Fig. 5.

Prolonged hypoxia causes death of astrocytes with greater mortality after hypoxia in high glucose. A – F. Micrographs of astrocyte cultures subjected to the indicated periods of hypoxia and concentrations of glucose followed by 6 h reoxygenation in normal glucose. Hoechst 33462 nuclear stain and Rhodamine B dextran are used to identify cells and indicate membrane break down. D. Significant death occurs after 6h hypoxia in 5 mM glucose and 6 h reoxygenation. E. More death occurs when the hypoxia is in 27 mM glucose. F. The connexin hemichannel blocker Gap26 prevents cell death. G. Cell death quantified as a function of duration of hypoxia, glucose concentration, and time of reoxygenation. H. Connexin hemichannel blockers (Gap26, Gap27, and La³⁺) are protective, as is the p38 MAPK inhibitor, SB202190, applied before the hypoxia. Pannexin hemichannel blockers (¹⁰panx1, E1b, and probenecid (Prob)) are not protective. I. The connexin hemichannel blocker, Gap26, applied during the reoxygenation period reduces coupling in control cells, presumably by preventing formation of new cell-cell channels that would compensate for internalization of existing channels. Gap26 did not affect the reduction in dye coupling produced by 6h hypoxia in high glucose followed by 6 h reoxygenation. * p < 0.05 hypoxia/reoxygenation vs. control, *** p < 0.001 hypoxia/reoxygenation without vs. with addition of indicated connexin hemichannel blockers or SB202190. From Orellana et al., 2010.