Genetic control of sensitivity to hippocampal cell death induced by kainic acid: a quantitative trait loci analysis

Host genetic factors are likely to contribute to differences in individual susceptibility to seizure-induced excitotoxic neuronal damage. Similarly, inbred strains of mice differ in their susceptibility to the kainic acid (KA) model of seizure-induced cell death, but the genes responsible for the differences are not known. Here, we define the inheritance patterns of susceptibility to KA-induced neurodegeneration in the hippocampus by assessing 331 back-cross (N2) progeny of two inbred mouse strains, C57BL/6 and FVB/N, previously shown to display resistance and sensitivity to KA-induced cell death, respectively. Results of phenotypic analysis suggest that the difference in susceptibility between these two strains is conferred by a single dominant gene. Therefore, we used an N2 back-cross between the inbred C57BL/6 and FVB/N strains for a genome-wide search for quantitative trait loci (QTLs), which are chromosomal sites containing genes influencing the magnitude of susceptibility. Genome-wide interval mapping in N2 progeny identified a locus on distal chromosome (Chr) 18 with a peak LOD score of 4.9 localized between D18Mit186 and D18Mit4 as having the strongest and most significant effect in this model. QTLs of minor effect were detected on Chr 15 (D15Mit174-D15Mit156) and Chr 4 (D4Mit264-D4Mit91), with peak LOD scores of 3.02 and 2.46, respectively. The three significant QTLs (Chrs 4, 15, 18) together account for nearly 25% of the trait variance for both genders combined. Reduced KA-induced cell death susceptibility was observed in a congenic strain in which the highly susceptible FVB/N strain carried putative resistance alleles from the C57BL/6 strain on Chr 18.     

Differences in ionotropic glutamate receptor subunit expression are not responsible for strain-dependent susceptibility to excitotoxin-induced injury

Systemic administration of kainic acid in C57BL/6 and FVB/N mice induces a comparable level of seizure induction yet results in differential susceptibility to seizure-induced cell death. While kainate administration causes severe hippocampal damage in mice of the FVB/N strain, C57BL/6 mice display no demonstrable cell loss or damage. At present, while the cellular mechanisms underlying strain-dependent differences in susceptibility remain unclear, some of this variation is assumed to have a genetic basis. As glutamate receptors are thought to participate in seizure induction and the subsequent neuronal degeneration that ensues, previous studies have proposed that variation in the precise subunit composition of glutamate receptors may result in differential susceptibility to excitotoxic cell death. Thus, we chose to examine the relationship between the cellular distribution and expression of glutamate receptor subunit proteins and cell loss within the hippocampus in mouse strains resistant and susceptible to kainate-induced excitotoxicity. Using semi-quantitative Western blot techniques and immunohistochemistry with the use of antibodies that recognize subunits of the KA (GluR5,6,7), AMPA (GluR1, GluR2, and GluR4), and NMDA (NMDAR1 and NMDAR2A/2B) receptors, we found no significant strain-dependent differences in the expression or distribution of these glutamate receptor subunits in the intact hippocampus. Following kainate administration, expression changes in ionotropic glutamate receptor subunits paralleled the development of susceptibility to cell death in the FVB/N strain only. Strain differences in hippocampal vulnerability to kainate-induced status epilepticus are not due to glutamate receptor protein expression.     

Comparison of seizure phenotype and neurodegeneration induced by systemic kainic acid in inbred, outbred, and hybrid mouse strains

We assessed inbred, outbred and hybrid mouse strains for susceptibility to seizures and neurodegeneration induced by systemic administration of kainic acid (KA). Each strain showed a unique pattern of susceptibility to seizures as assessed by the dose necessary to induce continuous tonic clonic seizures, progression through six seizure levels, the number of mice that failed to satisfy seizure criteria, and seizure-induced mortality. In general, the C57BL/6, ICR, FVB/N, and BALB/c strains were resistant to seizures while the C57BL/10, DBA/2 J, and F1 C57BL/6*CBA/J strains were vulnerable. Neuronal cell death was quantified in four subfields of the hippocampus: CA3, the hilus of the dentate gyrus, CA1, and the dentate granule cell layer. Neurodegeneration was also semiquantitatively assessed in other brain regions including the neocortex, striatum, thalamus, hypothalamus and amygdala. Although there was variability in the extent of cell death within strains, there were significant differences in the amount of hippocampal cell death between strains and also different patterns of neurodegeneration in affected brain areas. In general, the C57BL/6, C57BL/10, and F1 C57BL/6*CBA/J strains were resistant to neurodegeneration while the FVB/N, ICR and DBA/2 J strains were vulnerable. The BALB/c strain was unique in that neurodegeneration was confined to the hippocampus. Consistent with previous findings, the resistant neurodegeneration phenotype was dominant in an F1 cross of resistant and vulnerable inbred strains. Our results, using a large number of mouse strains, definitively demonstrate that a mouse strain's seizure phenotype is not related to its neurodegeneration phenotype.     

