Evidence for a Na+/Ca2+ exchanger in neuroblastoma x glioma hybrid NG108-15 cells.
To determine whether NG108-15 cells contain a functional Na+/Ca2+ exchanger, we isotonically replaced extracellular Na+ with N-methyl-D-glucamine (NMG) and measured the effect on cytosolic Ca2+ concentration ([Ca2+]i) using the fluorescent Ca2+ indicator fura 2. Replacement with NMG alone had no effect on basal [Ca2+]i or the rise in [Ca2+]i evoked by 80 mM K+ or 10 microM bradykinin, but caused a larger [Ca2+]i increase when thapsigargin and carbonyl cyanide p-trifluoromethoxyphenylhydrazone (FCCP) were added to the cells; this enhanced [Ca2+]i increase could be reversed by adding Na+ back to the bathing buffer. The elevation in [Ca2+]i induced by thapsigargin and FCCP was inversely proportional to extracellular Na+ concentration. Furthermore, the exchanger operated in the reverse mode, as measured by either [Ca2+]i change or 45Ca2+ uptake. An 810 bp cDNA fragment of the exchanger was amplified by PCR; it differed by a single amino acid residue from the corresponding segment of the rat brain Na+/Ca2+ exchanger. These data suggest that a functioning Na+/Ca2+ exchanger exists in NG108-15 cells.
Hsu
L S
LS
Department of Biochemistry, National Defense Medical Center, Taipei, Taiwan, Republic of China.
Chou
W Y
WY
Chueh
S H
SH
D001483:Base Sequence; D002118:Calcium / Q000378:metabolism; D000252:Calcium-Transporting ATPases / Q000037:antagonists & inhibitors; D002259:Carbonyl Cyanide p-Trifluoromethoxyphenylhydrazone / Q000494:pharmacology; D002352:Carrier Proteins / Q000037:antagonists & inhibitors / Q000378:metabolism*; D017931:DNA Primers; D018076:DNA, Complementary; D005910:Glioma / Q000378:metabolism*; D006822:Hybrid Cells; D008969:Molecular Sequence Data; D009447:Neuroblastoma / Q000378:metabolism*; D012964:Sodium / Q000378:metabolism; D019831:Sodium-Calcium Exchanger; D013729:Terpenes / Q000494:pharmacology; D019284:Thapsigargin; D014407:Tumor Cells, Cultured
D002352:Carrier Proteins; D017931:DNA Primers; D018076:DNA, Complementary; D019831:Sodium-Calcium Exchanger; D013729:Terpenes; D002259:Carbonyl Cyanide p-Trifluoromethoxyphenylhydrazone; D019284:Thapsigargin; D012964:Sodium; D000252:Calcium-Transporting ATPases; D002118:Calcium