Study of the SV2A protein role in Epilepsy. O. Bartholome*1, P.Van den Ackerveken*1, C. Menten1, M.E. Serrano Navacerrada2, V. Neirincks1, G. Becker2,A. Plainevaux2 , S. Wislet1 ,P. Leprince1 and B. Rogister1,3 Laboratory of Nervous System Diseases and Therapy, GIGA-Neuroscience, University of Liège, Liège, Belgium 2. University of Liège, GIGA-Research, Cyclotron Research Center 3. Neurology Department, University Hospital, Liège, Belgium. * These two authors equally contributed to this work Introduction The SV2A protein is a glycoprotein present in the membranes of most synaptic vesicles. Although it is highly conserved during evolution, its physiological role remains largely unknown. However, it has recently been demonstrated that a very effective anti-epileptic drug called levetiracetam (Keppra®) binds to the SV2A protein. As SV2A knock-out animals start to experiment seizures around post-natal day 7 and die in status epilepticus around post-natal day 15, a SV2A role in epilepsy has been suspected but is currently unproved. This project aims to better understand how the SV2A protein may be involved in the occurrence of epilepsy. For this purpose, we used a mouse line that allows the conditional invalidation of SV2A gene in hippocampal region (CA3 and dentate gyrus (DG)) from the postnatal day 15 to the postnatal day 56 (Sv2A-cKO or Grik4:Sv2a-cKO). Results 1. Absence of an epileptic phenotype in the validated Sv2A-cKO model. 3. 2D DIGE experiments of hippocampi proteomes revealed interesting patterns. A B 2'; 3'-cyclic-nucleotide 3'-phosphodiesterase ATP synthase subunit alpha; mitochondrial Aconitate hydratase Alpha-enolase Aspartate aminotransferase; mitochondrial Dihydrolipoyl dehydrogenase; mitochondrial Dihydrolipoyllysine-residue succinyltransferase component of 2-oxoglutarate dehydrogenase complex Dihydropyrimidinase-related protein 5 Fructose-bisphosphate aldolase A Gamma-enolase Glutamine synthetase Glyceraldehyde-3-phosphate dehydrogenase Heat shock cognate 71 kDa protein Heterogeneous nuclear ribonucleoprotein H Isocitrate dehydrogenase [NADP] cytoplasmic Isocitrate dehydrogenase [NAD] subunit alpha; mitochondrial Ketimine reductase mu-crystallin L-lactate dehydrogenase B chain Malate dehydrogenase; cytoplasmic Malate dehydrogenase; mitochondrial NADH dehydrogenase [ubiquinone] 1 alpha subcomplex subunit 10 Protein phosphatase 1 regulatory subunit 7 Pyruvate kinase PKM Stress-70 protein Succinyl-CoA:3-ketoacid coenzyme A transferase 1 Tubulin alpha-1A chain Tubulin beta-1 chain Tubulin beta-2A chain 1. A. Expression levels of SV2a genes determined by qRT-PCR on CA1, CA3 and DG extract. Gene expression was normalized on Gapdh level. Comparable reduction observed in western blot (Data not shown) B. SV2A-cKO adult animals do not exhibit spontaneous seizure (Data not shown) and have not a lowered epileptic threshold after PTZ treatment. 3. A. Several proteins upregulated in the hippocampus of Sv2A-cKO adult animals compared to WT were identified after mass spectrometry analysis and a protein-protein interaction network was build. 2. Absence of a compensation phenomenon by Sv2B and Sv2C. Sv2B Sv2C 2. No significant change in Sv2B or Sv2C transcripts concentrations, two other members of SV2 proteins family (paralogs of SV2A), in hippocampus of Sv2A-cKO animals in comparison with the wild-type (WT). Same observation for proteins expression (Data not shown) 3. B. Several biological functions were enriched in our network, the most significant ones being: Extracellular Exosome, Membrane Bounded Vesicle and Myelin Sheet. Conclusions Our preliminary results show that SV2A-cKO adult animals do not exhibit spontaneous seizure neither exhibit a lower epileptic threshold and do not compensate the absence of SV2A by overexpressing SV2B or C. However, the absence of SV2A in hippocampus seems to modulate the expression of several other proteins involved in a membranous metabolism (exosome and/or vesicles and/or myelin). Ongoing experiments are designed to confirm 2D-DIGE experiment and to investigate functions enriched in our network. Contact : odile.bartholome@ulg.ac.be