Simvastatin Restrains Exacerbation of Epilepsy Figure 2. Immunostaining of GFAP for reactive astrocytes in hippocampus at 4, 5 and 6 months post-lesion. (A1–A2) In the normal brain, GFAP staining (brown) showed slim astrocyte morphology and HE staining showed normal neuron (blue) number. Higher magnification of insert was presented in A2. (B1–B2) Simvastatin-treated group exhibited astrocytic hypertrophy and moderate reduction in neurons. (C1–C2) Animals with KA-lesion followed by saline administration showed pronounced up-regulation of GFAP expression. GFAP-positive astrocytes exhibited hypertrophy and atrophied processes. The number of neuron was also severely reduced. (D–E) Quantification of GFAP-positive cells in DH area of hippocampus. Scale bar = 400 mm in C1 (applies to A1, B1, C1); scale bar = 100 mm in C2 (applies to A2, B2, C2). (* P,0.05, vs. the control group; # P,0.05, vs. the saline-treated group). doi:10.1371/journal.pone.0024966.g002 greater than that of the saline-treated group, which persisted over time. These data suggest that simvastatin treatment can rescue the loss of neurons in CA3 and DH region. Simvastatin Treatment Restrains KA-lesion Induced Mossy Fiber Sprouting (MFS) Mossy fibers from granule cells in the DH undergo reorgani- zation of their terminal projections in both human epilepsy and animal models of epilepsy. In this study, KA lesion-induced aberrant MFS was shown by Timm’s method that selectively labeled synaptic terminals of mossy fibers due to their high zinc content. The effect of saline and simvastatin-treatment on KA- induced MFS was assessed at 6 months post-lesion. In the saline- treated group, Timm’s staining showed robust MFS that extended into the dentate supragranular layer (DSGL) (Fig. 4A1, A2). On the contrary, in the simvastatin-treated group (Fig. 4B1, B2), supragranular MFS was less intense and more dispersed, though it is still more dense than that in the normal brain (Fig. 4C1, C2). We further compared the average width and Timm’s staining density between simvastatin- and saline-treated groups. We found that simvastatin-treatment reduced MFS width and staining density in all upper blade (UB), lower blade (LB) and crest area of DH. These data suggest that simvastatin-treatment restrained KA-induced abberant MFS. Simvastatin Inhibits Abnormal Spike Discharges of the Epileptic Brain as Observed by EEG Monitoring The activity from the hippocampus was monitored with electronic encephalography (EEG) during 4 to 6 months after KA-lesion. Abnormal spikes in EEG showed high amplitude and sharp peak and they were manifested by asterisks (Fig. 5A–C). The EEG data from hippocampus of rats consisted of waves with a frequency ranging from 0 to 75 Hz and amplitude of 500 mV. Quantitative data showed the comparison of EEG data at 4, 5, and 6 months after KA-lesions. Simvastatin-treated group presented a significant lower frequency (4 months: 24.867.2/ min, 5 months: 28.164.2/min, 6 months: 31.065.1/min) of abnormal spikes than the saline-treated group (4 months: PLoS ONE | www.plosone.org 3 September 2011 | Volume 6 | Issue 9 | e24966