Fig. 1: Differential sensitivity of N2A- and N2B-containing NMDARs to DNRAbs.
a, b Direct DNRAb application to NMDARs. Upper panels, whole-cell currents from HEK293 cells expressing human NMDAR subunits, either hGluN1/hGluN2A (a) or hGluN1/hGluN2B (b) at 10 µg/mL. Currents were elicited by a 2.5 s application of glutamate (1 mM) in the continuous presence of glycine (0.1 mM) (holding potential, −70 mV). Lower panels, control antibody B1 (IgG1, gray circles) or human-derived DNRAb G11 (green circles) were added 75 s after a baseline recording of five sweeps, and were included in the bath throughout the remaining period. Current amplitudes for individual recordings were normalized to its baseline. Values are mean ± SEM (hN2A + B1, n = 6; hN2A + G11, n = 6; hN2B + B1, n = 5; hN2B + G11, n = 5; n = cells recorded). Example traces in upper panels show the +G11 recordings for the initial sweep during baseline (no antibody present) or for the last sweep during steady-state (in antibody). Ca2+ is not present in the extracellular solution to minimize run-down over time. c, d Peak current amplitudes in N2A-containing NMDAR are more strongly potentiated than those in N2B-containing receptors. Bar graphs (mean ± SEM with dots indicating individual values) (from left to right for hN1/hN2A, n = 6, 6, 6, 6, 5, 5 cells recorded; and for hN1/hN2B, n = 5, 5, 6, 5 cells recorded) showing normalized steady-state peak current amplitudes either for control antibody (B1) or DNRAbs (G11). Significance of DNRAb values are measured relative to their respective control (*p < 0.05 or **p < 0.01, two-sided t test) (left to right for N2A, p = 0.00137, 0.00932, 0.00906; left to right for N2B, p = 0.855, 0.0163). nt, not tested.
