Novel regulation by Rac1 of glucose- and forskolin-induced insulin secretion in INS-1 β-cells

2004 ◽  
Vol 286 (5) ◽  
pp. E818-E827 ◽  
Author(s):  
Jingsong Li ◽  
Ruihua Luo ◽  
Anjaneyulu Kowluru ◽  
GuoDong Li
Keyword(s):  
Insulin Secretion ◽  
Time Course ◽  
Morphological Changes ◽  
Secretory Granules ◽  
Late Phase ◽  
Inhibitory Effect ◽  
Dominant Negative ◽  
Secretory Process ◽  
Β Cells ◽  
Rac1 Gtpase

Stimulation of insulin secretion by glucose and other secretagogues from pancreatic islet β-cells is mediated by multiple signaling pathways. Rac1 is a member of Rho family GTPases regulating cytoskeletal organization, and recent evidence also implicates Rac1 in exocytotic processes. Herein, we report that exposure of insulin-secreting (INS) cells to stimulatory glucose concentrations caused translocation of Rac1 from cytosol to the membrane fraction (including the plasmalemma), an indication of Rac1 activation. Furthermore, glucose stimulation increased Rac1 GTPase activity. Time course study indicates that such an effect is demonstrable only after 15 min stimulation with glucose. Expression of a dominant-negative Rac1 mutant (N17Rac1) abolished glucose-induced translocation of Rac1 and significantly inhibited insulin secretion stimulated by glucose and forskolin. This inhibitory effect on glucose-stimulated insulin secretion was more apparent in the late phase of secretion. However, N17Rac1 expression did not significantly affect insulin secretion induced by high K+. INS-1 cells expressing N17Rac1 also displayed significant morphological changes and disappearance of F-actin structures. Expression of wild-type Rac1 or a constitutively active Rac1 mutant (V12Rac1) did not significantly affect either the stimulated insulin secretion or basal release, suggesting that Rac1 activation is essential, but not sufficient, for evoking secretory process. These data suggest, for the first time, that Rac1 may be involved in glucose- and forskolin-stimulated insulin secretion, possibly at the level of recruitment of secretory granules through actin cytoskeletal network reorganization.

scholarly journals Regulation of Insulin Secretion and β-Cell Mass by Activating Signal Cointegrator 2

2006 ◽  
Vol 26 (12) ◽  
pp. 4553-4563 ◽  
Author(s):  
Seon-Yong Yeom ◽  
Geun Hyang Kim ◽  
Chan Hee Kim ◽  
Heun Don Jung ◽  
So-Yeon Kim ◽  
...  
Keyword(s):  
Insulin Secretion ◽  
Endocrine Cells ◽  
Potential Role ◽  
Cell Mass ◽  
Dominant Negative ◽  
Transcriptional Coactivator ◽  
Β Cells ◽  
Β Cell ◽  
Islet Mass

ABSTRACT Activating signal cointegrator 2 (ASC-2) is a transcriptional coactivator of many nuclear receptors (NRs) and other transcription factors and contains two NR-interacting LXXLL motifs (NR boxes). In the pancreas, ASC-2 is expressed only in the endocrine cells of the islets of Langerhans, but not in the exocrine cells. Thus, we examined the potential role of ASC-2 in insulin secretion from pancreatic β-cells. Overexpressed ASC-2 increased glucose-elicited insulin secretion, whereas insulin secretion was decreased in islets from ASC-2+/− mice. DN1 and DN2 are two dominant-negative fragments of ASC-2 that contain NR boxes 1 and 2, respectively, and block the interactions of cognate NRs with the endogenous ASC-2. Primary rat islets ectopically expressing DN1 or DN2 exhibited decreased insulin secretion. Furthermore, relative to the wild type, ASC-2+/− mice showed reduced islet mass and number, which correlated with increased apoptosis and decreased proliferation of ASC-2+/− islets. These results suggest that ASC-2 regulates insulin secretion and β-cell survival and that the regulatory role of ASC-2 in insulin secretion appears to involve, at least in part, its interaction with NRs via its two NR boxes.