Mouse chromosome 11 harbors genetic determinants of hippocampal strain-specific nicotinic receptor expression

Differences between isogenic mouse strains in cellular expression of the neuronal nicotinic acetylcholine (ACh) receptor subunit alpha 4 (nAChR alpha 4) by the dorsal hippocampus are well known. To investigate further the genetic basis of these variations, expression of the nAChR alpha 4 subunit was measured in congenic mouse lines derived from two strains exhibiting notable divergence in the expression of this subunit: C3H and C57BL/6. Congenic lines carrying reciprocally introgressed regions (quantitative trait loci; QTL) from chromosomes 4, 5, and 12 each retained the phenotype most closely associated with the parental strain. However, in congenic lines harboring the reciprocal transfer of a chromosome 11 QTL, a characteristic difference in the ratio of interneurons versus astrocytes expressing nAChR alpha 4 in the CA1 region is reversed relative to the parental strain. These finding suggest that this chromosomal segment harbors genes that regulate strain distinct hippocampal morphology that is revealed by nAChR alpha 4 expression.     

Kainic Acid-Induced Neuronal Degeneration in Hippocampal Pyramidal Neurons Is Driven by Both Intrinsic and Extrinsic Factors: Analysis of FVB/N{leftrightarrow}C57BL/6 Chimeras

The excitotoxic effects of kainic acid (KA) in the mouse hippocampus is strain dependent. Following KA administration, the large majority of hippocampal pyramidal cells die in the FVB/N (FVB) mouse, while the pyramidal cells of the C57BL/6 (B6) strain are largely spared. We generated aggregation chimeras between the sensitive FVB and the resistant B6 strains to investigate whether intrinsic or extrinsic features of a neuron confer cell vulnerability or resistance to KA. The constitutive expression of transgenic green fluorescence protein (GFP) or β-galactosidase expressed from the ROSA26 locus was used to mark cells in FVB or B6 mice, respectively. These makers enable the identification of cells from each parental genotype while TUNEL (terminal deoxynucleotidyl transferase-mediated biotinylated dUTP nick end labeling)-staining labeled dying cells. The analysis of the percentage of dying cells in FVB-GFP ? B6-ROSA chimeras yielded an intriguing mix of both intrinsic and extrinsic factors in the readout of cell phenotype. Thus, normally resistant B6-ROSA pyramidal neurons demonstrated an increasing sensitivity to KA, in a linear fashion, when the percentage of FVB-GFP cells was increased, either across chimeras or in different regions of the same chimera. However, the death of B6-ROSA pyramidal cells never exceeded ～70% of the total amount of B6 neurons regardless of the amount of FVB cells in the chimeric hippocampus. In a similar manner, FVB-GFP cells show lower amounts of cell death in chimeras that are colonized by B6-ROSA cells, but again, are never fully rescued. These data indicate that both intrinsic and extrinsic factors modulate the sensitivity of hippocampal pyramidal cells to kainic acid.     

Genetic basis of kainate-induced excitotoxicity in mice: phenotypic modulation of seizure-induced cell death

Excitotoxicity, a process in which excessive excitation of glutamate receptors results in cell death, has been implicated in a number of neurological disorders. However, the genetic characteristics and molecular mechanisms that can modulate the extent of cell death are unclear. Previously, we had reported that the extent of excitotoxic cell death is conferred by differences in the genetic background of several mouse strains. As a first step in the identification of loci that can modulate the extent of excitotoxin-induced cell death, we tested C57BL/6 and FVB/N mice, their F1 hybrids and backcross progeny for differences in apparent excitotoxic cell death induced by kainic acid (KA). While no strain dependent differences in seizure duration were observed, phenotypic analysis of cell death indicated that C57BL/6 mice showed no seizure-induced cell death, while FVB/N mice exhibited extensive cell death. Studies of seizure-induced cell death in hybrid and backcross progeny revealed an association between seizure-induced cell death and genotype. Mice from the F1 cross exhibited little to no seizure-induced cell death, indicative that the extent of cell death is conferred as a dominant genetic trait. Phenotypic assessment of cell death in backcross progeny suggests that differences in apparent cell death are conferred by a single gene locus. These findings implicate genetic factors in individual differences in excitotoxin-induced cell death.     

