Figure 5.

Phosphorylation of CREB in human primary PBMC. (A) Time-course with HU308 (1 μM) and vehicle control. 10–50 min time-points are significantly different from time-matched vehicle controls (p < 0.0001–0.0022; two-way RM ANOVA with Holm-Šídák posthoc test). The graph shows mean ± SEM of three independent experiments performed in technical triplicate, each with cells from a separate subject (three subjects in total). (B) p-CREB stimulated by HU308 (1 μM) for 40 min in the presence of G protein inhibitors gallein, NF023, NF449, PTX, or vehicle control; data are normalized to responses in the absence of HU308 (100%); statistical comparisons are to the HU308 response without inhibitors (one-way RM ANOVA with Holm-Šídák posthoc test). (C) p-CREB stimulated by a CB₂-selective agonist HU308 (100 nM) or vehicle control at 40 min in the presence of a CB₂-selective inverse agonist/antagonist SR144528 (SR2, 1 μM), a CB1/CB2 neutral antagonist WIN55212-3 (WIN-3, 5 μM), or vehicle control; data are normalized to vehicle control (100%). Two-way RM ANOVA with Holm-Šídák posthoc test. Graphs B and C show independent experiment means (from technical triplicate) as well as overall mean ± SEM of these three independent experiments each performed with cells from a separate subject (three subjects in total).