Abstracts

Rationale: Epilepsy is a neurological disorder affecting 2% of the population. Levetiracetam (LEV) is a new antiepileptic drug that binds to presynaptic vesicular protein SV2A; however, the mechanism of action remains unknown. Abnormal presynaptic release of glutamate has been considered one of the seizure-induced alterations in epilepsy. Previous studies have shown abnormally enhanced vesicular release in hippocampal presynaptic boutons in epilepsy. Here, we investigate if chronic treatment with LEV will modify abnormal presynaptic vesicle release and synaptic vesicle proteins (SV2A, SV2B, SV2C) and transcripts expression after pilocarpine-induced status epilepticus. Methods: Different groups of control and epileptic mice were treated with LEV (experimental, n=7) or saline solution (control, n=6). To induce chronic epilepsy, the pilocarpine model of temporal lobe epilepsy was developed in synaptopHluorin (SpH)-expressing transgenic mice. Protein samples were extracted and immoblottings were developed to assess expression of SV2A, SV2B, and SV2C. Gene expression was analyzed using TaqMan real time quantitative PCR assays. Imaging of electrically-evoked release was performed by confocal imaging in brain slices. Time-lapsed images were obtained from the CA3 stratum lucidum in hippocampus. For this analysis, we included 92 synaptic boutons from 7 slices in the control non-treated group, 148 boutons from 12 slices in control (treated with LEV), 115 boutons from 7 slices in epileptic (non-treated group) and 100 boutons from 6 slices in epileptic group chronically treated with LEV. Results: As previously reported, epileptic SpH mice exhibited an increase in vesicular release when compared to control group. In contrast, LEV-treated SpH epileptic mice exhibited a reduction in release when compared to control animals treated with same. SV2A protein expression was down-regulated 22.9% in epileptic mice but after treatment with LEV SV2A expression increased 40% above controls. In contrast, sv2a transcripts were not upregulated in LEV-treated status epilepticus group.No significant changes were detected in sv2b gene expression among the groups. However, a significant change was observed for sv2c gene expression (ANOVA, p>0.0001, F=11.87) where status epilepticus induced a significant sv2c upregulation when compared to saline injected control group. Interestingly, in contrast to SV2C protein expression, chronic treatment with LEV induced a significant 33.8% and 11.7% reduction of sv2c levels in treated status epilepticus and control groups respectively when compared to saline-injected status epilepticus and control groups. Conclusions: Levetiracetam partially corrected abnormal SV2A and SV2C expression and inhibited abnormally enhanced vesicular release in epileptic SpH mice indicating that the anti-epileptic action and effectiveness in chronically epileptic tissue may be associated with changes in the presynaptic vesicular release machinery and its own pharmacological targets in mesial temporal lobe epilepsy.