Magnesium requirement for somatostatin inhibition of insulin secretion

1984 ◽  
Vol 105 (1) ◽  
pp. 83-86 ◽  
Author(s):  
Donald L. Curry ◽  
Leslie L. Bennett
Keyword(s):  
Insulin Secretion ◽  
Calcium Concentration ◽  
Inhibitory Effect ◽  
Secretory Process ◽  
Magnesium Ion ◽  
Second Phase ◽  
Intracellular Membrane ◽  
Calcium Magnesium ◽  
First Phase Insulin Secretion ◽  
Insulin Secretory Response

Abstract. Rat pancreas perfusions were performed using a perfusate with a fixed calcium concentration of 5 mEq/l and magnesium varying from 0 to 0.6 mEq/dl. Insulin secretion was stimulated by a constant glucose infusion of 300 mg/dl. This glucose concentration produces the typical biphasic insulin secretory response. We observed that in the absence of magnesium, somatostatin concentrations of 0.5 and 2.0 ng/ml were without effect on first phase insulin secretion. However, these same somatostatin levels produced 50% or more inhibition of insulin secretion in the presence of magnesium at 0.3 or 0.6 mEq/l. Similarly, in the absence of magnesium, somatostatin at 50 ng/ml failed to inhibit second phase insulin secretion, whereas this same somatostatin level produced about 50% inhibition of insulin secretion in the presence of magnesium at 0.3 mEq/l. Thus, altering perfusate magnesium concentrations without changing calcium is an important determinant of the degree of inhibition of secretion produced by somatostatin. In particular, in the absence of magnesium ion, somatostatin concentrations which would 'normally' produce 50% inhibition of secretion (ID50) are without effect. Therefore, magnesium ion is necessary for the full inhibitory effect of somatostatin to occur. These results suggest that inhibitors, as well as potentiators, of the insulin secretory process may act by altering intracellular/membrane calcium-magnesium ratios, but in opposite directions.

scholarly journals Decreased Insulin Secretion and Accumulation of Triglyceride in β Cells Overexpressing a Dominant-negative Form of AMP-activated Protein Kinase

2010 ◽  
Vol 57 (2) ◽  
pp. 141-152 ◽  
Author(s):  
Yukiko OKAZAKI ◽  
Kazuhiro ETO ◽  
Tokuyuki YAMASHITA ◽  
Masayuki OKAMOTO ◽  
Mitsuru OHSUGI ◽  
...  
Keyword(s):  
Insulin Secretion ◽  
Protein Kinase ◽  
Dominant Negative ◽  
Β Cells ◽  
Dominant Negative Form ◽  
Amp Activated Protein Kinase ◽  
Negative Form

Preptin derived from proinsulin-like growth factor II (proIGF-II) is secreted from pancreatic islet β-cells and enhances insulin secretion

2001 ◽  
Vol 360 (2) ◽  
pp. 431-439 ◽  
Author(s):  
Christina M. BUCHANAN ◽  
Anthony R. J. PHILLIPS ◽  
Garth J. S. COOPER
Keyword(s):  
Insulin Secretion ◽  
Growth Factor ◽  
Pancreatic Islet ◽  
Secretory Granules ◽  
Second Phase ◽  
Β Cells ◽  
Perfused Rat Pancreas ◽  
Amino Acid Peptide ◽  
Factor Ii ◽  
Second Phases

Pancreatic islet β-cells secrete the hormones insulin, amylin and pancreastatin. To search for further β-cell hormones, we purified peptides from secretory granules isolated from cultured murine βTC6-F7 β-cells. We identified a 34-amino-acid peptide (3948Da), corresponding to Asp69–Leu102 of the proinsulin-like growth factor II E-peptide, which we have termed ‘preptin’. Preptin, is present in islet β-cells and undergoes glucose-mediated co-secretion with insulin. Synthetic preptin increases insulin secretion from glucose-stimulated βTC6-F7 cells in a concentration-dependent and saturable manner. Preptin infusion into the isolated, perfused rat pancreas increases the second phase of glucose-mediated insulin secretion by 30%, while anti-preptin immunoglobulin infusion decreases the first and second phases of insulin secretion by 29 and 26% respectively. These findings suggest that preptin is a physiological amplifier of glucose-mediated insulin secretion.

