Levetiracetam has an activity-dependent effect on inhibitory transmission.

PURPOSE: Previous work has shown that levetiracetam (LEV) binds the vesicular protein SV2A and reduces excitatory neurotransmitter release during trains of high-frequency activity, most likely by accessing its binding site through vesicular endocytosis into excitatory synaptic terminals. Because there are differences in excitatory and inhibitory transmitter release mechanisms, and there are suggestions that neurons differ in their SV2A expression, we were curious whether LEV also reduces inhibitory transmission. METHODS: We used patch-clamp recording from CA1 neurons in rat brain slices to quantify the effects of LEV on inhibitory postsynaptic currents (IPSCs). We were able to elicit pure IPSCs by stimulating inhibitory terminals close to neuronal soma and blocking excitatory postsynaptic currents with specific antagonists. KEY FINDINGS: We found that LEV reduces inhibitory currents in a frequency-dependent manner, with the largest relative effect on the later IPSCs in the highest frequency trains. However, in contrast to excitatory postsynaptic currents (EPSCs), LEV reduced IPSC trains after a briefer, 30 min incubation. When spontaneous activity during incubation was blocked with antagonists of excitatory transmission, LEV no longer reduced IPSCs. If slices were returned to LEV-free artificial cerebrospinal fluid (ACSF) after LEV incubation, but prior to recording, the IPSC reduction failed to appear. However, if synaptic activity was limited by treating with excitatory transmitter antagonists, after the initial LEV exposure, LEV still diminished trains of IPSC. The concentration required to diminish IPSC trains was lower than for EPSCs. SIGNIFICANCE: LEV exerts a qualitatively similar, frequency-dependent effect on both IPSCs and EPSCs. The much shorter latency for IPSC reduction is consistent with the greater levels of spontaneous inhibition in brain slices, supporting the hypothesis that vesicular uptake is necessary for the entry of LEVs into terminals. The vesicular entry of LEV resembles the cell entry pathways for tetanus and botulinum neurotoxins, but is unique for small, neuroactive drugs. Although the reduction of IPSC trains by LEV initially seems counterintuitive for an antiepileptic drug, there are multiple reasons that disruption of γ-aminobutyric acid (GABA) release could ultimately attenuate pathologic discharges.

A new mechanism for antiepileptic drug action: vesicular entry may mediate the effects of levetiracetam.

Levetiracetam (LEV) is one of the most commonly prescribed antiepileptic drugs, but its mechanism of action is uncertain. Based on prior information that LEV binds to the vesicular protein synaptic vesicle protein 2A and reduces presynaptic neurotransmitter release, we wanted to more rigorously characterize its effect on transmitter release and explain the requirement for a prolonged incubation period for its full effect to manifest. During whole cell patch recordings from rat hippocampal pyramidal neurons in vitro, we found that LEV decreased synaptic currents in a frequency-dependent manner and reduced the readily releasable pool of vesicles. When we manipulated spontaneous activity and stimulation paradigms, we found that synaptic activity during LEV incubation alters the time at which LEV's effect appears, as well as its magnitude. We believe that synaptic activity and concomitant vesicular release allow LEV to enter recycling vesicles to reach its binding site, synaptic vesicle protein 2A. In support of this hypothesis, a vesicular "load-unload" protocol using hypertonic sucrose in the presence of LEV quickly induced LEV's effect. The effect rapidly disappeared after unloading in the absence of LEV. These findings are compatible with LEV acting at an intravesicular binding site to modulate the release of transmitter and with its most marked effect on rapidly discharging neurons. Our results identify a unique neurobiological explanation for LEV's highly selective antiepileptic effect and suggest that synaptic vesicle proteins might be appropriate targets for the development of other neuroactive drugs.

Seletracetam enhances short term depression in vitro.

Seletracetam (SEL), an analog of the antiepileptic drug levetiracetam (LEV), decreases seizure activity in a number of epilepsy models and binds to the synaptic vesicle protein SV2A with a higher affinity than LEV. Experiments were performed to determine if SEL, like LEV, reduces the later EPSPs in long trains of stimuli in a manner dependent upon access to the interior of synaptic vesicles and SV2A binding. When hippocampal slices were incubated in 3-30µM SEL for 3h, but not 30 min, the relative amplitude of the CA1 field excitatory synaptic potentials decreased over the course of a train of high frequency stimuli more than for control slices. This short term depression was frequency and dose dependent and largely disappeared when the spontaneous activity during the loading period was removed by cutting the Schaffer collaterals. The SEL effect was also observed in slices loaded during prolonged stimulation at 1Hz, but not 10Hz. Hippocampal slices loaded with both SEL and FM1-43 to visualize synaptic boutons released the FM1-43 in response to prolonged stimulation much more slowly than control slices during prolonged stimulation. Like LEV, SEL produced a frequency-dependent decrement of synaptic transmission that was dependent upon the drug entering recycling synaptic vesicles and compatible with SV2A binding. Previous observations of SV2A binding affinity correlated with the current effect of SEL and the previously reported effect of LEV on synaptic transmission validate SV2A as an extremely attractive target for future antiepileptic drug development.