated by 8-16 mM-CIO4-. The spontaneous inactivation of K+-induced (20.9 mM-K+) insulin release was delayed by 8 mM-CIO4-. The anion potentiated the 45Ca2+ uptake induced by glibenclamide, which is known to depolarize the beta-cell. Insulin release was not affected by 1-10 mM-trichloroacetate. It is suggested that CIO4- stimulates the beta-cell by affecting the gating of voltage-controlled Ca2+ channels.

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Interleukin 10 inhibits insulin release from and nitric oxide production in rat pancreatic islets

Acta Endocrinologica ◽

10.1530/eje.0.1350374 ◽

1996 ◽

Vol 135 (3) ◽

pp. 374-378 ◽

Cited By ~ 6

Author(s):

Renato Laffranchi ◽

Giatgen A Spinas

Keyword(s):

Nitric Oxide ◽

Pancreatic Islets ◽

Insulin Release ◽

Interleukin 10 ◽

Interleukin 1Β ◽

Nitric Oxide Production ◽

Β Cell ◽

Cell Dysfunction ◽

Rat Pancreatic Islets ◽

Oxide Production

Laffranchi R, Spinas GA. Interleukin 10 inhibits insulin release from and nitric oxide production in rat pancreatic islets. Eur J Endocrinol 1996;135:374–8. ISSN 0804–4643 Interleukin 10 was found to prevent cytokine-induced nitric oxide production in murine macrophages. Because, in rat islets, interleukin 1β induces β-cell dysfunction, mainly due to overproduction of nitric oxide, we studied if this effect could be counteracted by interleukin 10. Rat pancreatic islets were cultured for 24 h in the presence or absence of 0.02–20 ng/ml recombinant human interleukin 10. Interleukin 10 dose-dependently inhibited insulin secretion with maximal inhibition (27 ±4%, p < 0.05) at 2 ng/ml without impairment of islet cell viability. However, incubation of pancreatic islets with interleukin 10 resulted in a 61.5% decrease of nitric oxide production. Co-incubation of islets with interleukin 10 (2 ng/ml) and recombinant human interleukin 1β (0.15 ng/ml) resulted in a more pronounced suppression of basal insulin release than with interleukin 1β alone (55 ± 3.6% vs 44 ± 3.6% with interleukin 1β alone, p < 0.05) but did not reduce interleukin 1β-stimulated NO production or reverse the effect of interleukin 1β on cell viability. Thus, in pancreatic islets interleukin 10 is not capable of counteracting the interleukin 1β-induced β-cell dysfunction, but rather enhances the inhibitory effect of interleukin 1β by a different mechanism. Renato Laffranchi, Division of Endocrinology and Metabolism, Department of Internal Medicine, University Hospital, Rämistrasse 100, CH-8091 Zürich, Switzerland

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The Stimulus-Secretion Coupling of Amino Acid-induced Insulin Release: Secretory and Oxidative Response of Pancreatic Islets to L-Asparagine

Diabetes ◽

10.2337/diab.33.5.464 ◽

1984 ◽

Vol 33 (5) ◽

pp. 464-469 ◽

Cited By ~ 7

Author(s):

F. Malaisse-Lagae ◽

M. Welsh ◽

P. Lebrun ◽

A. Herchuelz ◽

A. Sener ◽

...

Keyword(s):

Amino Acid ◽

Pancreatic Islets ◽

Insulin Release ◽

Oxidative Response ◽

Stimulus Secretion Coupling

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Evidence for the participation of calmodulin in stimulus–secretion coupling in the pancreatic β-cell

Biochemical Journal ◽

10.1042/bj1920919 ◽

1980 ◽

Vol 192 (3) ◽

pp. 919-927 ◽

Cited By ~ 56

Author(s):

Juan J. Gagliardino ◽

Donna E. Harrison ◽

Michael R. Christie ◽

Elma E. Gagliardino ◽

Stephen J. H. Ashcroft

Keyword(s):

Insulin Release ◽

Kinase Inhibitor ◽

Potent Inhibitor ◽

Polyacrylamide Gel Electrophoresis ◽

Sodium Dodecyl ◽

Β Cell ◽

Batch Type ◽

Dependent Inhibition ◽

Stimulus Secretion Coupling ◽

Rat Islets Of Langerhans

1. The ability of a range of phenothiazines to inhibit activation of brain phosphodiesterase by purified calmodulin was studied. Trifluoperazine, prochlorperazine and 8-hydroxyprochlorperazine produced equipotent dose-dependent inhibition with half-maximum inhibition at 12μm. When tested at 10 or 50μm, 7-hydroxyprochlorperazine was a similarly potent inhibitor. However, trifluoperazine-5-oxide and N-methyl-2-(trifluoromethyl)phenothiazine were ineffective at concentrations up to 50μm, and produced only a modest inhibition at 100μm. 2. The same phenothiazines were tested for their ability to inhibit activation of brain phosphodiesterase by boiled extracts of rat islets of Langerhans. At a concentration of 20μm, 70–80% inhibition was observed with trifluoperazine, prochlorperazine, 7-hydroxyprochlorperazine or 8-hydroxyprochlorperazine, whereas trifluoperazine-5-oxide and N-methyl-2-(trifluoromethyl)phenothiazine were less effective. 3. The effect of these phenothiazines on insulin release from pancreatic islets was studied in batch-type incubations. Insulin release stimulated by glucose (20mm) was markedly inhibited by 10μm-trifluoperazine or -prochlorperazine and further inhibited at a concentration of 20μm. 8-Hydroxyprochlorperazine (20μm) was also a potent inhibitor but 7-hydroxyprochlorperazine (20μm) elicited only a modest inhibition of glucose-stimulated insulin release; no inhibition was observed with trifluoperazine-5-oxide or N-methyl-2-(trifluoromethyl)phenothiazine. 4. Trifluoperazine (20μm) markedly inhibited insulin release stimulated by leucine or 4-methyl-2-oxopentanoate in the absence of glucose, and both trifluoperazine and prochlorperazine (20μm) decreased insulin release stimulated by glibenclamide in the presence of 3.3mm-glucose. 5. None of the phenothiazines affected basal insulin release in the presence of 2mm-glucose. 6. Trifluoperazine (20μm) did not inhibit islet glucose utilization nor the incorporation of [3H]leucine into (pro)insulin or total islet protein. 7. Islet extracts catalysed the incorporation of 32P from [γ-32P]ATP into endogenous protein substrates. Sodium dodecyl sulphate/polyacrylamide-gel electrophoresis resolved several phosphorylated bands, but incorporation was slight. However, calmodulin in the presence of Ca2+ greatly enhanced incorporation: the predominant phosphorylated band had an estimated mol.wt. of 55000. This enhanced incorporation was abolished by trifluoperazine, but not by cyclic AMP-dependent protein kinase inhibitor protein. 8. These results suggest that islet phosphodiesterase-stimulating activity is similar to, although not necessarily identical with, calmodulin from skeletal muscle; that islet calmodulin may play an important role in Ca2+-dependent stimulus–secretion coupling in the β-cell; and that calmodulin may exert part at least of its effect on secretion via phosphorylation of endogenous islet proteins.

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