Endoplasmic reticulum Ca2+ signaling and mitochondrial Cyt c release in astrocytes following oxygen and glucose deprivation.

J. Neurochem. (2010) 114, 1436–1446. Abstract In the present study, we investigated changes of cytosolic Ca2+([Ca2+]cyt), endoplasmic reticulum Ca2+([Ca2+]ER) and mitochondrial Ca2+(Ca2+m) in astrocytes following oxygen/glucose deprivation and reoxygenation (OGD/REOX). Two hours OGD did not cause changes in [Ca2+]cyt, but led to a significant increase in [Ca2+]ER. The elevation in [Ca2+]ER continued and reached a peak level (130 ± 2 μM) by 90 min REOX. An abrupt release of Ca2+ER occurred during 1.5–2.5 h REOX, which was accompanied with a delayed and sustained rise in [Ca2+]cyt. Moreover, Ca2+m content was increased significantly within 15 min REOX followed by a secondary rise (∼4.5-fold) and a release of mitochondrial cytochrome c (Cyt c). Astrocytes exhibited translocation of Cyt c from mitochondria to endoplasmic reticulum (ER) and up regulation of ER stress protein p-eIF2α. Blocking Na+-K+-Cl− cotransporter isoform 1 activity, either by its potent inhibitor bumetanide or genetic ablation, abolished release of ER Ca2+, delayed rise in [Ca2+]cyt and Ca2+m. Inhibition of the reverse mode operation of the Na+/Ca2+ exchanger significantly attenuated OGD/REOX-mediated Cyt c release. In summary, this study illustrates that OGD/REOX triggers a time-dependent loss of Ca2+ homeostasis in cytosol and organelles (ER and mitochondria) in astrocytes. Collective stimulation of Na+-K+-Cl− cotransporter isoform 1 and reverse mode function of Na+/Ca2+ exchanger contributes to these changes.