Regulation of intracellular calcium in N1E-115 neuroblastoma cells: the role of Na(+)/Ca(2+) exchange.
In fura 2-loaded N1E-115 cells, regulation of intracellular Ca(2+) concentration ([Ca(2+)](i)) following a Ca(2+) load induced by 1 microM thapsigargin and 10 microM carbonylcyanide p-trifluoromethyoxyphenylhydrazone (FCCP) was Na(+) dependent and inhibited by 5 mM Ni(2+). In cells with normal intracellular Na(+) concentration ([Na(+)](i)), removal of bath Na(+), which should result in reversal of Na(+)/Ca(2+) exchange, did not increase [Ca(2+)](i) unless cell Ca(2+) buffer capacity was reduced. When N1E-115 cells were Na(+) loaded using 100 microM veratridine and 4 microg/ml scorpion venom, the rate of the reverse mode of the Na(+)/Ca(2+) exchanger was apparently enhanced, since an approximately 4- to 6-fold increase in [Ca(2+)](i) occurred despite normal cell Ca(2+) buffering. In SBFI-loaded cells, we were able to demonstrate forward operation of the Na(+)/Ca(2+) exchanger (net efflux of Ca(2+)) by observing increases (approximately 6 mM) in [Na(+)](i). These Ni(2+) (5 mM)-inhibited increases in [Na(+)](i) could only be observed when a continuous ionomycin-induced influx of Ca(2+) occurred. The voltage-sensitive dye bis-(1,3-diethylthiobarbituric acid) trimethine oxonol was used to measure changes in membrane potential. Ionomycin (1 microM) depolarized N1E-115 cells (approximately 25 mV). This depolarization was Na(+) dependent and blocked by 5 mM Ni(2+) and 250-500 microM benzamil. These data provide evidence for the presence of an electrogenic Na(+)/Ca(2+) exchanger that is capable of regulating [Ca(2+)](i) after release of Ca(2+) from cell stores.
Kopper
Kara L
KL
Allergan, Inc., Department of Biological Sciences, Irvine, California 92612, USA.
Adorante
Joseph S
JS
D000818:Animals; D001932:Brain Neoplasms / Q000378:metabolism*; D002118:Calcium / Q000378:metabolism*; D002259:Carbonyl Cyanide p-Trifluoromethoxyphenylhydrazone / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D015759:Ionomycin / Q000494:pharmacology; D007476:Ionophores / Q000494:pharmacology; D008564:Membrane Potentials / Q000502:physiology; D051379:Mice; D009447:Neuroblastoma / Q000378:metabolism*; D009532:Nickel / Q000378:metabolism; D010042:Ouabain / Q000494:pharmacology; D012604:Scorpion Venoms / Q000494:pharmacology; D012964:Sodium / Q000378:metabolism*; D019831:Sodium-Calcium Exchanger / Q000037:antagonists & inhibitors / Q000378:metabolism*; D019284:Thapsigargin / Q000494:pharmacology; D014407:Tumor Cells, Cultured; D014701:Veratridine / Q000494:pharmacology
D004791:Enzyme Inhibitors; D007476:Ionophores; D012604:Scorpion Venoms; D019831:Sodium-Calcium Exchanger; D002259:Carbonyl Cyanide p-Trifluoromethoxyphenylhydrazone; D015759:Ionomycin; D010042:Ouabain; D019284:Thapsigargin; D014701:Veratridine; D009532:Nickel; D012964:Sodium; D002118:Calcium