Epileptogenic effects of NMDAR antibodies in a passive transfer mouse model (1)
N-methyl D-aspartate receptor (NMDAR) antibody-mediated encephalitis commonly presents with features of psychiatric disturbance, cognitive problems, seizures and encephalopathy, usually followed by movement disorder, autonomic dysfunction, hypoventilation and coma. The NMDAR antibodies are pathogenic, with in vitro and in vivo experiments showing loss of NMDAR, most evident in the hippocampus, and demonstrated also in patient brain tissue post-mortem.
Eight C57BL/6 female mice were implanted with EEG transmitters (Day 1) and injected intracerebroventricularly with IgG from NMDAR antibody Patient 1 (four mice) and healthy Control subject 1 (four mice) on Day 7 and again on Day 14 post-implantation. A small increase in delta power (1–4 Hz) was observed in both groups after the second injection, with no difference between the patient and healthy control groups. Seizure number was higher in the NMDAR antibody-injected mice [median seizures/mouse (range) 10 (2–33), n = 9] compared to the healthy control IgG-injected mice [median seizures/mouse (range) 3 (1–5), n = 6; Mann Whitney P = 0.004]. The more severe stage 3 seizures were seen in all the NMDAR antibody IgG-injected mice compared to only three of six of the healthy control IgG-injected mice (P = 0.04), and the number of stage 3 seizures per mouse was higher in the NMDAR antibody IgG-injected mice [mean ± standard error of the mean (SEM) (range) 7.7 ± 2.8 (1–29)] compared to the healthy control IgG-injected group [0.83 ± 0.4 (0–2); P = 0.003]. Finally, a higher total seizure score per mouse was seen in the NMDAR antibody IgG-injected animals (38.3 ± 9.8 versus 7.5 ± 1.67; P = 0.003).
Using the concurrent video and EEG from the 60 min following the injection of PTZ, segments of EEG were classified according to six metrics and manually assigned to categories for the library. Epileptic ‘spike’ events were not seen in the animals in Experiment 1, or before PTZ seizure induction, but after PTZ the spike events were more numerous in NMDAR antibody IgG-injected mice than in healthy control IgG-injected mice.
Human IgG was easily detected in the hippocampi of NMDAR antibody IgG-injected mice, with little in the healthy control IgG-injected tissue, or in other brain regions where NMDAR expression is lower. There was variability in the IgG fluorescence intensity across the hippocampi but overall the bound IgG was higher in NMDAR- antibody injected mice in each of the three hippocampal regions analysed, particularly in the CA3, and on pooling the data for all three regions (P < 0.001). The seizure scores in NMDAR antibody injected mice correlated with the amount of IgG bound (n = 5) (R2 = 0.8, P = 0.04), with a similar trend for the CA3 regions only (R2 = 0.7, P = 0.07).
A commercial anti-NR1 antibody, which binds an intracellular NMDAR epitope on the NR1 subunit, and required tissue permeabilization; the results were not different between NMDAR antibody IgG- and healthy control IgG-injected brains. The total NMDAR were measured in parallel and were not different between test and control hippocampi. However, as determined on adjacent sections for direct comparison, a proportion of the surface NMDAR was already bound by human IgG with, consequently, a 19.7% reduction in the number of NMDAR free of bound IgG (P = 0.035).
