Fig. 5
NMDA-CB2R heteromer levels and functionality in APPSw/Ind neurons. A–F Expression of NMDA-CB2R heteromers in mouse primary neurons (A, B) and microglia (D, E) of wild type (A, D) and APPSw/Ind transgenic mice (B, E) as determined by PLA (see Materials and Methods) using specific primary antibodies against the GluN1 subunit and against the CB2R receptor. Confocal microscopy images (stacks of 3 consecutive panels) show heteroreceptor complexes as red clusters over Hoechst-stained nuclei (blue). Three independent experiments were performed using, for each condition, 5 preparations per session. Bar graphs show the amount of red dots/cell in APPSw/Ind mice and control animals (*p < 0.05, **p < 0.01; Student’s t test versus the control condition). G–L Primary neurons from control and APPSw/Ind mice were treated with selective agonists (15 μM NMDA for NMDA channel, and/or 100 nM JWH-133 for CB2R) and cAMP levels (G, H), ERK1/2 phosphorylation (I, J), and DMR (K, L) assays were determined. When indicated cells were pretreated with selective receptor antagonists (1 μM MK-801 for NMDA or 1 μM SR-144528 for CB2R). Values are the mean ± S.E.M. of 5 independent experiments performed in triplicates. One-way ANOVA followed by Bonferroni’s multiple comparison post hoc test were used for statistical analysis (*p < 0.05, **p < 0.01, ***p < 0.001 versus forskolin treatment in cAMP determinations or versus vehicle treatment (basal) in ERK phosphorylation assays). ANOVA summary: G F: 14.5, p<0.001; H F: 25.3, p<0.001, I F: 5.6, p<0.001, J 13.2, p<0.001