Anoctamin 1 (Ano1) is required for glucose-induced membrane potential oscillations and insulin secretion by murine β-cells

• Published in: Pflügers Archiv 2016-04, Vol.468 (4), p.573-591
• Main Authors: Crutzen, Raphaël, Virreira, Myrna, Markadieu, Nicolas, Shlyonsky, Vadim, Sener, Abdullah, Malaisse, Willy J., Beauwens, Renaud, Boom, Alain, Golstein, Philippe E.
• Format: Article
• Language: English
• Subjects: Animals; Anoctamin-1; Biomedical and Life Sciences; Biomedicine; Calcium - metabolism; Cell Biology; Cells, Cultured; Chloride Channels - antagonists & inhibitors; Chloride Channels - metabolism; Chlorides - metabolism; Exocytosis; Glucose - metabolism; Human Physiology; Humans; Insulin - metabolism; Insulin-Secreting Cells - metabolism; Insulin-Secreting Cells - physiology; Ion Channels; Ion Channels, Receptors and Transporters; Membrane Potentials; Mice; Mice, Inbred C57BL; Molecular Medicine; Neurosciences; Rats; Rats, Wistar; Receptors; Receptors and Transporters
• ISSN: 0031-6768; 1432-2013
• DOI: 10.1007/s00424-015-1758-5

Anions such as Cl − and HCO 3 − 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 NO 3 − (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 HCO 3 − 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.