Figure 6.

D318L and L320A mutations abolish the potentiating effect of CK on ATP-induced cell death. Cells were seeded at 5 × 10³ in a 96-well plate 24 h prior to stimulation with a range of ATP concentrations (1 µM–3 mM) or 3 mM ATP + AZ10606120 (10 µM) for a further 24 h. AlamarBlue was added for 2 h prior to measurement of fluorescence (RFU) using a Flexstation 3 plate reader. (A) Data is presented as the percentage mean cell viability compared to control media. (B) Cell viability was determined in the presence of 500 μM ATP + CK compared to 500 μM ATP alone for WT hP2X7 and S59A hP2X7. (C) Cell viability was determined in the presence of 50 μM ATP + CK, 100 μM ATP + CK, and 500 μM ATP + CK compared to each concentration of ATP alone for WT hP2X7, S60A, D318L and L320A hP2X7 mutants.