Critical role of sodium in cytosolic [Ca2+] elevations in cultured hippocampal CA1 neurons during anoxic depolarization

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Abstract
Although the extent of ischemic brain damage is directly proportional to the duration of anoxic depolarization (AD), the mechanism of cytosolic [Ca2+] ([Ca2+]c) elevation during AD is poorly understood. To address the mechanism in this study, [Ca2+]c was monitored in cultured rat hippocampal CA1 neurons loaded with a Ca‐sensitive dye, fura‐2FF, and exposed to an AD‐simulating medium containing (in mmol/L): K+ 65, Na+ 50, Ca2+ 0.13, glutamate 0.1, and pH reduced to 6.6. Application of this medium promptly elevated [Ca2+]c to about 30 µmol/L, but only if oxygen was removed, the respiratory chain was inhibited, or if the mitochondria were uncoupled. These high [Ca2+]c elevations depended on external Ca2+ and could not be prevented by inhibiting NMDA or α‐amino‐3‐hydroxy‐5‐methyl‐4‐isoxazolepropionate (AMPA)/kainate receptors, or gadolinium‐sensitive channels. However, they could be prevented by removing external Na+ or simultaneously inhibiting NMDA and AMPA/kainate receptors; 2‐[2‐[4‐(4‐nitrobenzyloxy)phenyl]ethyl]isothiourea methanesulfonate (KB‐R7943), an inhibitor of plasmalemmal Na+/Ca2+ exchanger, partly suppressed them. The data indicate that the [Ca2+]c elevations to 30 µmol/L during AD result from Na+ influx. Activation of either NMDA or AMPA/kainate channels provides adequate Na+ influx to induce these [Ca2+]c elevations, which are mediated by KB‐R7943‐sensitive and KB‐R7943‐resistant mechanisms.