Pharmacological validation of a semi-automated in vitro hippocampal brain slice assay for assessment of seizure liability.
BACKGROUND
Drug-induced seizures are a serious, life-threatening adverse drug reaction (ADR) that can result in the failure of drugs to be licensed for clinical use or withdrawn from the market. Seizure liability of potential drugs is traditionally assessed using animal models run during the later phases of the drug discovery process. Given the low throughput, high animal usage and high compound requirement associated with these assays, it would be advantageous to identify higher throughput, in vitro models that could be used to give an earlier assessment of seizure liability. The hippocampal brain slice is one possibility but conventionally allows recording from only one slice at a time. The aim of this study was to validate a semi-automated system (Slicemaster, Scientifica UK Ltd) which allows concurrent electrophysiological recording from multiple brain slices.


METHODS
Conventional electrophysiological recording techniques were used to record electrically evoked synaptic activity from rat hippocampal brain slices. Population spikes (PS) were evoked at 30 s intervals by electrical stimulation of the Schaffer collateral pathway and were recorded using extracellular electrodes positioned in the CA1 cell body layer. Responses were quantified as PS areas (the area above and below the 0 mV line). The effects of eight validation compounds known to cause seizures in vivo and/or in the clinic were assessed.


RESULTS
Seven out of eight compounds evoked a concentration-dependent increase in population spike (PS) area that was statistically significant at higher concentrations (P<0.05; ANOVA). At the highest test concentration the percentage effects (mean+/-s.e.m.), relative to vehicle, were: picrotoxin 212.9+/-28.8, pentylenetrazole (PTZ) 181.4+/-24.7, 4-AP 328.9+/-48.6, aminophylline 124.5+/-5.9, chlorpromazine 122.1+/-9.8, SNC-80 132.1+/-12.6 and penicillin 174.7+/-14.1. Physostigmine had no significant effect on PS area although a concentration-dependent change in the morphology of the response was evident.


CONCLUSIONS
All validation compounds evoked a statistically significant effect on synaptic activity in the rat hippocampal slice. Although similar effects have been described previously, this is the first time that the effects of a pharmacologically diverse set of compounds have been assessed using a standardised brain slice assay. Given the low compound usage and relatively high throughput associated with this assay, the hippocampal brain slice assay may facilitate earlier testing of convulsant liability than is currently possible using in vivo models.
Easter
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Safety Assessment UK, AstraZeneca R&D Alderley Park, Macclesfield, SK10 4TG, UK. alison.easter@astrazeneca.com
Sharp
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Valentin
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Pollard
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D015761:4-Aminopyridine / Q000633:toxicity; D000200:Action Potentials / Q000187:drug effects / Q000502:physiology*; D000818:Animals; D001549:Benzamides / Q000633:toxicity; D002746:Chlorpromazine / Q000633:toxicity; D003292:Convulsants / Q000145:classification / Q000633:toxicity; D004121:Dimethyl Sulfoxide / Q000633:toxicity; D004305:Dose-Response Relationship, Drug; D004353:Drug Evaluation, Preclinical / Q000295:instrumentation / Q000379:methods; D004594:Electrophysiology / Q000295:instrumentation / Q000379:methods*; D005260:Female; D006624:Hippocampus / Q000187:drug effects / Q000502:physiology*; D066298:In Vitro Techniques; D008297:Male; D010400:Penicillin G / Q000633:toxicity; D010433:Pentylenetetrazole / Q000633:toxicity; D010830:Physostigmine / Q000633:toxicity; D010852:Picrotoxin / Q000633:toxicity; D010879:Piperazines / Q000633:toxicity; D051381:Rats; D017208:Rats, Wistar; D015203:Reproducibility of Results; D012640:Seizures / Q000175:diagnosis / Q000503:physiopathology*
D015761:4-Aminopyridine; D001549:Benzamides; D002746:Chlorpromazine; D003292:Convulsants; D004121:Dimethyl Sulfoxide; D010400:Penicillin G; D010433:Pentylenetetrazole; D010830:Physostigmine; D010852:Picrotoxin; D010879:Piperazines; C088464:4-(alpha-(4-allyl-2,5-dimethyl-1-piperazinyl)-3-methoxybenzyl)-N,N-diethylbenzamide