Biochimica et Biophysica Acta (BBA) - General Subjects
Calcium and pancreatic β-cell function 7. Evidence for cyclic AMP-induced translocation of intracellular calcium
Abstract
The effect of cyclic AMP on calcium movements in the pancreatic β-cell was evaluated using an experimental approach based on in situ labelling of intracellular organelles of ob/ob-mouse islets with 45Ca. Whereas the glucose-stimulated 45Ca incorporation by mitochondria and secretory granules was increased under a condition known to reduce cyclic AMP (starvation), raised levels of this nucleotide (addition of 3-isobutyl-1-methylxanthine or N6,O2′-dibutyryl adenosine 3′,5′-cyclic monophosphate) reduced the mitochondrial accumulation of 45Ca. Conditions with increased cyclic AMP were associated with a stimulated efflux of 45Ca from the secretory granules but not from the mitochondria. The microsomal fraction differed from both the mitochondrial and secretory granule fractions by accumulating more 45Ca after the addition of 3-isobutyl-1-methylxanthine. The results suggest that cyclic AMP potentiates glucose-stimulated insulin release by increasing cytoplasmic Ca2+ at the expense of the calcium taken up by the organelles of the pancreatic β-cells.
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Cited by (21)
Insulinotropic glucagon-like peptide-1-mediated activation of non- selective cation currents in insulinoma cells is mimicked by maitotoxin
1997, Journal of Biological ChemistryMaitotoxin (MTX) activates a Ca2+-dependent non-selective cation current (ICa-NS) in insulinoma cells whose time course is identical to non-selective cation currents activated by incretin hormones such as glucagon-like peptide-1 (GLP-1), which stimulate glucose-dependent insulin secretion by activating cAMP signaling pathways. We investigated the mechanism of activation of ICa-NS in insulinoma cells using specific pharmacological reagents, and these studies further support an identity between MTX- and GLP-1-activated currents. ICa-NS is inhibited by extracellular application of genistein, econazole, and SKF 96365. This inhibition by genistein suggests that tyrosine phophorylation may play a role in the activation of ICa-NS. ICa-NS is not inhibited by incubation of cells in glucose-free solution, by extracellular tetrodotoxin, nimodipine, or tetraethylammonium, or by intracellular dialysis with 4-aminopyridine, ATP, ryanodine, or heparin. ICa-NS is also not significantly inhibited by staurosporine, which does, however, partially inhibit the MTX-induced rise of intracellular Ca2+ concentration. These effects of staurosporine suggest that protein kinase C may not be involved in the activation of ICa-NS but that it may regulate intracellular Ca2+ release. Alternatively, ICa-NS may have a small component that is carried through separate divalent cation-selective channels that are inhibited by staurosporine. ICa-NS is neither activated nor inhibited by dialysis with KF, KF + AlF3 or GTPγS (guanosine 5′-O-(3-thiotriphosphate)), suggesting that GTP-binding proteins do not play a major role in the activation of this current.
Activation of a cAMP-regulated Ca<sup>2+</sup>-signaling pathway in pancreatic β-cells by the insulinotropic hormone glucagon-like peptide-1
1995, Journal of Biological ChemistryGlucagon-like peptide-1 (GLP-1) is an intestinally derived insulinotropic hormone that is currently under investigation for use in the treatment of diabetes mellitus. To investigate the Ca2+ signaling pathways by which GLP-1 may stimulate the secretion of insulin from pancreatic β-cells, we examined its effects on the concentration of free intracellular Ca2+ ([Ca2+]i) while simultaneously determining what action it exerts on ion channel function. Measurements of [Ca2+]i were obtained from single rat β-cells and from βTC6 and HIT-T15 insulinoma cells loaded with the Ca2+ indicator fura-2, and changes in membrane potential and current were monitored using the perforated patch clamp technique. We report a previously undocumented action of GLP-1 and analogs of cAMP (8-bromo-cAMP, Sp- or Rp-adenosine 3′,5′-cyclic monophosphothionate triethylamine) to raise [Ca2+]i that is attributable to the activation of a prolonged inward current designated here as IcAMP. Activation of IcAMP is associated with an increased membrane conductance, membrane depolarization, and triggers large increases of [Ca2+]i. IcAMP is primarily a Na+ current that is blocked by extracellularly applied La3+ or by intracellular administration of Ca2+ chelators (1,2-bis(2aminophenoxy)ethane-N,N,N′,N′-tetraacetic acid/acetoxymethyl, EGTA) and which exhibits a reversal potential of about −26 mV. We propose that IcAMP results from the opening of nonselective cation channels that are activated by intracellular Ca2+ and cAMP and which might play an important role in the regulation of insulin secretion from pancreatic β-cells.
Cyclic AMP raises cytoplasmic calcium in pancreatic α<inf>2</inf>-cells by mobilizing calcium incorporated in response to glucose
1989, Cell CalciumThe cytoplasmic Ca2+ concentration ([Ca2+]i) was monitored in individual guinea-pig pancreatic α2-cells exposed to modulators of glucagon release. Addition of the stimulatory amino acid arginine resulted in a sustained increase in [Ca2+]i, whereas the inhibitor glucose had the opposite effect. Epinephrine, the β-adrenergic agonist isoproterenol, the adenylate cyclase activator forskolin and 8-bromo-cAMP transiently raised [Ca2+]i provided that the cells had been pretreated with glucose. However, simultaneous presence of glucose was not required and the effect occurred even in the absence of extracellular Ca2+. Carbachol, the α2-adrenergic agonist clonidine and the sulfonylurea tolbutamide lacked effects on [Ca2+]i. In addition to providing support for the concept that glucagon release is positively modulated by [Ca2+]i, the results demonstrate that cAMP raises [Ca2+]i in the α2-cells by mobilizing calcium incorporated in response to glucose.
