Fig. 1

The NMDA receptor interacts with the cannabinoid CB₂ receptor in a heterologous expression system. A–C Immunocytochemistry performed in HEK-293T cells expressing CB₂-YFP (A) (1 μg cDNA), that was detected by its own fluorescence (green), and GluN1-Rluc receptor (1 μg cDNA) (B), that was detected by a mouse monoclonal anti-Rluc antibody and a secondary Cy3-conjugated anti-mouse IgG antibody (red). Colocalization is shown in yellow (C). Cell nuclei were stained with Hoechst (blue). Images are taken near the bottom of the cell, i.e. it mainly includes the membrane in contact with the glass of the plate. Scale bar: 20 μm. D Schematic representation of BRET assay: the occurrence of energy transfer depends on the distance between the BRET donor (Rluc) and the BRET acceptor (YFP). E BRET assays were performed in HEK-293T cells transfected with a constant amount of cDNAs for GluN1-Rluc (0.25 μg), GluN2B (0.15 μg), and increasing amounts of cDNA for CB₂R-YFP (0.25 to 1.25 μg) (left) or, as negative controls, using GHSR_1aR-YFP (0.25 to 1.25 μg) as acceptor  (right top) ), or using  AT₁R-Rluc (0.5 μg) as donor and CB₂R-YFP (0.25 to 1.25 μg) as acceptor (right bottom). Values are the mean ± S.E.M. of 6 independent experiments performed in duplicates