Barium influx mediated by the cardiac sodium-calcium exchanger in transfected Chinese hamster ovary cells.
We examined Ba2+ influx using isotopic and fura-2 techniques in transfected Chinese hamster ovary cells expressing the bovine cardiac Na+/Ca2+ exchanger (CK1.4 cells). Ba2+ competitively inhibited exchange-mediated 45Ca2+ uptake with a Ki approximately 3 mM. Ba2+ uptake was stimulated by pretreating the cells with ouabain and by removing extracellular Na+, as expected for Na+/Ba2+ exchange activity. The maximal velocity of Ba2+ accumulation was estimated to be 50% of that for Ca2+. When the monovalent cation ionophore gramicidin was used to equilibrate internal and external concentrations of Na+, Ba2+ influx was negligible in the absence of Na+ and increased to a maximum at 20-40 mM Na+. At higher Na+ concentrations, Ba2+ influx declined, presumably due to the competition between Na+ and Ba2+ for transport sites on the exchanger. Unlike Ca2+, Ba2+ did not appear to be taken up by intracellular organelles. Thus, 133Ba2+ uptake in ouabain-treated cells was not reduced by mitochondrial inhibitors such as-Cl-CCP or oligomycin-rotenone. Moreover, intracellular Ca2+ stores that had been depleted of Ca2+ by pretreatment of the cells with ionomycin (a Ca2+ ionophore) remained empty during a subsequent period of Ba2+ influx. Ca2+ uptake or release by intracellular organelles secondarily regulated exchange activity through alterations in [Ca2+]i. Exchange-mediated Ba2+ influx was inhibited when cytosolic [Ca2+] was reduced to 20 nM or less and was accelerated at cytosolic Ca2+ concentrations of 25-50 nM We conclude that (a) Ba2+ substitutes for Ca2+ as a transport substrate for the exchanger, (b) cytosolic Ba2+ does not appear to be sequestered by intracellular organelles, and (c) exchange-mediated Ba2+ influx is accelerated by low concentrations of cytosolic Ca2+.
Condrescu
M
M
Department of Physiology, Pharmacology and Toxicology, University of Medicine and Dentistry of New Jersey, New Jersey Medical School, Newark 07103, USA.
Chernaya
G
G
Kalaria
V
V
Reeves
J P
JP
D000818:Animals; D001464:Barium / Q000378:metabolism*; D016466:CHO Cells / Q000378:metabolism* / Q000502:physiology; D002352:Carrier Proteins / Q000235:genetics* / Q000378:metabolism*; D002417:Cattle; D006224:Cricetinae; D007425:Intracellular Membranes / Q000378:metabolism; D009206:Myocardium / Q000378:metabolism*; D015388:Organelles / Q000378:metabolism; D009994:Osmolar Concentration; D012964:Sodium / Q000378:metabolism; D019831:Sodium-Calcium Exchanger; D014162:Transfection
D002352:Carrier Proteins; D019831:Sodium-Calcium Exchanger; D001464:Barium; D012964:Sodium