Brain Malformations in C57BL/6 Mice Affect Seizure Onset
Authors: Keever Katherine M. , Li Ying , Womble Paige D. , Sullens D. Greg , Lugo Joaquin N. , Ramos Raddy L. 
Published: May 2022

Background

Brain malformations are known to cause cognitive delay and seizures in both humans and mice. It has been demonstrated that the neocortex and cerebellum of the brain are vulnerable to malformations affecting neuronal lamination and synaptic connectivity. Our objective was to determine if the presence of sporadic neocortical and/or cerebellar brain malformation affected time to seizure onset in a large cohort of C57BL/6 mice, as this strain is widely used in neuroscience and epilepsy research. We hypothesized that the presence of cerebellar and/or neocortical malformations in mice would reduce seizure onset in both male and female mice.

Methods

Flurothyl was used to induce seizures in C57BL/6J mice, and the time to seizure onset was recorded. Following seizures, mice were euthanized and brains were placed in 4% paraformaldehyde and then into 30% sucrose. Brains underwent cryosectioning at 45um thickness, with every other coronal forebrain section and 15 cerebellum sagittal sections mounted on gelatin‐coated slides. After Nissl staining, high and low power brightfield microscopy were used to screen the stained slides for presence of cerebellar and neocortical malformations. Only malformations present in 2 or more sections in the neocortex or cerebellum were classified as a malformation. Statistical analysis was performed using two‐tailed independent t‐tests with significance at p < 0.05.

Results

Out of 132 mouse brains, 82 (62.1%) had either neocortical and/or cerebellar malformations and 49 (37.71%) had no malformations. Of 62 female mice, 40 (64.5%) had malformations, and 22 (35.48%) did not have malformations. Of 69 male mice, 42 (67.74%) had malformations, and 27 (43.54%) did not have malformations. In the female malformations group, 7 of 40 (17.5%) were exclusively neocortical, 26 of 40 (65%) were cerebellar, and 7 of 40 (17.5%) had both neocortical and cerebellar malformations. Statistically significant differences in seizure onset were found in the female cohort between those with malformations and without malformations (p = 0.016). Comparison of female mice with both cerebellar and neocortical malformations to female mice with no malformations also demonstrated statistically significant differences (p = 0.017). For the male malformations cohort, 7 of 42 (16.67%) were neocortical, 31 of 42 were cerebellar, and 4 of 42 (9.52%) had both neocortical and cerebellar malformations. Comparison of male mice with only neocortical malformations to those with no malformations yielded significant results (p = 0.04). Differences in time to seizure onset were also found when comparing male mice with only cerebellar malformations to those with only neocortical malformations (p = 0.016).

Conclusions

Significant differences in seizure onset time were present in both genders of C57BL/6 mice with either cerebellar and/or neocortical malformations. This raises important questions regarding pre‐existing malformations in mice used to study diseases such as epilepsy. For example, sporadic malformations influencing seizure onset could affect the efficacy of anti‐seizure medications being researched.