Fig. 4.

Direct activation of P2X7R triggers PC12 cell apoptosis and phosphorylation of p38 and JNK MAPKs. A) Viability of PC12 cells was assessed after 24 h incubation with BzATP (1 mM) in the presence or absence of the calcium chelator BAPTA.AM (1 μM), p38 inhibitor SB203580 (1 μM), the JNK inhibitor SP600125 (0.1 μM) and the selective caspase 3 inhibitor z-DVED-fmk (25 μM) The values represent the mean ± SD (n = 3–5 with 8 replicates). *p < 0.001 vs control by one-way ANOVA followed by Holm-Sidak's multiple comparison test. B–C) PC12 cells were cultured with BzATP before protein was harvested at the indicated times and processed for quantitative infrared western blot. B) Blots were stained for phospho-p38 (P-p38) and actin (red), and total p38 (green). MW: molecular weight marker. Bottom graph, quantitation of the infrared fluorescence signal ratio of P-p38 versus total p38. The columns represent the mean ± SD (n = 4, individual values show as dots). Control (Ctr): Untreated cells. *p < 0.02 vs. control by Kruskal-Wallis test followed by Dunn's multiple comparison test. C) Blots were stained for phospho-JNK 54 (PJNK54), phospho-JNK 46 (PJNK 46), and GADPH (red), and total JNK 56 and JNK 46 (green). Bottom graph, quantitation of the infrared fluorescence signal ratio of PJNK 54 versus total JNK 54. No changes were detected in the phosphorylation of JNK 46 at any timepoint. The columns represent the mean ± SD (n = 5, individual values show as dots). Control (Ctr): Untreated cells. *p < 0.02 vs. control by Kruskal-Wallis test followed by Dunn's multiple comparison test. (For interpretation of the references to colour in this figure legend, the reader is referred to the Web version of this article.)