To develop a mouse model of anti-N-methyl-D-aspartate receptor encephalitis (NMDARe) that allows comprehensive neuro-immunobiological investigations and the assessment of potential therapies.

Eight-week-old female C57BL/6J mice were immunized with GluN1_356-385 peptide (or saline) along with AddaVax (adjuvant that favors B-cell autoimmunity) and pertussis toxin, followed by systematically examinations for behavioral and neuro-immunobiological changes. Treatment groups received an anti-CD20, a positive allosteric modulator of NMDAR (NMDAR-PAM, SGE-301), or both. GluN1-antibody synthesis, epitope spreading, antibody effects on NMDAR density and function, brain immunological infiltrates, microglial activation, and antibody synthesis by cultured inguinal (ILN) and deep cervical lymph nodes (DCLN) were assessed by immunohistochemistry, calcium imaging, confocal and super-resolution microscopy, electrophysiology, and flow cytometry. Changes in memory and behavior were assessed with a panel of behavioral tests, and clinical/subclinical seizures with brain-implanted electrodes.

Immunized mice, but not controls, developed serum and CSF NMDAR-antibodies, showing epitope spreading and reduced synaptic NMDAR clusters and hippocampal plasticity. Additionally, they had brain-bound antibodies, inflammatory infiltrates (mainly B- and plasma cells), microglia activation, and presence of NMDAR/IgG complexes in microglial endosomes. Cultured DCLN showed NMDAR-antibody synthesis. These findings were associated with psychotic-like behavior, memory deficits, increased seizure susceptibility, and abnormal movements. Treatment with anti-CD20, NMDAR-PAM or both, reversed most neurobiological and behavioral abnormalities. Repopulation of B cells was associated with re-emergence of clinical-neurobiological alterations, which were abrogated by the NMDAR-PAM.

This model offers an all-inclusive neuro-immunobiology of the disease, allowing testing novel treatments, supporting the therapeutic potential of NMDAR-PAM, and suggesting an immunological paradigm of brain NMDAR-epitope spreading, which along the DCLN might contribute to fine-tuning the immune response.