Cannabinoids inhibit microglial CD40 expression induced by IFN-γ. A, Mouse primary microglial cells were cultured in 6-well tissue-culture plates (5 × 105/well) and treated with THC (0.6 μM), CP55940 (5 μM) or selective cannabinoid CB2 agonist (JWH015; 5 μM) in the presence or absence of IFN-γ (100 U/mL), or treated with vehicle (1% DMSO Control) or IFN-γ alone (100 U/mL); B, In parallel 6-well tissue-culture plates, microglial cells were incubated with IFN-γ (100 U/mL) in the presence or absence of JWH-015 at the indicated doses. After 12 hr-treatments, these cells were prepared for FACS analysis of CD40 expression as described in Materials and methods. For A, ANOVA and post hoc testing showed significant differences of mean fluorescence (+/- SD with n = 3 for each condition) between IFN-γ treatment and IFN-γ treatment in the presence of THC, CP55940 or JWH-015 (p < 0.001). However, there was not a significant difference between IFN-γ/THC and either IFN-γ/CP55940 or IFN-γ/JWH-015 (p > 0.05). For B, ANOVA and post hoc testing showed significant differences of mean fluorescence (+/- SD with n = 3 for each condition) between IFN-γ treatment and IFN-γ treatment in the presence of JWH-015 at 5 μM, 2.5 μM and 1.25 μM (** p < 0.001). C, Western blot analysis by anti-mouse CD40 antibody shows CD40 protein expression and, by anti-β-actin antibody, shows β-actin protein (internal reference). D, Densitometric quantification of Western immunoblotting analysis from independent experiments (n = 2 for IFN-γ; n = 3 for IFN-γ/JWH-015 treatment) indicated that doses of JWH-015 of 1.25 μM or greater significantly (** p < 0.05) reduced IFN-γ-induced CD40 expression. CD40 expression is shown normalized to β-actin.