Role of Na+-H+ and Na+-Ca2+ exchange in hypoxia-related acute astrocyte death

Cultured astrocytes do not succumb to hypoxia/zero glucose for up to 24 h, yet astrocyte death following injury can occur within 1 h. It was previously demonstrated that astrocyte loss can occur quickly when the gaseous and interstitial ionic changes of transient brain ischemia are simulated: After a 20-40-min exposure to hypoxic, acidic, ion-shifted Ringer (HAIR), most cells died within 30 min after return to normal saline (i.e., "reperfusion"). Astrocyte death required external Ca2+ and was blocked by KB-117943, an inhibitor of reversed Na+-Ca2+ exchange, suggesting that injury was triggered by a rise in [Ca2+](i). In the present study, we confirmed the elevation of [Ca2+](i) during reperfusion and studied the role of Na+-Ca2+ and Na+-H+ exchange in this process. Upon reperfusion, elevation of [Ca2+](i) was detectable by Fura-2 and was blocked by KB-117943. The low-affinity Ca2+ indicator Fura-FF indicated a mean [Ca2+](i) rise to 4.8 +/- 0.4 V,M. Loading astrocytes with Fura-2 provided significant protection from injury, presumably due to the high affinity of the dye for Ca2+. Injury was prevented by the Na+-H+ exchange inhibitors ethyl isopropyl amiloride or HOE-694, and the rise of [Ca2+](i) at the onset of reperfusion was blocked by HOE-694. Acidic reperfusion media was also protective. These data are consistent with Na+ loading via Na+-H+ exchange, fostering reversal of Na+-Ca2+ exchange and cytotoxic elevation of [Ca2+](i). The results indicate that mechanisms involved in pH regulation may play a role in the fate of astrocytes following acute CNS injuries. (C) 2004 Wiley-Liss, Inc.