Cellular electrophysiological recording techniques are used to study drug modulation of neurotransmitter-gated and voltage-activated ion channels in brain slices, cultured neurons and heterologous cells transfected with cloned ion channel subunit genes. Correlative studies are carried out in animal models. ? ? In a series of experiments we revealed the roles of kainate receptors (KAR) that contain the GluR5 subunit (KAR-G5) in seizure induction and epileptogenesis. These receptors are a subgroup of ionotropic glutamate receptors which are distinguishable from NMDA and AMPA receptors. We used KAR-G5 receptor knockout mice and the KAR-G5 agonist ATPA to determine the behavioral, clinical seizure and electrophysiological effects of KAR-G5 receptor activation. By performing slow tail vain infusion of the selective GluR 5 agonist ATPA during Video-EEG recordings we demonstrated that the selective activation of KAR-G5 mediates a behavioral arrest followed by myoclonic jerks which show a sharp wave as EEG correlate. Clonic and tonic seizures occurring at higher doses of ATPA are not mediated by KAR-G5 and are most likely due to AMPA receptor activation as ATPA loses its selectivity at higher concentrations. These data were confirmed in in vitro slice recordings and behavioral testing for locomotor activity. ? ? In a different approach we triggered epilepsy in rats by induction of SE with intraperitoneal injections of kainic acid. Seven to thirteen days after SE we performed 24h Video-EEG monitoring, which revealed signs of epileptogenesis in all animals (EEG seizures, clinical seizures, sharp waves). The following day the rats were sacrificed for in vitro slice recordings (whole cell patch clamp, extracellular recordings) to characterize potential changes in the functional effects of KAR-G5 activation on GABA transmission. GABA is the main inhibitory neurotransmitter in the adult brain of humans as well as rodents.? ? Experiments published by our collaborator Maria Braga showed that activation of KAR-G5 in the amygdala slice preparation of healthy rats has a bidirectional effect. Low concentrations of ATPA (1uM) increase GABAergic transmission but higher doses (10uM) rather decrease GABAergic transmission. We found that in the epileptic amygdala the response to low concentrations of ATPA is reduced, reflecting less enhancement of GABAergic transmission ( disinhibition). However, 10 uM ATPA did still induce bursting activity in the slices of epileptic animals. This suggests that the function of KARG5 receptors may be changed in a way that excitation outweighs inhibition. Further studies investigating the cellular location and detailed subunit composition that may have changed during epileptogenesis are underway. ? ? Furthermore we performed a therapeutic study using a competitive antagonist at the AMPA receptor, also an ionotropic glutamate receptor. AMPA receptors carry the vast majority of the fast excitatory currents induced by glutamate (main excitatory neurotransmitter). This makes them a promising target as treatment for status epilepticus. The study was performed in mice under Video-EEG recordings. We chose two arbitrary time points (early and late) for treatment with the AMPA antagonists and compared it with vehicle and diazepam.? ? At both time points early (after the first 5 min of continuous EEG seizure activity) and late (25 min later) the AMPA antagonist was superior to both vehicle and diazepam in successfully interrupting the ongoing seizure activity without later reoccurrence of seizures. We did not see any mortality after the AMPA antagonist treatment. The mortality rate in the diazepam group (57%) exceeded even the vehicle group what is most likely due to the depressant effect on the respiratory and cardiovascular system.? ? The potential antiepileptogenic (protection against epilepsy after SE) effect of the AMPA antagonist treatment is currently being evaluated by screening for the occurrence of spontaneous seizures with 24h Video-EEG- monitoring sessions up to 20 weeks after SE.
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