Abstracts

Rationale: Mapping seizure susceptibility loci in mice provides a framework for identifying potentially novel candidate genes for complex human disorders like human epilepsy. Using C57BL/6J (B6, host) X A/J (donor) chromosome substitution strains (CSS), we previously identified loci conferring susceptibility to pilocarpine-induced (limbic) seizures on mouse chromosomes 10 and 18. Here we report further genetic mapping of pilocarpine quantitative trait loci (QTLs) on ch10 and 18. Methods: We bred F2 populations for both CSS10 and CSS18. Male 10-11 week old F2 mice were tested for seizure susceptibility to pilocarpine (250 mg/kg, i.p.) and seizure behaviors were observed for 2.5 hours. Using a seizure staging system adapted from established (Racine) rodent seizure scales, highest stage reached and latency to each stage were recorded for all mice. We used the R/qtl software to map susceptibility loci in the F2 populations and used permutation to calculate the empirical significance threshold for each F2 population. Since relatively few animals in the F2 mapping population reached higher seizure stages (stage 4: rearing/hyperactivity or tonic extension/clonic seizures with loss of posture, and stage 5: continuous tonic/clonic seizures or tonic hindlimb extension), we also used survival methods to map QTLs. Latency to stages and highest stage reached were regressed on the number of B6 alleles carried at each locus, and stages not reached replaced with the time to the end of the observation period. Results: We tested 87 10F2 animals for seizure susceptibility and genotyped them at 6 markers on chromosome 10. QTL analysis identified significant, overlapping QTLs for latency to stage 3 seizures (partial status epilepticus) and duration of stage 3 seizures on distal chromosome 10. The chromosome 10 QTL appears to act in an additive fashion, and the A/J allele confers short latency to stage 3 and long duration of partial status. Survival analysis identified a correlation of time to stage 3 and 4 seizures to marker rs13480719, within the significant linkage region identified by rQTL. In an independent sample of 10F2 animals (N=76), association was found with latency to stages 4 and 5 and with the highest stage reached, with the strongest effect at rs29348118, also within the locus for seizure susceptibility identified by rQTL. We tested 92 18F2 animals for pilocarpine seizure susceptibility and genotyped them at 9 markers on Chromosome 18. QTL analysis revealed a significant linkage peak for duration of partial status near marker rs13483373. The chromosome 18 QTL appears to act in a dominant-recessive fashion, with greater susceptibility conferred by the A/J allele. Conclusions: QTL mapping can identify loci that make a quantitative contribution to a trait, and eventually identify the causative DNA-sequence polymorphisms. We have mapped loci on mouse chromosomes 10 and 18 for pilocarpine-induced seizures, a model for limbic seizure susceptibility. Novel candidate genes identified in mice can be investigated in functional studies, and tested for their role in human epilepsy populations.