Intracellular calcium dynamics and cellular energetics in ischemic NG108-15 cells studied by concurrent P-31/F-19 and Na-23 double-quantum filtered NMR spectroscopy

The role of voltage-sensitive Ca2+ channels in mediating Ca2+ influx during ischemia was investigated in NG108-15 cells, a neuronal cell line that does not express glutamate-sensitive receptor-mediated Ca2+ channels, Concurrent P-31/F-19 and Na-23 double-quantum filtered (DQF) NMR spectra were used to monitor cellular energy status, intracellular [Ca2+] ([Ca2(+)](i)), and intracellular Na+ content in cells loaded with the calcium indicator 1,2-bis-(2-amino-5-fluorophenoxy)ethane-N,N,N',N'-tetraacetic acid (5FBAPTA) during ischemia and reperfusion, Cells loaded with 5FBAPTA were indistinguishable from unloaded cells except for small immediate decreases in levels of phosphocreatine (PCr) and ATP. Ischemia induced a steady decrease in intracellular pH and PCr and ATP levels, and a steady increase in intracellular Na+ content; however, a substantial increase in [Ca2+](i) (about threefold) was seen only following marked impairment of cellular energy status, when PCr was undetectable and ATP content was reduced to 55% of control fevers. A depolarization-induced increase in [Ca2+](i) could be completely blocked by 1 mu M nifedipine, whereas up to 20 mu M nifedipine had no effect on the increase in [Ca2+](i) seen during ischemia. These data demonstrate that voltage-gated Ca2+ channels do not mediate significant Ca2+ flux during ischemia in this cell line and suggest an important role for Ca-i(2+) stores, the Na+/Ca2+ antiporter, or other processes linked to cellular energy status in the increase in cytosolic Ca2+ level during ischemia.