ALKALINIZATION PROLONGS RECOVERY FROM GLUTAMATE-INDUCED INCREASES IN INTRACELLULAR CA2+ CONCENTRATION BY ENHANCING CA2+ EFFLUX THROUGH THE MITOCHONDRIAL NA+ CA2+ EXCHANGER IN CULTURED RAT FOREBRAIN NEURONS/

Increasing extracellular pH from 7.4 to 8.5 caused a dramatic increase in the time required to recover from a glutamate (3 mu M, for 15 s)-i nduced increase in intracellular Ca2+ concentration ([Ca2+](i)) in ind o-1-loaded cultured cortical neurons. Recovery time in pH 7.4 HERES-bu ffered saline solution (HBSS) was 126 +/- 30 s, whereas recovery time was 216 +/- 19 s when the pH was increased to 8.5, Removal of extracel lular Ca2+ did not inhibit the prolongation of recovery caused by incr easing pH. Extracellular alkalinization caused rapid intracellular alk alinization following glutamate exposure, suggesting that pH 8.5 HBSS may delay Ca2+ recovery by affecting intraneuronal Ca2+ buffering mech anisms, rather than an exclusively extracellular effect. The effect of pH 8.5 HBSS on Ca2+ recovery was similar to the effect of the mitocho ndrial uncoupler carbonyl cyanide p-(trifluoromethoxyphenyl) hydrazone (FCCP; 750 nM). However, pH 8.5 HBSS did not have a quantitative effec t on mitochondrial membrane potential comparable to that of FCCP in ne urons loaded with a potential-sensitive fluorescent indicator, 5,5',6, 6 loro-1,1',3,3'-tetraethylbenzimidazolocarbocyanine iodide (JC-I). We found that the effect of pH 8.5 HBSS on Ca2+ recovery was completely inhibited by the mitochondrial Na+/Ca2+ exchange inhibitor CGP-37157 ( 25 mu M). This suggests that increased mitochondrial Ca2+ afflux via t he mitochondrial Na+/Ca2+ exchanger is responsible for the prolongatio n of [Ca2+](i) recovery caused by alkaline pH following glutamate expo sure.