title: Anoctamin 1 (Ano1) is required for glucose-induced membrane potential oscillations and insulin secretion by murine β-cells.
abstract: Anions such as Cl(-) and HCO3 (-) are well known to play an important role in glucose-stimulated insulin secretion (GSIS). In this study, we demonstrate that glucose-induced Cl(-) efflux from β-cells is mediated by the Ca(2+)-activated Cl(-) channel anoctamin 1 (Ano1). Ano1 expression in rat β-cells is demonstrated by reverse transcriptase-polymerase chain reaction, western blotting, and immunohistochemistry. Typical Ano1 currents are observed in whole-cell and inside-out patches in the presence of intracellular Ca(++): at 1 μM, the Cl(-) current is outwardly rectifying, and at 2 μM, it becomes almost linear. The relative permeabilities of monovalent anions are NO3 (-) (1.83 ± 0.10) > Br(-) (1.42 ± 0.07) > Cl(-) (1.0). A linear single-channel current-voltage relationship shows a conductance of 8.37 pS. These currents are nearly abolished by blocking Ano1 antibodies or by the inhibitors 2-(5-ethyl-4-hydroxy-6-methylpyrimidin-2-ylthio)-N-(4-(4-methoxyphenyl)thiazol-2-yl)acetamide (T-AO1) and tannic acid (TA). These inhibitors induce a strong decrease of 16.7-mM glucose-stimulated action potential rate (at least 87 % on dispersed cells) and a partial membrane repolarization with T-AO1. They abolish or strongly inhibit the GSIS increment at 8.3 mM and at 16.7 mM glucose. Blocking Ano1 antibodies also abolish the 16.7-mM GSIS increment. Combined treatment with bumetanide and acetazolamide in low Cl(-) and HCO3 (-) media provokes a 65 % reduction in action potential (AP) amplitude and a 15-mV AP peak repolarization. Although the mechanism triggering Ano1 opening remains to be established, the present data demonstrate that Ano1 is required to sustain glucose-stimulated membrane potential oscillations and insulin secretion.
authors.affiliation: Laboratory of Cell and Molecular Physiology, Université Libre de Bruxelles, Campus Erasme, Brussels, Belgium.
authors.forename: Raphaël
authors.initials: R
authors.lastname: Crutzen
authors.affiliation: Laboratory of Cell and Molecular Physiology, Université Libre de Bruxelles, Campus Erasme, Brussels, Belgium.
authors.forename: Myrna
authors.initials: M
authors.lastname: Virreira
authors.affiliation: Laboratory of Cell and Molecular Physiology, Université Libre de Bruxelles, Campus Erasme, Brussels, Belgium.
authors.forename: Nicolas
authors.initials: N
authors.lastname: Markadieu
authors.affiliation: Laboratory of Physiology and Pharmacology, Université Libre de Bruxelles, Campus Erasme, Brussels, Belgium.
authors.forename: Vadim
authors.initials: V
authors.lastname: Shlyonsky
authors.affiliation: Laboratory of Physiology and Pharmacology, Université Libre de Bruxelles, Campus Erasme, Brussels, Belgium.
authors.forename: Abdullah
authors.initials: A
authors.lastname: Sener
authors.affiliation: Department of Biochemistry, Université Libre de Bruxelles, Campus Erasme, Brussels, Belgium.
authors.forename: Willy J
authors.initials: WJ
authors.lastname: Malaisse
authors.affiliation: Laboratory of Cell and Molecular Physiology, Université Libre de Bruxelles, Campus Erasme, Brussels, Belgium. renbeau@ulb.ac.be.
authors.forename: Renaud
authors.initials: R
authors.lastname: Beauwens
authors.affiliation: Laboratory of Histology, Histopathology and Neuroanatomy, Université Libre de Bruxelles, Campus Erasme, Brussels, Belgium.
authors.forename: Alain
authors.initials: A
authors.lastname: Boom
authors.affiliation: Laboratory of Physiology and Pharmacology, Université Libre de Bruxelles, Campus Erasme, Brussels, Belgium.
authors.forename: Philippe E
authors.initials: PE
authors.lastname: Golstein
mesh_terms: D000818:Animals; D000075369:Anoctamin-1; D002118:Calcium / Q000378:metabolism; D002478:Cells, Cultured; D018118:Chloride Channels / Q000037:antagonists & inhibitors / Q000378:metabolism*; D002712:Chlorides / Q000378:metabolism; D005089:Exocytosis; D005947:Glucose / Q000378:metabolism*; D006801:Humans; D007328:Insulin / Q000378:metabolism*; D050417:Insulin-Secreting Cells / Q000378:metabolism* / Q000502:physiology; D008564:Membrane Potentials; D051379:Mice; D008810:Mice, Inbred C57BL; D051381:Rats; D017208:Rats, Wistar
chemical_list: C531181:ANO1 protein, mouse; D000075369:Anoctamin-1; D018118:Chloride Channels; D002712:Chlorides; D007328:Insulin; D005947:Glucose; D002118:Calcium
keywords: Chloride; Islet; TMEM16A