Fig. 1
Increased polyubiquitination after status epilepticus. a. Representative pictures of coronal brain sections at the level of the dorsal ipsilateral and contralateral hippocampus 24 h after status epilepticus showing FjB-positive cells restricted mainly to the ipsilateral CA3 subfield (arrows and insert). Note, the ipsilateral CA1 and dentate gyrus (DG) and the contralateral hippocampus are mainly spared from cell death. Scale bar, 500 μm b. Representative Western (n = 1 per lane) and graph showing increased polyubiquitination levels (FK2 antibody) in the ipsilateral hippocampus after intra-amygdala KA induced-status epilepticus (mean ± sem, *p <0.05 by one-way ANOVA with Fisher’s post hoc test; n = 4 per group). c. Representative Western (n = 1 per lane) and graph showing increased polyubiquitination levels (FK2 antibody) in the hippocampus after pilocarpine-induced status epilepticus (mean ± sem, *p <0.05 by student’s two-tailed t-test; n = 4 per group). d. Schematic showing transgenic approach to monitor UPS impairment in vivo. UbG76V-GFP transgenic mice over-express a modified green fluorescent protein (GFP) constitutively targeted for ubiquitin-dependent proteasome degradation. Under physiological conditions GFP is immediately degraded by the proteasome and no intracellular accumulation of GFP can be detected; however, if UPS function is compromised, GFP accumulates inside the cell where it can be detected. e. Representative photomicrographs (5x lens) showing subfield-specific increased GFP-reporter accumulation in the ipsilateral hippocampus of transgenic UbG76V-GFP mice subjected to intra-amygdala KA induced status epilepticus. Note, strong GFP signal in the DG and CA1 hippocampal subfield shortly after status epilepticus (4 h and 8 h) and scattered GFP staining throughout the hippocampus involving all subfields 24 h after seizure suppression. Scale bar, 500 μm f. Table showing hippocampal (DG, CA1 and CA3) subfield-specific increase in GFP signal in transgenic UbG76V-GFP mice after status epilepticus. Subfield-specific GFP signal was quantified according to a semi-quantitative scale after staining with GFP antibody (− baseline (absent/very low signal), + increase above baseline, ++ strong increase above baseline; n = 3 per group)