In this study we used fluorescence imaging techniques to characterize the effects of an acute short-term hypoxia on intracellular [Ca2+] signalling in primary cultures of astrocytes from the rat brain stem, midbrain, hippocampus and cortex. It was found that astrocytes respond with strong elevation in intracellular [Ca2+] when PO2 in the incubation media decreases below 15 mmHg. Removal of external Ca2+ had no effect on hypoxia-evoked [Ca2+]i signalling in astrocytes, while application of SERCA inhibitor thapsigargin completely blocked the responses. Cellular mechanism of PO2 sensing involves activation of the PLC pathway as demonstrated in the experiments showing blockade of the hypoxia-induced responses by various inhibitors of PLC downstream processes (U73122, 2-APB or Xestospongine C). Hypoxia-induced [Ca2+]i responses were found to be completely blocked by depolarization of astroglial mitochondria using uncoupler FCCP and, therefore, appear to be dependent on the mitochondrial membrane potential. Ca2+ responses induced in astrocytes by decreases in PO2 lead to fusion of ATP and polyphosphate containing vesicular compartments. These data indicate that astrocytes are sensitive to physiological changes in PO2 and this sensitivity may have a functional significance in the control of local blood flow and the neuronal activity.
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