Quantitative trait locus on distal chromosome 1 regulates the occurrence of spontaneous spike-wave discharges in DBA/2 mice

Purpose: Most common forms of human epilepsy result from a complex combination of polygenetic and environmental factors. Quantitative trait locus (QTL) mapping is a first step toward the nonbiased discovery of epilepsy-related candidate genes. QTL studies of susceptibility to induced seizures in mouse strains have consistently converged on a distal region of chromosome 1 as a major phenotypic determinant; however, its influence on spontaneous epilepsy remains unclear. In the present study we characterized the influence of allelic variations within this QTL, termed Szs1, on the occurrence of spontaneous spike-wave discharges (SWDs) characteristic of absence seizures in DBA/2 (D2) mice. Methods: We analyzed SWD occurrence and patterns in freely behaving D2, C57BL/6 (B6) and the congenic strains D2.B6-Szs1 and B6.D2-Szs1. Key Findings: We showed that congenic manipulation of the Szs1 locus drastically reduced the number and the duration of SWDs in D2.B6-Szs1 mice, which are homozygous for Szs1 from B6 strain on a D2 strain background. However, it failed to induce the full expression of SWDs in the reverse congenic animals B6.D2-Szs1. Significance: Our results demonstrate that the occurrence of SWDs in D2 animals is under polygenic control and, therefore, the D2 and B6 strains might be a useful model to dissect the genetic determinants of polygenic SWDs characteristic of typical absence seizures. Furthermore, we point to the existence of epistatic interactions between at least one modifier gene within Szs1 and genes within unlinked QTLs in regulating the occurrence of spontaneous nonconvulsive forms of epilepsies.     

Neuroprotection by glutamate receptor antagonists against seizure-induced excitotoxic cell death in the aging brain

We previously have identified phenotypic differences in susceptibility to hippocampal seizure-induced cell death among two inbred strains of mice. We have also reported that the age-related increased susceptibility to the neurotoxic effects of seizure-induced injury is regulated in a strain-dependent manner. In the present study, we wanted to begin to determine the pharmacological mechanism that contributes to variability in the response to the neurotoxic effects of kainate. Thus, we compared the effects of the NMDA receptor antagonist, MK-801 and of the AMPA receptor antagonist NBQX on hippocampal damage in the kainate model of seizure-induced excitotoxic cell death in young, middle-aged, and aged C57BL/6 and FVB/N mice, when given 90 min following kainate-induced status epilepticus. Following kainate injections, mice were scored for seizure activity and brains from mice in each age and antagonist group were processed for light microscopic histopathologic evaluation 7 days following kainate administration to evaluate the severity of seizure-induced injury. Administration of MK-801 significantly reduced the extent of hippocampal damage in young, mature and aged FVB/N mice, while application of NBQX was only effective at attenuating cell death in young and aged mice throughout all hippocampal subfields. Our results suggest that both NMDA and non-NMDA receptors are involved in kainate-induced cell death in the mouse and suggest that aging may differentially affect the ability of neuroprotectants to protect against hippocampal damage. Differences in the effectiveness of these two antagonists could result from differential regulation of glutamatergic neurotransmitter systems or ion channel specificity.     

