Cloning of the promoters for the beta-cell ATP-sensitive K-channel subunits Kir6.2 and SUR1

The nature of ATP-sensitive K+ (K+ATP) channel-independent, insulinotropic action of glucose was investigated using non-glucose-primed pancreatic islets. When the beta-cell was depolarized with K+, glucose dose dependently stimulated insulin release despite inhibition of the K+ATP channel closure by diazoxide. K+ depolarization could be replaced with BAY K 8644, a calcium channel agonist. Prior fasting of rats and lowering ambient temperature greatly suppressed glucose oxidation and utilization by the islet cells and abolished insulin release in response to high glucose alone. However, under these conditions, the K+ATP channel-independent, glucose-induced insulin release was clearly demonstrable. p-Nitrophenyl-alpha-D-glucopyranoside (sweet taste inhibitor) but not its beta-isomer, neomycin (phospholipase C inhibitor) and staurosporine (C kinase blocker) inhibited the K+ATP channel-independent, insulinotropic action of glucose. For the K+ATP channel-independent glucose-induced insulin release 1) elevation of cytosolic calcium is required, 2) minute glucose metabolism is enough, if glucose metabolism is necessary, and 3) direct recognition of glucose molecule, phospholipase C, and protein kinase C appear to be involved.

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Metabolic Regulation of the Pancreatic Beta-Cell ATP-Sensitive K+ Channel: A Pas de Deux

Diabetes ◽

10.2337/diabetes.53.suppl_3.s113 ◽

2004 ◽

Vol 53 (Supplement 3) ◽

pp. S113-S122 ◽

Cited By ~ 91

Author(s):

A. Tarasov ◽

J. Dusonchet ◽

F. Ashcroft

Keyword(s):

Beta Cell ◽

Metabolic Regulation ◽

Pancreatic Beta Cell ◽

K Channel

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Glutamate is not a Major Conveyer of ATP-sensitive K+ Channel-Independent Glucose Action in Pancreatic Islet .BETA. Cell.

Endocrine Journal ◽

10.1507/endocrj.48.391 ◽

2001 ◽

Vol 48 (3) ◽

pp. 391-395 ◽

Cited By ~ 21

Author(s):

SATOKO YAMADA ◽

MITSUHISA KOMATSU ◽

YOSHIHIKO SATO ◽

KEISHI YAMAUCHI ◽

TORU AIZAWA ◽

...

Keyword(s):

Beta Cell ◽

Pancreatic Islet ◽

K Channel ◽

Islet Beta Cell ◽

Pancreatic Islet Beta Cell

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The structure and function of the ATP-sensitive K+ channel in insulin-secreting pancreatic beta-cells

Journal of Molecular Endocrinology ◽

10.1677/jme.0.0220113 ◽

1999 ◽

Vol 22 (2) ◽

pp. 113-123 ◽

Cited By ~ 97

Author(s):

T Miki ◽

K Nagashima ◽

S Seino

Keyword(s):

Molecular Structure ◽

Insulin Secretion ◽

Beta Cell ◽

Beta Cells ◽

Katp Channel ◽

Pancreatic Beta Cells ◽

Pancreatic Beta Cell ◽

Molecular Genetic ◽

Katp Channels ◽

K Channel

ATP-sensitive K+ channels (KATP channels) play important roles in many cellular functions by coupling cell metabolism to electrical activity. The KATP channels in pancreatic beta-cells are thought to be critical in the regulation of glucose-induced and sulfonylurea-induced insulin secretion. Until recently, however, the molecular structure of the KATP channel was not known. Cloning members of the novel inwardly rectifying K+ channel subfamily Kir6.0 (Kir6.1 and Kir6.2) and the sulfonylurea receptors (SUR1 and SUR2) has clarified the molecular structure of KATP channels. The pancreatic beta-cell KATP channel comprises two subunits: a Kir6.2 subunit and an SUR1 subunit. Molecular biological and molecular genetic studies have provided insights into the physiological and pathophysiological roles of the pancreatic beta-cell KATP channel in insulin secretion.

