PERTURBATION OF INTRACELLULAR CALCIUM AND HYDROGEN-ION REGULATION IN CULTURED MOUSE HIPPOCAMPAL-NEURONS BY REDUCTION OF THE SODIUM-ION CONCENTRATION GRADIENT

Na+-Ca2+ exchange has been identified as a mechanism for regulation of intracellular Ca ion concentration ([Ca2+](i)) in neurons of inverteb rates and vertebrates, but for mammalian central neurons its role in r estoration of resting [Ca2+](i) after transient increases induced by s timulation has been less clear. We have examined the recovery of [Ca2](i) following K+ depolarization and glutamate receptor activation of cultured mouse hippocampal neurons using the Ca2+-sensitive dye Fura-2 . Reduction of the transmembrane Na+ gradient by removal of external N af slowed the recovery of neurons from imposed Ca2+ loads. We observed that [Ca2+](i) regulation was disrupted more severely when N-methyl-D -glucamine (N-MG), Tris, or choline rather than Li+ replaced external Na+. Additional disruption of intracellular pH regulation by substitut es other than Li+ may account for this difference. Measurement of [Ca2 +],and [H+],(using the H+-sensitive dye BCECF) during glutamate recept or activation indicated that Ca2+ influx resulted in production of int racellular H+, and that Li+ but not N-MG could prevent cytoplasmic aci dification on removal of external Na+. We also observed that intracell ular acidification alone was sufficient to slow recovery from Ca2+ loa d. We conclude, therefore, that Na+-Ca2+ exchange contributes to recov ery of [Ca2+](i) after stimulation leading to Ca2+ entry into hippocam pal neurons, and that Na+-H+ exchange limits the acidification (and se condary increase in [Ca2+](i)) that accompanies Ca2+ influx. We sugges t that because both Na+-Ca2+ and Na+-H+ exchangers will be compromised during ischemia and hypoglycemia, increased intracellular HC may syne rgize with cytoplasmic Ca2+ to potentiate excitotoxic neuronal death.