The interrelationship between mediators of hormone action (cAMP and calcium) and induction of ornithine decarboxylase (ODC) activity was investigated in bone and bone cells. Stimuli that enhanced the cAMP level in both osteoblastlike (BL) and osteoclastlike (CL) cells and in intact calvaria only induced ODC activity in the isolated cell populations. Moreover, addition of fresh medium induced ODC activity in BL cells and not in calvaria, without an effect on the cAMP level. The tumor-promoting agent 12-O-tetradecanoyl-phorbol-13-acetate (TPA), however, induced in both systems ODC activity, without an effect on the cAMP concentration. From these data it was concluded that induction of ODC activity may not be mediated by cAMP. The role of Ca in ODC induction was also investigated. Experiments with BL cells, incubated in media with various Ca concentrations, as well as experiments with the Ca blocker D600 showed that basal and hormone-induced ODC activity was dependent on the intracellular Ca concentration. A regulatory role of cAMP, however, in concert with Ca, cannot be excluded.
La relation entre les médiateurs de l'action des hormones (AMP cyclique et Ca) et l'induction de l'activité de l'omithine decarboxylase (ODC) a étéétudiée dans l'os et les cellules osseuses. Les stimull qui accroissent le taux de l'AMPc dans les cellules ostéobiastiques (OBL) et les cellules ostéoclastiques (OCL) dans des calvaria intacts n'Induisant une activité de l'ODC que dans les populations cellulaires Isolées. De plus, l'addition de milieu frais qui Induit une activité de l'ODC dans les cellules OBL mais pas dans les calvaria reste sans effet sur le taux d'AMPc. Le 12-O tetradecanoyl phorbol-13-acetate (TPA) qui est un agent carcinogénique induit cependant dans les deux systèmes l'activité de l'ODC, mais est sans effet sur la concentration en AMPc. Ces données amènent è conclure que l'Induction de l'activité ODC peut ne pas être médlée par l'AMPc. Pour évaluer l'effet d'autres médiateurs de l'action hormonale que l'AMPc, le rôle du Ca dans l'Induction de l'ODC a été aussi étudié. Des essais sur des cellules OBL, incubées dans des milieux ayant différantes concentrations en Ca, tout comme des études avec te calcium-bloquant D 600, ont montré que l'activité ODC de base et celte induite par tes hormones dépendent de Is concentration intracellulaire en calcium. Un rôle régulateur de l'AMPc, s'exerçant de concert avec te Ca, ne peut cependant pas être exclu.
Opposing actions of methylxanthines and dibutyryl cyclic AMP on 1,25 dihydroxyvitamin D<inf>3</inf> production and calcium fluxes in isolated chick renal tubules
1984, Biochemical and Biophysical Research CommunicationsIn contrast to dibuturyl cyclic AMP, the methylxanthine phosphodiesterase inhibitors theophylline and caffeine were found to inhibit the conversion of 25 hydroxyvitamin D3 to 1,25 dihydroxyvitamin D3 in isolated renal tubules from vitamin D deficient chicks. This inhibition occurred at concentrations of methylxanthines which were shown to increase renal tubule cyclic AMP levels. No effect of theophylline or caffeine on 25 hydroxyvitamin D3 metabolism in isolated chick renal mitochondria was detected. Because of a demonstrated inhibitory action of calcium (10 and 20 μmol/l) on renal mitochondrial conversion of 25 hydroxyvitamin D3 to 1,25 dihydroxyvitamin D3, the effect of theophylline and dibutyryl cyclic AMP on cellular calcium-45 efflux and total renal tubule calcium content was estimated. Theophylline 10 mmol/l was found to inhibit renal tubular calcium efflux and to increase total cellular calcium content, while dibutyryl cyclic AMP 1 mmol/l had the reverse effect on both parameters. Divergent actions of the methylxanthines and dibutyryl cyclic AMP on the formation of 1,25 dihydroxyvitamin D3 and renal tubule calcium efflux and content support the hypothesis that intracellular calcium is an important regulator of renal vitamin D metabolism. The results indicate that observed actions of methylxanthines cannot always be ascribed to cyclic AMP accumulation.
Glucose inhibits <sup>45</sup>Ca efflux from pancreatic β-cells also in the absence of Na<sup>+</sup>-Ca<sup>2+</sup> countertransport
1984, BBA - BiomembranesDuring perifusion with medium deprived of Ca2+, addition of glucose or omission of Na+ resulted in prompt and quantitatively similar inhibitions of 45Ca efflux from β-cell rich pancreatic islets microdissected from ob / ob mice. Glucose had no additional inhibitory effect when Na+ was isoosmotically replaced by sucrose or choline+. When K+ was used as a substitute for Na+, the inhibitory effect of Na+ removal on 45Ca efflux became additive to that of glucose. The observation that glucose can be equally effective in inhibiting 45Ca efflux in the presence or absence of Na+ is difficult to reconcile with the postulate that the Na+-Ca2+ countertransport mechanism is a primary site of action for glucose.