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Characterization of the solubilized glibenclamide receptor in a hamster pancreatic β-cell line, HIT T15

Biochemical Journal ◽

10.1042/bj2770619 ◽

1991 ◽

Vol 277 (3) ◽

pp. 619-624 ◽

Cited By ~ 16

Author(s):

I Niki ◽

M Welsh ◽

P O Berggren ◽

P Hubbard ◽

S J H Ashcroft

Keyword(s):

Beta Cell ◽

Cell Line ◽

Beta Cells ◽

Specific Binding ◽

Pancreatic Beta Cell ◽

Active Form ◽

K Channel ◽

Scatchard Analysis ◽

Beta Cell Line ◽

20 Nm

The glibenclamide receptor, a putative ATP-sensitive K+ channel in the hamster pancreatic beta-cell line HIT T15, was solubilized by using the zwitterionic detergent CHAPS. [3H]Glibenclamide binding was dependent on the incubation time and on the concentration of soluble membrane protein. Over 80% of [3H]glibenclamide bound could be displaced with 1 microM non-labelled glibenclamide. The curve relating specific binding to the concentration of [3H]glibenclamide (1-20 nM) showed saturation kinetics. Scatchard analysis suggested a single class of non-interacting binding sites with a Kd of 3.3 nM and a Bmax. of 90 fmol/mg of protein. [3H]Glibenclamide binding to solubilized membranes was inhibited by glibenclamide, tolbutamide and meglitinide. The relative potency of these agents on binding of [3H]glibenclamide to solubilized membranes was similar to that observed with microsomal preparations and paralleled their effects on K-ATP channel activity, measured as 86Rb efflux. These data show that the sulphonylurea receptor in the pancreatic beta-cell can be solubilized in an active form retaining specificity for sulphonylureas. ADP, which inhibits [3H]glibenclamide binding to microsomal preparations or intact HIT beta-cells, did not inhibit binding to the solubilized receptor. Incubation of intact HIT beta-cells with 125I-glibenclamide derivative followed by exposure to u.v. light resulted in covalent labelling of a peptide of 65 kDa on SDS/PAGE. The extent of labelling increased with 125I-glibenclamide derivative concentration (1-20 nM) and was inhibited in the presence of excess unlabelled glibenclamide.

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The β-cell glibenclamide receptor is an ADP-binding protein

Biochemical Journal ◽

10.1042/bj2680713 ◽

1990 ◽

Vol 268 (3) ◽

pp. 713-718 ◽

Cited By ~ 33

Author(s):

I Niki ◽

J L Nicks ◽

S J H Ashcroft

Keyword(s):

Beta Cell ◽

Insulin Release ◽

Plasma Membranes ◽

Pancreatic Beta Cell ◽

Atp Depletion ◽

K Channel ◽

Whole Cells ◽

Cell Plasma ◽

Beta Cell Line ◽

Extracellular Side

The effects of ADP on [3H]glibenclamide binding to membranes and whole cells, the activity of the ATP-sensitive K+ channel (K-ATP channel), intracellular Ca2+ concentration and insulin secretion were studied in a hamster pancreatic beta-cell line, HIT T15. ADP dose-dependently inhibited [3H]glibenclamide binding to membranes and to whole cells in a competitive manner. ADP-agarose also inhibited the binding to whole cells. The activity of the K-ATP channel was assayed by measuring 86Rb efflux from whole cells. ADP inhibited the 86Rb efflux elicited either by diazoxide or by ATP depletion. In the presence, but not in the absence, of extracellular Ca2+, ADP evoked a rapid and sustained increase in intracellular Ca2+ concentration as estimated with the fluorescent dye quin 2. Insulin release from HIT cells was also increased by 0.5-2 mM-ADP in the presence of 0.5 mM-glucose. These effects of ADP on glibenclamide binding, K-ATP channel activity and insulin release were specific for ADP, and were not reproduced by any other nucleotide so far tested. The present findings strongly suggest that ADP and sulphonylureas have common binding sites on the extracellular side of beta-cell plasma membranes, where they inhibit the activity of the K-ATP channel, resulting in an increase in intracellular Ca2+ concentration and insulin release.

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