Cytosolic free calcium concentrations in synaptosomes during histotoxic hypoxia

Altered cytosolic free calcium concentrations (Ca²⁺]_i) accompany impaired brain metabolism and may mediate subsequent effects on brain function and cell death. The current experiments examined whether hypoxia-induced elevations in Ca²⁺]_i are from external or internal sources. In the absence of external calcium, neither KCl depolarization, histotoxic hypoxia (KCN), nor the combination changed Ca²⁺]_i. However, with external CaCl₂ concentrations as small as 13 M, KCl depolarization increased Ca²⁺]_i instantaneously while hypoxia gradually raised Ca²⁺]_i. The combination of KCN and KCl was additive. Increasing external calcium concentrations up to 2.6 mM exaggerated the effects of K⁺ and KCN on Ca²⁺]_i, but raising medium calcium to 5.2 mM did not further augment the rise. Diminishing the sodium in the media, which alters the activity and perhaps the direction of the Na/Ca exchanger, reduced the increase in Ca²⁺]_i due to hypoxia, but enhanced the KCl response. The changes in ATP following K⁺ depolarization, KCN or their combination in the presence of physiological calcium concentrations did not parallel alterations in Ca²⁺]_i, which suggests that diminished activity of the calcium dependent ATPase does not underlie the elevation in Ca²⁺]_i. Valinomycin, an ionophore which reduces the mitochondrial membrane potential, elevated Ca²⁺]_i and the effects were additive with K⁺ depolariration in a calcium dependent manner that paralleled the effects of hypoxia. Together these results suggest that hypoxia-induced elevations of synaptosomal Ca²]_i are due to an inability of the synaptosome to buffer entering calcium.