scholarly journals N-Type Ca2+-Channels in Murine Pancreatic β-Cells Are Inhibited by an Exclusive Coupling with Somatostatin Receptor Subtype 1

Endocrinology ◽  
2009 ◽  
Vol 150 (2) ◽  
pp. 741-748 ◽  
Author(s):  
Paul A. Smith
Keyword(s):  
Insulin Secretion ◽  
Somatostatin Receptor ◽  
Inhibitory Effect ◽  
Receptor Subtype ◽  
Receptor Subtypes ◽  
Β Cells ◽  
Pancreatic Β Cells ◽  
Somatostatin Receptor Subtypes ◽  
Perforated Patch Clamp ◽  
Ca2 Channels

Somatostatin (SRIF) is a well-established inhibitor of insulin secretion, an effect in part mediated by a direct inhibition of voltage-operated Ca2+-channels. However, the identity of the somatostatin receptor subtypes (SSTRs) and voltage-operated Ca2+-channels involved in this process are unknown. Whole-cell perforated patch-clamp methods were applied to the murine pancreatic β-cell line, MIN6, to explore the molecular pharmacology of this problem. SRIF-14 inhibited voltage-gated Ca2+ currents (ICa2+) by 19 ± 3% (n=24) with a pEC50 = 9.05 (95% confidence limits 9–9.1). This action was mimicked solely by 100 nm CH-275, a selective agonist at the somatostatin type 1 receptor (SSTR1), but not by 100 nm BIM-23027, L-362855, or NNC-269100; agonists selective for the other four SSTRs known to exist in MIN6. The inhibition of ICa2+ produced by SRIF and CH-275 was insensitive to pertussis toxin but was reversed by a prepulse to +100 mV. The inhibition of ICa2+ by SRIF-14 was unaffected by 20 μm nifedipine, an inhibitor of L-type Ca2+ channels. Application of the specific N-type Ca2+ channel (Cav2.2) inhibitor ω-conotoxin GV1A at 100 nm mimicked, and as a consequence abolished, the inhibitory effect of SRIF-14 on ICa2+. SRIF selectively inhibits N-type Ca2+-channels in murine pancreatic β-cells via exclusive coupling with SSTR1. These findings help explain how SSTR1 activation can inhibit insulin secretion in pancreatic β-cells and suggest a possible new therapeutic lead for treatment of hyperinsulinemia. In pancreatic β-cells, somatostatin selectively inhibits N-type, but not other, Ca2+-channels via a direct and exclusive coupling with somatostatin receptor subtype 1.

scholarly journals Human stem cell model of HNF1A deficiency shows uncoupled insulin to C-peptide secretion with accumulation of abnormal insulin granules

2021 ◽  
Author(s):  
Bryan J. González ◽  
Haoquan Zhao ◽  
Jacqueline Niu ◽  
Damian J. Williams ◽  
Jaeyop Lee ◽  
...  
Keyword(s):  
Stem Cells ◽  
Stem Cell ◽  
Insulin Secretion ◽  
Intracellular Calcium ◽  
Cell Fate ◽  
Endocrine Cells ◽  
Secretory Granules ◽  
Β Cells ◽  
C Peptide ◽  
Pancreatic Endocrine Cells

