Patients with Systemic lupus erythematosus (SLE) experience various peripheral and central nervous system manifestations including spatial memory impairment. A subset of autoantibodies (DNRAbs) cross-react with the GluN2A and GluN2B subunits of the NMDA receptor (NMDAR). We find that these DNRAbs act as positive allosteric modulators on NMDARs with GluN2A-containing NMDARs, even those containing a single GluN2A subunit, exhibiting a much greater sensitivity to DNRAbs than those with exclusively GluN2B. Accordingly, GluN2A-specific antagonists provide greater protection from DNRAb-mediated neuronal cell death than GluN2B antagonists. Using transgenic mice to perturb expression of either GluN2A or GluN2B in vivo, we find that DNRAb-mediated disruption of spatial memory characterized by early neuronal cell death and subsequent microglia-dependent pathologies requires GluN2A-containing NMDARs. Our results indicate that GluN2A-specific antagonists or negative allosteric modulators are strong candidates to treat SLE patients with nervous system dysfunction.

Highlights d NMDARs transition through two distinct structural conformations before opening d The efficiency of NMDAR opening depends on which intermediate pathway is entered d Fast NMDAR opening requires glutamate-induced displacement of the GluN2 pre-M1 helix d Glutamate-induced displacement of the S2-M4/M4 axis also facilitates rapid channel gating

No abstract

Patients with Systemic lupus erythematosus (SLE) experience various peripheral and central nervous system manifestations including spatial memory impairment. A subset of autoantibodies (DNRAbs) cross-react with the GluN2A and GluN2B subunits of the NMDA receptor (NMDAR). We find that these DNRAbs act as positive allosteric modulators on NMDARs with GluN2A-containing NMDARs, even those containing a single GluN2A subunit, exhibiting a much greater sensitivity to DNRAbs than those with exclusively GluN2B. Accordingly, GluN2A-specific antagonists provide greater protection from DNRAb-mediated neuronal cell death than GluN2B antagonists. Using transgenic mice to perturb expression of either GluN2A or GluN2B in vivo, we find that DNRAb-mediated disruption of spatial memory characterized by early neuronal cell death and subsequent microglia-dependent pathologies requires GluN2Acontaining NMDARs. Our results indicate that GluN2A-specific antagonists or negative allosteric modulators are strong candidates to treat SLE patients with nervous system dysfunction.

NMDA receptors (NMDAR) are glutamate-gated ion channels that mediate the majority of fast excitatory synaptic transmission in the nervous system. A central feature of NMDAR physiology is the opening of the ion channel driven by presynaptically-released glutamate. Using glutamate applications to outside-out patches containing a single NMDAR in the continuous presence of the co-agonist glycine, we find that agonist-bound receptors transition to the open state via two conformations, an 'unconstrained pre-active' state that can rapidly transition to the open state and contributes to synaptic events, and a 'constrained pre-active' state that requires more energy and hence time to open and does not contribute to fast signaling. To define how agonist binding might drive these conformations, we decoupled the ligand-binding domains from specific transmembrane segments for the GluN1 and GluN2A subunits. Displacements of the central pore-forming M3 segments define the energy of fast channel opening. However, to enter the unconstrained conformation and contribute to fast signaling, a peripheral helix, the GluN2 pre-M1, must be displaced before the M3 segments move. This pre-M1 displacement is facilitated by the flexibility of another nearby peripheral element, the GluN1 and GluN2A S2-M4. We conclude that peripheral structural elements -pre-M1 and S2-M4 -work in concert to remove constraints and prime the channel for rapid opening, thus facilitating fast synaptic transmission. 214 words. McDaniel et al., 2020). The S2-M4 linker/M4 segment also regulates gating (Amin et al., 2017;Yelshanskaya et al., 2017;Shi et al., 2019). Still, the energetics and timing of these non-pore forming elements to receptor gating remains incomplete.Here, we address the gating mechanism in NMDARs. At most synapses, NMDARs are obligate hetero-tetramers composed of two GluN1 and two GluN2 (A-D) subunits. To define conformational changes between agonist binding and pore opening, we rapidly applied glutamate to outside-out patches containing single NMDARs and assessed the time between when glutamate was first applied to when the channel first opened ('latency to 1 st opening', see Materials and Methods) (Aldrich et al., 1983). With this approach, we identified that wild type GluN1/GluN2A NMDARs, in the transition from agonist-bound closed to the open state, transitions through one of two pre-active states: an 'unconstrained' state that can rapidly transition to the open state and contributes to fast synaptic signaling, and a 'constrained' state that slowly transitions to the open state, presumably due to higher energy requirements, and does not contribute to fast signaling. Further, by decoupling the LBD from the TMD for each specific LBD-TMD linker (S1-M1, M3-S2, and S2-M4, Figure 1B & 1C) (Niu et al., 2004;Kazi et al., 2014), we show that the non-pore forming linkers uniquely influence pore-opening by altering the stability of these pre-active conformations. Thus, our results highlight the temporal and coordinated movements from agonist binding to io...