Genetic dissection of the signals that induce synaptic reorganization

Synaptic reorganization of mossy fibers following kainic acid (KA) administration has been reported to contribute to the formation of recurrent excitatory circuits, resulting in an epileptogenic state. It is unclear, however, whether KA-induced mossy fiber sprouting results from neuronal cell loss or the seizure activity that KA induces. We have recently demonstrated that certain strains of mice are resistant to excitotoxic cell death, yet exhibit seizure activity similar to what has been observed in rodents susceptible to KA. The present study takes advantage of these strain differences to explore the roles of seizure activity vs cell loss in triggering mossy fiber sprouting. In order to understand the relationships between gene induction, cell death, and the sprouting response, we assessed the regulation of two molecules associated with the sprouting response, c-fos and GAP-43, in mice resistant (C57BL/6) and susceptible (FVB/N) to KA-induced cell death. Following administration of KA, increases in c-fos immunoreactivity were observed in both strains, although prolonged induction of c-fos was present only in the hippocampal neurons of FVB/N mice. Mossy fiber sprouting following KA administration was also only observed in FVB/N mice, while induction of GAP-43, a marker associated with mossy fiber sprouting, was not observed in either strain. These results indicate that: (i) KA-induced seizure activity alone is insufficient to induce mossy fiber sprouting; (ii) mossy fiber sprouting may be due to the loss of hilar neurons following kainate administration; and (iii) induction of GAP-43 is not a necessary component of the sprouting response that occurs following KA in mice.     

Genes on distal chromosome 18 determine vulnerability to excitotoxic neurodegeneration following status epilepticus, but not striatal neurodegeneration induced by quinolinic acid

Previous studies have provided evidence that a quantitative trait locus (QTL) on the distal part of chromosome 18 (chr18) is a major determinant of vulnerability to hippocampal neurodegeneration following kainic acid (KA)-induced seizures in inbred mouse strains. We assessed excitotoxic vulnerability in two congenic, “genome tagged” mouse strains carrying segments of either distal or proximal/medial chr18 from vulnerable DBA/2J mice on a resistant C57BL/6 background. Systemic KA injections triggered brain-wide neurodegeneration in the distal chr18 congenic strain, and specifically in the hilus of the dentate gyrus, but not in CA3. In contrast, the proximal/medial chr18 congenic strain exhibited enhanced degeneration in CA1 and CA3, but little neurodegeneration elsewhere. Both strains exhibited low levels of QUIN-induced striatal neurodegeneration comparable to what is seen in C57BL/6 mice. These results suggest that gene(s) on distal chr18 are important determinants of vulnerability to KA-induced hippocampal neurodegeneration, but not QUIN-induced striatal neurodegeneration.     

Susceptibility to excitotoxic and metabolic striatal neurodegeneration in the mouse is genotype dependent

Previously, we had reported that hippocampal susceptibility to the neurotoxic effects of excitotoxin administration is strain dependent [Schauwecker and Steward, Proc. Natl. Acad. Sci. U.S.A. 94 (1997) 4103]. However, it has been unclear whether strain-related gene products may play a similar role in providing protection against drugs that produce striatal lesions. The present series of experiments sought to elucidate whether genetic background alters neuronal viability within the striatum following metabolic or excitotoxic injury. Thus, we have examined the effect of mouse strain on susceptibility to striatal injury using well-characterized animal models of Huntington's disease by examining whether C57BL/6 mice, previously identified as resistant to excitotoxin-induced hippocampal cell death, are resistant to quinolinate, malonate, and 3-nitropropionic acid (3-NP). Intrastriatal injection of either malonate or quinolinate and systemic administration of 3-NP resulted in significantly smaller striatal lesions in C57BL/6 mice as compared to FVB/N mice, previously identified as susceptible to hippocampal excitotoxic injury. The existence of an animal strain with decreased resistance to striatal lesions suggests that there are mediating factors involved in the preferential vulnerability of the striatum to neurotoxic lesioning. The identification of these factors could provide strategies for therapeutic intervention in Huntington's disease.     

Identification of a single QTL,Mptp1, for susceptibility to MPTP-induced substantia nigra pars compacta neuron loss in mice

The loss of substantia nigra pars compacta (SNpc) neurons seen in idiopathic Parkinson's disease is hypothesized to result from a genetic susceptibility to an unknown environmental toxin. MPTP has been used as a prototypical toxin, since exposure to this drug results in variable SNpc cell death in several vertebrate species, including man and mouse. Previously, we have shown that C57BL/6J mice are sensitive to this compound, while Swiss-Webster mice are resistant. In this study, we intercrossed these mouse strains to map quantitative trait loci (QTL) for MPTP sensitivity. Using genome wide PCR analysis, we found that a single major QTLs, Mptp1, located near the distal end of chromosome 1 between D1Mit113 and D1Mit293, accounts for the majority of the strain sensitivity to MPTP.     