AbstractMutations in HNF1A cause Maturity Onset Diabetes of the Young type 3 (MODY3), the most prevalent form of monogenic diabetes. We generated stem cell-derived pancreatic endocrine cells from human embryonic stem cells (hESCs) with induced hypomorphic mutations in HNF1A. Using these cells, we show that HNF1A orchestrates a transcriptional program required for distinct aspects of β-cell fate and function. During islet cell differentiation, HNF1A deficiency biases islet endocrine cells towards an α-cell fate associated with PAX4 down-regulation. HNF1A- deficient β-cells display impaired basal and glucose stimulated-insulin secretion in association with a reduction in CACNA1A and intracellular calcium levels, and impaired insulin granule exocytosis in association with SYT13 down-regulation. Knockout of PAX4, CACNA1A and SYT13 reproduce the relevant phenotypes. Reduction of insulin secretion is associated with accumulation of enlarged secretory granules, and altered stoichiometry of secreted insulin to C-peptide. In HNF1A deficient β-cells, glibenclamide, a sulfonylurea drug used in the treatment of MODY3 patients, increases intracellular calcium to levels beyond those achieved by glucose, and restores C-peptide and insulin secretion to a normal stoichiometric ratio. To study HNF1A deficiency in the context of a human disease model, we also generated stem cell-derived pancreatic endocrine cells from two MODY3 patient’s induced pluripotent stem cells (iPSCs). While insulin secretion defects are constitutive in cells with complete HNF1A loss of function, β-cells heterozygous for hypomorphic HNF1A mutations are initially normal, but lose the ability to secrete insulin and acquire abnormal stoichiometric secretion ratios. Importantly, the defects observed in these stem cell models are also seen in circulating proportions of insulin:C-peptide in nine MODY3 patients.One sentence of summaryDeficiency of the transcription factor HNF1A biases islet endocrine cell fate towards α-cells, impairs intracellular calcium homeostasis and insulin exocytosis, alters the stoichiometry of insulin to C-peptide release, and leads to an accumulation of abnormal insulin secretory granules in β-cells.

Structural finding of R/S-3,4-dihydro-2,2-dimethyl-6-halo-4-(substituted phenylaminocarbonylamino)-2H-1-benzopyrans as selective pancreatic β-cells KATP-pβ channel openers

2007 ◽  
Vol 85 (12) ◽  
pp. 1053-1063 ◽  
Author(s):  
Sk. Mahasin Alam ◽  
Soma Samanta ◽  
Amit Kumar Halder ◽  
Soumya Basu ◽  
Tarun Jha
Keyword(s):  
Insulin Secretion ◽  
Negative Impact ◽  
Inhibitory Effect ◽  
Van Der Waals Interactions ◽  
Β Cells ◽  
Pancreatic Β Cells ◽  
Qsar Study ◽  
Water Partition Coefficient ◽  
Qsar Models ◽  
Leave One Out

R/S-3,4-Dihydro-2,2-dimethyl-6-halo-4-(substituted phenylaminocarbonyl-amino)-2H-1-benzopyrans are pancreatic β-cells potassium (KATP-pβ) channel openers with inhibitory effect on insulin secretion. To find the more active and effective benzopyrans as selective potassium (KATP-pβ) channel openers towards the pancreatic tissues, quantitative structure–activity relationships (QSAR) study was performed using E-state and R-state indices along with Wang–Ford charges, n-octanol/water partition coefficient, molar refractivity, and indicator parameters. QSAR models were developed by statistical techniques, e.g., multiple linear regression (MLR), principle component regression analysis (PCRA), and partial least squares (PLS) analysis. The generated equations were validated by the leave-one-out cross-validation method. The models show the importance of ETSA indices of atom numbers 16, 17, 18, 19, 21 as well as 22. The positive coefficient of S16, S17, S18, S19, S21, and S22 indicate that with the increase of the value of E-state indices, desired activity decreases. RTSA index is also important for the biological activity, and the atom numbers 16, 17, 18, 19, 20 and 22 are involved in van der Waals interactions. RTSA index also possesses negative impact on the inhibition of residual insulin secretion. Wang–Ford charges of some particular atoms are also important for the inhibition. Increase of n-octanol/water partition coefficients of compounds inhibit insulin secretion, and the presence of chlorine atom at m- and p- positions of the phenyl ring B is necessary for the inhibition of residual insulin secretion.Key words: benzopyran derivatives, potassium channel openers, PCRA, PLS, QSAR.

