Abstracts

Rationale: Brivaracetam (BRV) is a novel synaptic vesicle protein 2A (SV2A) ligand, 10 fold more potent than levetiracetam (LEV) in animal models of epilepsy. Previously, we demonstrated that LEV decreases synaptic transmission in a time- and stimulation frequency-dependent manner and vesicular entry may mediate the effect of LEV. In the present study, we examine if the effect of LEV on the synaptic response is reproducible by BRV and if BRV. Methods: Hippocampal slices (500 μm, 3-4 wk old SD male rats) were maintained in a submerged perfusion chamber with bipolar stimulation of Schaffer collateral fibers and extracellular recordings from the dendritic layer of CA1. We examined BRV action at 1, 3, 10 and 30 µM concentrations applied for either 30 minutes or 3 hours prior to stimulation (N=8 each group). Three trains of stimuli (20 pulses at 5, 10, 20, and 40 Hz) were delivered, each 6 min apart, and the effect of BRV on the ratio of EPSPn/EPSP1 was measured. To determine if the effect of BRV is mediated by getting into the synaptic vesicle and binding to SV2A, we used a load-unload protocol. After 3hr incubation (loading), hippocampal slices were rinsed for 15 minutes with artificial cerebrospinal fluid (ACSF) to remove extracellular BRV (washout). To unload BRV from presynaptic vesicles, train stimulation (4 Min, 10 Hz) was then delivered to the slices followed by another 15 min ACSF washout (unloading). After each stage (loading, washout, unloading) the ratio of EPSPn/EPSP1 was measured using three trains of stimuli (20 pulses at 40 Hz). Statistical analyses were performed using two way repeated-measures ANOVA. Results: BRV decreases synaptic transmission in a time-, stimulation frequency-, and concentration-dependent manner. At 40 Hz train stimulation, 3 hours incubation with 1-30 µM BRV and 30 minute incubation with ≥10µM BRV significantly decreased the ratio of EPSPn/EPSP1 compared to control beginning at pulse 2 or 3 (P< 0.05). At 20 Hz train stimulation, 3-hour incubation with ≥10µM BRV incubation and 30 minute incubation with 30µM BRV reduced the ratio of EPSPn/EPSP1 compared to control beginning at pulse 3 or 4 (P< 0.05). At 10 Hz train stimulation, only 3-hour 30µM BRV incubation significantly decreased the EPSPn/EPSP1 ratio compared to control starting at pulse 13 (P<0.05). The remaining conditions, we did not see any significant decrease in EPSPn/EPSP1 ratio. Previous studies with LEV showed that the effect of 3-hour 300µM LEV incubation on decreasing synaptic responses could not be altered after a 15-minute washout to remove the drug from the medium, but it could be eliminated by unloading the drug using train stimulation (4 Min, 10 Hz) plus 15-minute washout. But BRV affect cannot be eliminated by either washout or unloading stimulation followed by subsequent washout. Conclusions: Our data suggest that BRV resembles LEV in its effect on synaptic transmission, but is more potent in decreasing synaptic transmission than LEV. The load-unload results are consistent with our hypothesis that these drugs reduce synaptic transmission by getting into synaptic vesicles and binding to SV2A.