Suppression of sulfonylurea- and glucose-induced insulin secretion in vitro and in vivo in mice lacking the chloride transport protein ClC-3.
Priming of insulin secretory granules for release requires intragranular acidification and depends on vesicular Cl(-)-fluxes, but the identity of the chloride transporter/ion channel involved is unknown. We tested the hypothesis that the chloride transport protein ClC-3 fulfills these actions in pancreatic beta cells. In ClC-3(-/-) mice, insulin secretion evoked by membrane depolarization (high extracellular K(+), sulfonylureas), or glucose was >60% reduced compared to WT animals. This effect was mirrored by a approximately 80% reduction in depolarization-evoked beta cell exocytosis (monitored as increases in cell capacitance) in single ClC-3(-/-) beta cells, as well as a 44% reduction in proton transport across the granule membrane. ClC-3 expression in the insulin granule was demonstrated by immunoblotting, immunostaining, and negative immuno-EM in a high-purification fraction of large dense-core vesicles (LDCVs) obtained by phogrin-EGFP labeling. The data establish the importance of granular Cl(-) fluxes in granule priming and provide direct evidence for the involvement of ClC-3 in the process.
Li
Dai-Qing
DQ
Lund University Diabetes Center, Department of Clinical Sciences Malmö, Lund University, Malmö, SE-205 02 Malmö, Sweden.
Jing
Xingjun
X
Salehi
Albert
A
Collins
Stephan C
SC
Hoppa
Michael B
MB
Rosengren
Anders H
AH
Zhang
Enming
E
Lundquist
Ingmar
I
Olofsson
Charlotta S
CS
Mörgelin
Matthias
M
Eliasson
Lena
L
Rorsman
Patrik
P
Renström
Erik
E
D000818:Animals; D002118:Calcium / Q000378:metabolism; D018118:Chloride Channels / Q000235:genetics / Q000378:metabolism*; D002712:Chlorides / Q000378:metabolism; D003594:Cytoplasmic Granules / Q000378:metabolism; D052216:Glucagon-Like Peptide 1 / Q000378:metabolism; D005947:Glucose / Q000378:metabolism*; D007328:Insulin / Q000378:metabolism*; D000078790:Insulin Secretion; D050417:Insulin-Secreting Cells / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism*; D051379:Mice; D018345:Mice, Knockout; D034622:RNA Interference; D013453:Sulfonylurea Compounds / Q000494:pharmacology*
D018118:Chloride Channels; D002712:Chlorides; C115376:ClC-3 channel; D007328:Insulin; D013453:Sulfonylurea Compounds; D052216:Glucagon-Like Peptide 1; D005947:Glucose; D002118:Calcium