Figure 1: Generation of experimental FCD-associated convulsive seizures.
(a) Simplified diagram of the constitutively active Rheb (RhebCA) upstream of mTORC1. (b) Diagram of the electrode placement around an E15±0.5 fetal brain (left). Photograph of a P0 mouse head with GFP fluorescence in the medial prefrontal cortex (mPFC) following electroporation at E15 (right). The ellipse highlights the location of the somatosensory cortex (SSC). (c) Confocal images of pS6 immunostaining (red) and GFP and tdTomato fluorescence (green) in coronal sections from 2-month-old mice containing neurons electroporated with either RhebCA+tdTomatoCAG or GFPCAG alone in the mPFC. Scale bar, 40 μm. (d) Bar graphs of total pS6 immunoreactivity per cell (FU: arbitrary fluorescence unit, N=4 per condition) from data shown in (c). **P<0.01 (Student’s t-test). (e–g) Images of coronal sections (top) containing neurons electroporated with control plasmid in the mPFC (e), with RhebCA in the mPFC (f) or in the SSC (g), and corresponding representative examples of EEG (black) and EMG (grey) traces (bottom). Scale bar, 1 mm. (h) Expanded time window (10 s) view of events found in the EEG trace shown in f, numbered correspondingly to the blue boxes. (i) Bar graphs show the number (#) of seizures/day and seizure duration in animals with seizures (N=8) from the mPFC RhebCA-expressing cohort. Control animals (N=9) electroporated with a control plasmid did not exhibit any seizures. *P<0.05 (Student’s t-test). (j) Three-dimensional reconstruction of mouse brains containing neurons electroporated with either RhebCA+tdTomatoCAG or a control plasmid GFPCAG, both pseudocolored green to ease visualization. ACC, anterior cingulate cortex; CC, corpus callosum; MC, motor cortex. Error bars, s.e.m.