Adrenomedullin inhibits insulin exocytosis via pertussis toxin-sensitive G protein-coupled mechanism

2006 ◽  
Vol 291 (1) ◽  
pp. E9-E14 ◽  
Author(s):  
Nobuo Sekine ◽  
Koji Takano ◽  
Nako Kimata-Hayashi ◽  
Takashi Kadowaki ◽  
Toshiro Fujita
Keyword(s):  
Insulin Secretion ◽  
G Proteins ◽  
Pertussis Toxin ◽  
Colorimetric Assay ◽  
Calcium Ionophore ◽  
Cytosolic Calcium ◽  
Inhibitory Effect ◽  
Calcium Currents ◽  
Β Cells ◽  
Insulin Exocytosis

Direct effects of adrenomedullin on insulin secretion from pancreatic β-cells were investigated using a differentiated insulin-secreting cell line INS-1. Adrenomedullin (1–100 pM) inhibited insulin secretion at both basal (3 mM) and high (15 mM) glucose concentrations, although this inhibitory effect was not observed at higher concentrations of adrenomedullin. The inhibition of glucose-induced insulin secretion by adrenomedullin was restored with 12-h pretreatment with 1 μg/ml pertussis toxin (PTX), suggesting that this effect could be mediated by PTX-sensitive G proteins. Cellular glucose metabolism evaluated by 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide colorimetric assay was not affected by adrenomedullin at concentrations that inhibited insulin secretion. Moreover, electrophysiological studies revealed that 10 pM adrenomedullin had no effect on membrane potential, voltage-gated calcium currents, or cytosolic calcium concentration induced by 15 mM glucose. Finally, insulin release induced by cAMP-raising agents, such as forskolin plus 3-isobutyl-1-methylxanthine or the calcium ionophore ionomycin, was significantly inhibited by 10 and 100 pM adrenomedullin. In conclusion, adrenomedullin at picomolar concentrations directly inhibited insulin secretion from β-cells. This effect is likely due to the inhibition of insulin exocytosis through the activation of PTX-sensitive G proteins.

scholarly journals In pancreatic β-cells myosin 1b regulates glucose-stimulated insulin secretion by modulating an early step in insulin granule trafficking from the Golgi

2021 ◽  
pp. mbc.E21-03-0094
Author(s):  
Hiroshi Tokuo ◽  
Shigeru Komaba ◽  
Lynne M. Coluccio
Keyword(s):  
Insulin Secretion ◽  
Islet Cells ◽  
Early Stage ◽  
Secretory Granules ◽  
Β Cells ◽  
Pancreatic Β Cells ◽  
Glucose Levels ◽  
Insulin Granule ◽  
Glucose Stimulated Insulin Secretion ◽  
Insulinoma Cells

Pancreatic β-cells secrete insulin, which controls blood glucose levels, and defects in insulin secretion are responsible for diabetes mellitus. The actin cytoskeleton and some myosins support insulin granule trafficking and release, although a role for the class I myosin Myo1b, an actin- and membrane-associated load-sensitive motor, in insulin biology is unknown. We found by immunohistochemistry that Myo1b is expressed in islet cells of rat pancreas. In cultured rat insulinoma 832/13 cells Myo1b localized near actin patches, the trans-Golgi network (TGN) marker TGN38, and insulin granules in the perinuclear region. Myo1b depletion by siRNA in 832/13 cells reduced intracellular proinsulin and insulin content and glucose-stimulated insulin secretion (GSIS), and led to the accumulation of (pro)insulin SGs at the TGN. Using an in situ fluorescent pulse-chase strategy to track nascent proinsulin (Bearrows et al., 2019), Myo1b depletion in insulinoma cells reduced the number of (pro)insulin-containing secretory granules budding from the TGN. The studies indicate for the first time that in pancreatic β-cells Myo1b controls GSIS at least in part by mediating an early stage in insulin granule trafficking from the TGN.

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