Abstracts

Inheritance patterns between mouse strains suggest that a robust genetic influence determines individual susceptibility to kainic acid-induced hippocampal cell death. Previous quantitative trait loci (QTL) mapping studies had located three significant loci on chromosomes 18, 15, and 4 in the mouse genome, responsible for seizure-induced cell death susceptibility. The QTL with the largest effect on susceptibility to seizure-induced cell death maps to the distal region of chromosome 18 (Schauwecker et al. 2004). In order to localize the loci more precisely, we generated a series of congenic and subcongenic strains by introgressing segments of seizure-induced cell death loci from the C57BL/6 resistant mouse into the background of the FVB/N susceptible mouse. Interval-specific congenic lines (ISCLs) were derived by identifying mice that carried recombination events in the C57BL/6 interval, during the backcrossing for congenics. Recombinant mice were backcrossed to FVB/N, and progeny that carried the reduced Chr 18, Chr 15, or Chr 4 region were tested for the effect on susceptibility to seizure-induced cell death. Mice were injected with kainate and observed for 3-4 hours for the onset and extent of seizures. Seven days following kainate, animals were perfused and hippocampi were assessed for susceptibility to kainate-induced cell death. A candidate gene approach was used to identify strong candidate genes within our introgressed regions by comparing the expression of candidate genes between C57BL/6 and FVB/N mice strains using quantitative RT-PCR. Genomic DNA from C57BL/6 and FVB/N mice was amplified and sequenced to identify potential single nucleotide polymorphisms (SNPs) so as to detect genetic variants in both susceptible (FVB/N) and resistant (C57BL/6) murine populations. Three congenic strains have been produced through introgression of various segments of chromosome 18, 15, or 4 from C57BL/6 mice into the recipient FVB/N mouse, and vice versa. In all cases, the extent of cell death was dramatically affected in that damage was significantly reduced in those congenic strains that contained a C57BL/6 resistant region in a FVB/N strain. We have developed multiple ISCLs which span our 3 QTL intervals, which have allowed us to begin to test each chromosomal segment statistically for linkage to kainate-induced cell death. Two of these share the C57BL/6 phenotype of reduced cell death. Differential expression of hippocampal mRNA for several candidate genes using quantitative RT-PCR and nucleotide variants have been identified between FVB/N and C57BL/6 strains as well. Overall, these data confirm the existence of the genetic loci that confer susceptibility to seizure-induced cell death. The resolution of individual recombinants obtained in this study should allow us to narrow our originally defined critical fragments down to a size amenable to facilitate the positional cloning of the loci. (Supported by NS038696 (PES).)