Contribution of Na/Ca exchange to Ca2+ outflow and entry in the rat pancreatic beta-cell: studies with antisense oligonucleotides.
To characterize the role played by Na/Ca exchange in the pancreatic beta-cell, phosphorothioated antisense oligonucleotides (AS-oligos) were used to knock down the exchanger in rat pancreatic beta-cells. Na/Ca exchange activity was evaluated by measuring cytosolic free Ca2+ concentration ([Ca2+]i) in single cells using fura-2. Exposure of beta-cells to 500 nmol/l of the AS-oligos for 24 h inhibited Na/Ca exchange activity by approximately 77%. In contrast, control oligonucleotides (scrambled and mismatched) did not affect Na/Ca exchange activity. In AS-oligo-treated cells, the increase in [Ca2+]i induced by membrane depolarization (K+ or the hypoglycemic sulfonylurea, tolbutamide) was reduced by 28 or 40%, respectively. Likewise, the rate of [Ca2+]i decrease after K+ or tolbutamide removal was reduced by 72 or 40%, respectively. AS-oligos treatment also abolished the nifedipine-resistant increase in [Ca2+]i induced by K+ and profoundly altered the oscillatory or sustained increases in [Ca2+]i induced by 11.1 mmol/l glucose. The present study shows that AS-oligos may specifically inhibit Na/Ca exchange in rat pancreatic beta-cells. In those cells, Na/Ca exchange appears to mediate Ca2+ entry in response to membrane depolarization and to be responsible for up to 70% of Ca2+ removal from the cytoplasm upon membrane repolarization.
Van Eylen
F
F
Laboratory of Pharmacology, Université Libre de Bruxelles, School of Medicine, Brussels, Belgium.
Lebeau
C
C
Albuquerque-Silva
J
J
Herchuelz
A
A
D000818:Animals; D002118:Calcium / Q000378:metabolism*; D002121:Calcium Channel Blockers / Q000494:pharmacology; D002448:Cell Adhesion / Q000187:drug effects; D005947:Glucose / Q000494:pharmacology; D007004:Hypoglycemic Agents / Q000494:pharmacology; D017136:Ion Transport / Q000187:drug effects; D007515:Islets of Langerhans / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D009543:Nifedipine / Q000494:pharmacology; D016376:Oligonucleotides, Antisense / Q000494:pharmacology; D011189:Potassium Chloride / Q000494:pharmacology; D012333:RNA, Messenger / Q000187:drug effects / Q000378:metabolism; D051381:Rats; D017208:Rats, Wistar; D012964:Sodium / Q000378:metabolism*; D019831:Sodium-Calcium Exchanger / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D014044:Tolbutamide / Q000494:pharmacology
D002121:Calcium Channel Blockers; D007004:Hypoglycemic Agents; D016376:Oligonucleotides, Antisense; D012333:RNA, Messenger; D019831:Sodium-Calcium Exchanger; D011189:Potassium Chloride; D014044:Tolbutamide; D012964:Sodium; D009543:Nifedipine; D005947:Glucose; D002118:Calcium