A knock‐in mouse model for KCNQ2‐related epileptic encephalopathy displays spontaneous generalized seizures and cognitive impairment

FIGURE 1. Electroencephalographic (EEG) recordings of the female carrying the p.(Thr274Met) variant at day 9, day 12, day 15, and 3 months of life. The first three EEGs display a suppression‐burst pattern, with periods of high‐amplitude bursts of paroxysmal activity, alternating with periods of flattening of the traces. At 3 months of age, the EEG became continuous, with delta rhythms and no spike

FIGURE 2. A, Schematic representation (not to scale) of the Kcnq2 endogenous, recombined, and Cre‐excised loci. The binding sites of the genotyping primers 284‐Oht and 281‐Oht are shown. Asterisks show the position of Thr274 in exon 3d of the mouse Kcnq2 gene. B, Representative example of a genotyping polymerase chain reaction (PCR) using mouse genomic DNA as a template and primers 284‐Oht and 281‐Oht. PCR without DNA served as a negative control (C−). The results are shown for 10 different animals, including six heterozygous knock‐in mice. Multiplexing is performed with SRY genotyping primers to confirm gender. MWM, Molecular Weight Marker. C, Kernel density estimates computed using R, with the time of spontaneous death as a variable. The graph shows the time of death in 175 knock‐in animals. Ninety‐one percent of spontaneous death in our colony occurred in heterozygous knock‐in animals. D, Kaplan‐Meier survival plot for the two genotypes from postnatal day 0 (P0) to P300

FIGURE 3. A, Schematic representation of a section at lateral +0.60 mm. Colored regions indicate the presence of at least one significant parameter within the brain region at the 0.05 level. White coloring indicates P > .05, and gray indicates that there were not enough data to calculate a P value. The left panel is for males, and the right is for females. B, Histograms for males (left) and females (right) showing variation (decreased, minus scale; increased, positive scale) in areas and lengths expressed as percentage of wild type (WT) together with a color map indicating the significance level. Numbers on the x‐axis correspond to brain regions illustrated in A, and bars are listed in the order of parameters provided in Table S1. C, Illustrating example of Kcnq2 +/+ (WT) and Kcnq2 Thr274Met/+ (knock‐in [KI]) male brain sizes in parasagittal sections double‐stained for Nissl and Luxol. D, Representative image of the corpus callosum enlargement in Kcnq2 Thr274Met/+ male brains shown across a parasagittal section

FIGURE 4. Representative example of the different phases of a spontaneous generalized tonic‐clonic seizure observed in the Kcnq2 Thr274Met/+ mouse model. These images are extracted from a video where a heterozygous knock‐in mouse is housed with a wild‐type animal (Video S3)

FIGURE 5. Prefrontal cortex electrocorticogram of a P32 Kcnq2 Thr274Met/+ mouse during a seizure. Artifacts are caused by violent seizure movements

FIGURE 6. Cognitive differences between Kcnq2 Thr274Met/+ and Kcnq2 +/+ mice. A, Time to reach the platform in the Morris Water Maze task. The Kcnq2 +/+ wild‐type mice (WT) are represented by blue squares, and Kcnq2 Thr274Met/+ mice (KI) are represented by red diamonds. The time is expressed in seconds, and the figure indicates the median value of a group of 10 mice for each block. Plus signs beside the curve indicate the significance of the within‐group, between‐blocks variation (χ2, ++ P < .001). The asterisk beside the left bracket indicates the learning slopes difference (t, *P < .01), and the x beside the right bracket indicates the probe test difference (t, x P < .01). B, Time to enter into the shelter in the Barnes Maze task. Time is expressed in seconds, and the figure shows the median value of a group of 11 mice for each block. Plus signs beside the curve indicate the significance of the within‐group, between‐blocks variation (χ2, +++ P < .001). The asterisk beside the bracket indicates the learning slopes difference (t, * P < .01). C, Number of errors before entering into the shelter in the Barnes Maze task. The median value of the blocks is reported. Plus signs beside the curve…