Effect of P2RX7 agonists on pCREB in human monocytic cells. (A) Isolated peripheral human blood monocytes were treated with vehicle (2.5 mM HEPES), 100 μM BzATP, or 100 ng/mL LPS for the indicated times. A representative immunoblot displaying pCREB is shown (n=3). The pCREB antibody recognizes pCREB and phosphorylated ATF-1 (pATF1); thus, it is also shown. Immunoblotting was performed with anti-Grb2 antibody as a loading control. (B) THP-1 human monocytes were treated with vehicle (2.5 mM HEPES), 100 μM PMA, 1 mM ATP, or 250 μM BzATP for 15 min. A representative immunoblot displaying pCREB/pATF1 is shown (n=3). Immunoblotting was performed with anti-Grb2 antibody as a loading control. (C) Murine RAW 264.7 macrophages were stimulated with vehicle (2.5 mM HEPES), 250 μM BzATP, or 1 μg/mL LPS for the indicated time-points. Immunoblotting was performed with anti-pCREB/pATF1 and anti-β-tubulin (loading control) antibodies. Results of the three independent experiments were combined and graphed ± sem.