Mutations in the glycine-binding site in the GluN1 subunit alter the surface expression and desensitisation kinetics of GluN1/GluN3A receptors. (a) Schematic representation of the glycine-binding site in the GluN1 subunit (PDB code: 1PB7); the amino acid residues studied here, and a glycine molecule, are shown. (b) Representative images of COS-7 cells transfected with the indicated wild-type or mutant GluN1-4a subunits together with GFP-GluN3A (GluN3A) subunit and labelled 24 h after transfection. (c) Summary of the relative surface expression of the indicated GluN subunits measured using fluorescence microscopy (n ≥ 24 cells per group); *p < 0.05 vs. GluN1-4a/GluN3A (ANOVA). (d) Quantification of relative surface expression of the indicated GluN1-4a/GFP-GluN3A (GluN1-4a/GluN3A) receptors using quantitative colorimetric assay is shown (n = 6); *p < 0.05 relative to GluN1-4a/GluN3A (ANOVA). (e) Representative whole-cell patch-clamp recordings from HEK293 cells transfected with the indicated wild-type or mutant GluN1-4a/GluN3A receptors. Currents were elicited by applying glycine at the indicated concentrations. (f) Summary of the τw of desensitisation for the indicated GluN1-4a/GluN3A receptors (n ≥ 5 cells per group). (g) Peak concentration-response curves for the indicated GluN1-4a/GluN3A receptors. Each data point represents the mean relative current from ≥5 independent cells. The EC50 values and Hill coefficients are listed in Table 1.