Do preclinical seizure models preselect certain adverse effects of antiepileptic drugs

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Abstract

Classical screening tests (maximal electroshock, MES, and threshold pentylenetetrazol, PTZ) employ non-epileptic rodents and identify antiepileptic drugs (AEDs) with mechanisms of action associated with significant CNS side effects. Thus MES identifies drugs acting on Na+ channels that produce cerebellar toxicity. It may be possible to produce novel AEDs more selectively targeted at voltage-sensitive (VS) ion channels. There is little specific evidence for the likely success of this strategy with subunit selective agents targeted at the different VS Na+ channels. Drugs targeted at specific VS Ca++ channels (T, N, P/Q types) may be useful in generalised seizures. There are many as yet unexplored possibilities relating to K+ channels. GABA related drugs acting on PTZ clonic seizures tend to induce sedation and muscle hypotonia. Studies in mice, particularly with knock-in mutations, but also with subunit selective agents acting via the GABAA benzodiazepine site, suggest that it is possible to produce agents which do or do not induce particular side effects (sedative, hypnotic, anxiolytic, muscle relaxant, amnesia, anaesthesia). Whether these findings transfer to man has yet to be established. Acquired epilepsy in rodents (e.g. kindling or spontaneous seizures following chemically- or electrically-induced status epilepticus) or acquired epilepsy in man (following prolonged febrile seizures or traumatic brain injury) is associated with multiple changes in the function and subunit composition of ion channels and receptor molecules. Optimal screening of novel AEDs, both for efficacy and side effects, requires models with receptor and ion channel changes similar to those in the target human syndrome.

Introduction

This review begins by observing that the classical preclinical screening models have consistently selected drugs with significant CNS side effects, apparently as a result of the models identifying compounds with specific molecular targets. The review then discusses immediate and more remote strategies for overcoming this problem. The screening tests that have dominated the search for novel anticonvulsant drugs within pharmaceutical companies, academia and governmental agencies for the last half century have evolved from seizures induced by (a) maximal electroshock (MES) (Merritt and Putnam, 1938) and (b) pentylenetetrazol (PTZ) (Richards and Everett, 1944). These two approaches have formed the basis of screening programmes because individually they identified different constellations of antiepileptic drugs (AEDs) and combined they were thought (until very recently) to identify all potential AEDs. We shall discuss for each test the link between mechanism of AED action and CNS side effect profile. The benefits of using animals with genetic or acquired forms of epilepsy for screening will be discussed.

Section snippets

Maximal electroshock (tonic extension)

Following Merritt and Putnam's original success selecting phenytoin with this test in cats, it evolved into a fast and efficient screening procedure in rats and mice (Krall et al., 1978). Comparison of the efficacy in mice against MES and against threshold PTZ showed that there was a category of drugs with a marked preferential action against MES, most notably phenytoin and carbamazepine which lacked activity against PTZ (Krall et al., 1978). These drugs also share a clinical spectrum of

VS Na+ channel mutations and epilepsy

Mutations in VS Na+ channels are associated with various diseases of muscle (such as periodic paralyses and paramyotonia congenita), the heart (long QT syndrome) and epilepsy (Catterall, 2000). In families with GEFS(+), a syndrome showing generalised seizures and febrile seizures, a variety of mutations have been reported in either the β1 or the α1 subunit. As this is a dominant mutation it is possible that the expressed mutant isoform could be used to screen for drugs that selectively

Is more selective targeting of AEDs for Na+ channels possible?

The AEDs act to prolong inactivation by binding to sites on the 6th transmembrane segments in domains III and IV (Ragsdale and Avoli, 1998, Yarov-Yarovoy et al., 2001, Rogawski, 2002). As first shown by Noda et al. (1986) these regions are highly conserved in the α-subunit, being identical in sequence in rat I and II. This suggests that it may be difficult to design AEDs that differentiate sharply between the Na+ channel isoforms. Nevertheless expression systems for the different isoforms could

Threshold PTZ (clonic seizure) test models

Suppression of clonic seizure responses in mice following the subcutaneous injection of PTZ identifies a group of AEDs that only partially overlaps the group identified by the MES test. In particular benzodiazepines and ethosuximide are much more potent against PTZ than against MES and phenobarbital is moderately more active against PTZ (Krall et al., 1978). Of newer drugs tiagabine and gabapentin are effective and lamotrigine is not (White et al., 2002). Potentiating GABA-mediated inhibition

The GABAA receptor, subunit subtypes and benzodiazepine selectivity

The GABAA receptor is a heteropentameric structure comprising subunits selected from five or more families ( α1–6, β1–4, γ1–4, δ, θ, ρ1–3). Although there are potentially thousands of possible pentameric combinations of 19 or so subunits that could constitute a GABAA receptor, about a dozen combinations make up 90% or more of the receptors expressed in the brain (see Table 2). Studies in cellular expression systems reveal that the subunit combinations determine the sensitivity of the receptor

GABAA receptor subunits: behavioural effects of genetic and pharmacological manipulation

It appears that the anticonvulsant action of diazepam and other major benzodiazepines involves actions at GABAA receptors showing BZ I and BZ II responses, i.e. containing α1, α2, α3 and α5 subunits. Behavioural experiments with genetically-modified mice (both knock-out and knock-in) and with some subtype selective benzodiazepines suggest that the other neurological effects of benzodiazepines are mediated by specific receptor subtypes. Interpretation of the effects in knock-out mice is

Prospects for novel GABA-related AEDs

Recombinant receptors with different subunits can be used to screen novel compounds acting at the BZ site or other sites on the GABAA receptor to identify potential drugs with appropriate selectivity of agonist–antagonist action. These can then be tested for anticonvulsant efficacy and for side effect profile in normal and knock-out and knock-in mice to prove that the effects required have been achieved, and the mechanism is as assumed. The animal data strongly suggest that it should be

Models using epileptic rather than normal animals

The two standard screening procedures (MES and PTZ) employ acutely-induced seizures in normal rodents. This is convenient but does not mimic spontaneous seizures occurring in the epileptic brain. We now know that epilepsy can arise from a single mutation involving an ion channel or receptor molecule or in the case of acquired epilepsy involve a multiplicity of changes in the expression or function of ion channels and receptor subunits. These changes may influence both the antiepileptic efficacy

Therapeutic index and side effect profile of NMDA antagonists

NMDA receptor antagonists, particularly the competitive antagonists acting at the glutamate site (such as AP7 and d-CPPene) and competitive antagonists acting at the glycine site (such as L-687,414) have been shown to be powerful anticonvulsants in reflex epilepsies (e.g. sound-induced seizures in DBA/2 mice and GEPrats, and photically-induced responses in Papio papio (Chapman et al., 1990, Patel et al., 1990, Chapman et al., 1991, Smith and Meldrum, 1992). They are also effective against MES

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