2002/05/01 by Kara L. Kopper, Joseph S. Adorante
paper · doi:10.1152/ajpcell.00182.2001
In fura 2-loaded N1E-115 cells, regulation of intracellular Ca 2+ concentration ([Ca 2+ ] i ) following a Ca 2+ load induced by 1 μM thapsigargin and 10 μM 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 μM veratridine and 4 μg/ml scorpion venom, the rate of the reverse mode of the Na + /Ca 2+ exchanger was apparently enhanced, since an ∼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 (∼ 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 μM) depolarized N1E-115 cells (∼25 mV). This depolarization was Na + dependent and blocked by 5 mM Ni 2+ and 250–500 μM 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.