Fig. 5

TNFα induces calcium influx via TNFα receptor 1 (TNFR1). a TNFα induced increase in intracellular free Ca²⁺, as indicated by the change in Fura2 emission. This increase was blocked in Ca²⁺-free medium (0 Ca²⁺ + 0.5 mM EGTA, green), and the intracellular calcium response profile was changed by neutralizing TNFR1 antibody (TNFR1 ab, red). IgG antibody (IgG ab, blue) was used as a control. Arrow denotes addition of TNFα (15 ng/ml) to the astrocyte cultures. The Ca²⁺ ionophore, A23187-Br (10 μM) was added at the end of each experiment (arrow head) to provide a calibration point. b TNFα-induced intracellular free Ca²⁺ (Fura2 ratio) increase was prevented in Ca²⁺-free medium but was not affected in cells pre-treated with thapsigargin (2 μM). ^* p < 0.05 compared to control at each time point; ^# p < 0.05 compared to TNFα at each designated time point, n = 5–6. c, d TNFR1 neutralizing antibody (TNFR1 ab) delayed calcium influx rise onset and reduced the influx plateau time, but the calcium influx maximum of the level of intracellular free Ca²⁺ (Fura2 ratio) was not affected between TNFR1 ab and IgG ab-pre-treated TNFα-stimulated cells. ^* p < 0.05, n = 5. e PARP-1 activation profile detected by PAR formation was also changed when TNFR1 was blocked. TNFR1 neutralizing antibody reduced the length and level of PAR formation, mimicking effects of PARP inhibitor. ^* p < 0.05, n = 3