The Effects of Hypoxia/Reoxygenation on the Physiological Behaviour of U373-Mg Astrocytes
This work characterizes the anoxia-evoked changes in the content of bound calcium (Cab) in brain cortex membraneous structures, studied in vivo, on living brain cortex preparation and in vitro, on subcellular fractions (synaptosomes, microsomes and mitochondria) in anoxic conditions. The chlorotetracycline (CTC) fluorescent chelate probe was used to monitor changes of Cab content in hydrophobic domains of intracellular membranes. In in vivo experiments the bioelectric activity of single neurons was recorded simultaneously with measurements of Ca-CTC fluorescence. In vitro experiments were supplemented with determinations of synaptosomal 45Ca-uptake. It was found that the response of cortical neurons to anoxia is manifested in a decrease of a portion of Ca2+ bound with membrane hydrophobic domains. These in vivo changes preceded the noticeable disturbances of neuronal electric activity. An anoxia-evoked drop in Cab was also clearly demonstrated in vitro, irrespective of K+ (for synaptosomes) or Na+ (for mitochondria) concentrations in incubation media, although the additional effect of Cab displacement was noted when Na+/K+ concentrations were modified in order to simulate their changes occurring in anoxic conditions. It was found that the membranes of different neuronal compartments are not uniformly vulnerable to anoxia in vitro as the anoxic decrease in Cab content occurred in synaptosomes and microsomes much sooner than in mitochondria. Therefore, in vivo and in vitro experiments visualized high sensitivity of Ca2+-binding mechanisms in different neuronal membranes to anoxia. The anoxia-evoked displacement of a portion of Cab to the free ionic form may trigger a complex intracellular response determining anoxic reactions and post-anoxic recovery.