P2X7 Receptor Activation Induces Reactive Oxygen Species Formation and Cell Death in Murine EOC13 Microglia

Figure 4

P2X7 activation induces ROS formation in EOC13 microglial cells. (Left panels) Adherent DCF-loaded EOC13 cells or (right panels) suspended EOC13 cells in (a) NaCl medium containing 1 mM Ca²⁺ (preincubated in the absence (control) or presence of 10 μM AZ10606120 at 37°C for 15 min), (b) NaCl medium in the absence (control) or presence of 1 mM Ca²⁺, (c) NaCl medium in the absence (control) or presence of 100 μM EGTA, or (d) NaCl or KCl medium were (a–d) incubated in the absence (basal) or presence of 575 μM ATP⁴⁻ (2 mM or 1.4 mM ATP as explained in Section  2.8) at 37°C for (left panels) 15 min or (right panels) 5 min in the presence of 25 μM ethidium⁺. (a–d) Incubations were stopped by the addition of MgCl₂ medium and centrifugation. Mean fluorescence intensities (MFI) of (left panels) DCF (ROS formation) or (right panels) ethidium⁺ uptake (pore formation) were determined by flow cytometry and results shown as means ± SD, 𝑛=3; ***𝑃<0.001 or **𝑃<0.01 compared to corresponding basal; ^†††𝑃<0.001 compared to corresponding ATP.