Attenuation of maitotoxin-induced cytotoxicity in rat aortic smooth muscle cells by inhibitors of Na+/Ca2+ exchange, and calpain activation.
The highly potent marine toxin maitotoxin (MTX) evoked an increase in cytosolic Ca(2+) levels in fura-2 loaded rat aortic smooth muscle cells, which was dependent on extracellular Ca(2+). This increase was almost fully inhibited by KB-R7943, a potent selective inhibitor of the reverse mode of the Na(+)/Ca(2+) exchanger (NCX). Cell viability was assessed using ethidium bromide uptake and the alamarBlue cytotoxicity assay. In both assays MTX-induced toxicity was attenuated by KB-R7943, as well as by MDL 28170, a membrane permeable calpain inhibitor. Maitotoxin-evoked contractions of rat aortic strip preparations in vitro, which persist following washout of the toxin, were relaxed by subsequent addition of KB-R7943 or MDL 28170, either in the presence of, or following washout of MTX. These results suggest that MTX targets the Na(+)/Ca(2+) exchanger and causes it to operate in reverse mode (Na(+) efflux/Ca(2+) influx), thus leading to calpain activation, NCX cleavage, secondary Ca(2+) overload and cell death.
Frew
Robert
R
Defence Research and Development Canada-Suffield, Medicine Hat, Alberta, Canada.
Wang
Yushan
Y
Weiss
Tracy M
TM
Nelson
Peggy
P
Sawyer
Thomas W
TW
D000818:Animals; D002118:Calcium / Q000378:metabolism*; D002154:Calpain / Q000378:metabolism*; D002478:Cells, Cultured; D004789:Enzyme Activation / Q000187:drug effects; D004996:Ethidium / Q000032:analysis; D005456:Fluorescent Dyes / Q000032:analysis; D005470:Fluorometry; D016257:Fura-2 / Q000032:analysis; D066298:In Vitro Techniques; D007202:Indicators and Reagents; D017136:Ion Transport / Q000187:drug effects*; D008387:Marine Toxins / Q000037:antagonists & inhibitors / Q000494:pharmacology*; D009119:Muscle Contraction; D009131:Muscle, Smooth, Vascular / Q000187:drug effects; D010078:Oxazines; D035964:Oxocins / Q000037:antagonists & inhibitors / Q000494:pharmacology*; D051381:Rats; D012964:Sodium / Q000378:metabolism*; D019831:Sodium-Calcium Exchanger / Q000037:antagonists & inhibitors; D013890:Thiourea / Q000031:analogs & derivatives* / Q000494:pharmacology; D014966:Xanthenes; D000095702:Polyether Toxins
D002118:Calcium; D002154:Calpain; D004996:Ethidium; D005456:Fluorescent Dyes; D016257:Fura-2; D007202:Indicators and Reagents; D008387:Marine Toxins; D010078:Oxazines; D035964:Oxocins; D012964:Sodium; D019831:Sodium-Calcium Exchanger; D013890:Thiourea; D014966:Xanthenes; C101670:2-(2-(4-(4-nitrobenzyloxy)phenyl)ethyl)isothiourea methanesulfonate; C005843:resazurin; C020394:maitotoxin; D000095702:Polyether Toxins