The sodium-calcium antiport of heart mitochondria is not electroneutral.
Heart mitochondria contain a nNa+/Ca2+ antiport that participates in the regulation of matrix [Ca2+]. Based largely on a single study (Brand, M. D. (1985) Biochem. J. 229, 161-166), there has been a consensus that this antiport promotes the electroneutral exchange of two Na+ for one Ca2+. However, a recent study in our laboratory (Baysal, K., Jung, D. W., Gunter, K. K., Gunter, T. P., and Brierley, G. P. (1994) Am. J. Physiol. 266, C800-C808) has shown that the Na(+)-dependent efflux of Ca2+ from heart mitochondria has more energy available to it than can be supplied by a passive 2Na+/Ca2+ exchange. We have therefore re-examined Brand's protocols using fluorescent probes to monitor matrix pH and free [Ca2+]. Respiring heart mitochondria, suspended in KCl and treated with ruthenium red to block Ca2+ influx, extrude Ca2+ and establish a large [Ca2+]out:[Ca2+]matrix gradient. The extrusion of Ca2+ under these conditions is Na(+)-dependent and diltiazem-sensitive and can be attributed to the nNa+/Ca2+ antiport. Addition of nigericin increases the membrane potential (delta psi) and decreases delta pH to 0.1 or less, but has virtually no effect on the magnitude of the [Ca2+] gradient. Under these conditions a gradient maintained by electroneutral 2Na+/Ca2+ antiport should be abolished because the mitochondrial Na+/H+ antiport keeps the [Na+] gradient equivalent to the [H+] gradient. The [Ca2+] gradient is abolished, however, when an uncoupler is added to dissipate delta psi or when the exogenous electroneutral antiport BrA23187 is added. In addition, [Ca2+] influx via the nNa+/Ca2+ antiport in nonrespiring mitochondria is enhanced when delta psi is abolished. These results are consistent with Ca2+ extrusion by an electrophoretic antiport that can respond to delta psi but not with an electroneutral antiport.
Jung
D W
DW
Department of Medical Biochemistry, Ohio State University, Columbus 43210.
Baysal
K
K
Brierley
G P
GP
D000818:Animals; D017920:Antiporters / Q000187:drug effects / Q000378:metabolism* / Q000502:physiology; D002118:Calcium / Q000378:metabolism*; D002259:Carbonyl Cyanide p-Trifluoromethoxyphenylhydrazone / Q000494:pharmacology; D002417:Cattle; D016572:Cyclosporine / Q000494:pharmacology; D004110:Diltiazem / Q000494:pharmacology; D005456:Fluorescent Dyes; D006863:Hydrogen-Ion Concentration; D008564:Membrane Potentials; D008929:Mitochondria, Heart / Q000187:drug effects / Q000378:metabolism* / Q000502:physiology; D009550:Nigericin / Q000494:pharmacology; D009861:Onium Compounds / Q000494:pharmacology; D009943:Organophosphorus Compounds / Q000494:pharmacology; D010100:Oxygen / Q000378:metabolism; D012964:Sodium / Q000378:metabolism*; D019831:Sodium-Calcium Exchanger; D013816:Thermodynamics; D014634:Valinomycin / Q000494:pharmacology
D017920:Antiporters; D005456:Fluorescent Dyes; D009861:Onium Compounds; D009943:Organophosphorus Compounds; D019831:Sodium-Calcium Exchanger; D014634:Valinomycin; D002259:Carbonyl Cyanide p-Trifluoromethoxyphenylhydrazone; D016572:Cyclosporine; D012964:Sodium; D004110:Diltiazem; D009550:Nigericin; D010100:Oxygen; D002118:Calcium; C013289:tetraphenylphosphonium