Mapping an X-linked locus that influences heat-induced febrile seizures in mice

Purpose: Febrile seizures (FS) are the most common seizure type in children between the age of 6 months and 5 years. Although FS are largely benign, recurrent FS are a major risk factor for developing temporal lobe epilepsy (TLE) later in life. The mechanisms underlying FS are largely unknown; however, family and twin studies indicate that FS susceptibility is under complex genetic control. We have recently developed a phenotypic screen to study the genetics of FS susceptibility in mice. Using this screen in a phenotype‐driven genetic strategy we analyzed the C57BL/6J‐Chr #^A/NaJ chromosome substitution strain (CSS) panel. In each CSS line one chromosome of the A/J strain is substituted in a genetically homogeneous C57BL/6J background. The analysis of the CSS panel revealed that A/J chromosomes 1, 2, 6, 10, 13, and X carry at least one quantitative trait locus (QTL) for heat‐induced FS susceptibility. The fact that many X‐linked genes are highly expressed in the brain and have been implicated in human developmental disorders often presenting with seizures (like fragile X mental retardation) prompted us to map the chromosome X QTL. Methods: C57BL/6J mice were mated with C57BL/6J‐Chr X^A/NaJ (CSSX) to generate F₂‐generations—CXBL6 and BL6CX—originating from CSSX or C57BL/6J mothers, respectively. Heat‐induced FS were elicited on postnatal day 14 by exposure to a controlled warm airstream of 50°C. The latency to heat‐induced FS is our phenotype. This phenotype has previously been validated by video–electroencephalography (EEG) monitoring. After phenotyping and genotyping the F₂‐population, QTL analysis was performed using R/QTL software. Key Findings: QTL analysis revealed a significant peak with an LOD‐score of 3.25. The 1‐LOD confidence interval (149,886,866–158,836,462 bp) comprises 52 protein coding genes, of which 34 are known to be brain expressed. Two of these brain‐expressed genes have previously been linked to X‐linked epilepsies, namely Cdkl5 and Pdha1. Significance: Our results show that the mouse genetics of X‐linked FS susceptibility is complex, and that our heat‐induced FS‐driven genetic approach is a powerful tool for use in unraveling the complexities of this trait in mice. Fine‐mapping and functional studies will be required to further identify the X‐linked FS susceptibility genes.     

Modulation of cell death by mouse genotype: differential vulnerability to excitatory amino acid-induced lesions

Previously we have demonstrated that mature inbred strains of mice differ significantly in their response to kainate-induced cell death. While both C57BL/6 and FVB/N mice exhibit similar seizure activity in response to kainate, only C57BL/6 mice can be characterized as resistant to kainate-induced cell death. To examine further the molecular pharmacological basis for this strain difference in hippocampal sensitivity, we assessed the ability of the ionotropic glutamate receptor agonists, kainic acid (KA), N-methyl-D-aspartate (NMDA), ibotenic acid (IBO), and quinolinic acid (QUIN), to promote excitotoxic damage. We examined seizure-related behavior and subsequent neurotoxicity in C57BL/6 and FVB/N mice following intrahippocampal administration of the kainate receptor agonist, KA, the NMDA receptor agonists NMDA or QUIN, or the NMDA and metabotropic glutamate receptor agonist, IBO. The time course and extent of cell death in mice were evaluated using Nissl and selective silver stains, and Fluoro-Jade, a fluorescent marker for dying neurons. In the present study, FVB/N mice were exquisitely sensitive to injection of KA at all doses, while susceptibility in C57BL/6 mice was dose dependent. In contrast, while hippocampal damage was present in both strains at all doses of QUIN, the extent of cell damage was significantly less in C57BL/6 mice at low doses (30 and 60 mM). Similarly, IBO administration resulted in differences in the extent of cell death when administered at the highest dose (126 mM). No strain-dependent differences in cell loss were observed following NMDA lesions. These results provide further evidence that susceptibility to excitotoxin-induced cell death is highly strain dependent and is kainate and NMDA receptor dependent.     

Mapping of a FEB3 homologous febrile seizure locus on mouse chromosome 2 containing candidate genes Scn1a and Scn3a

Febrile seizures (FS) are the most common seizure type in children. Recurrent FS are a risk factor for developing temporal lobe epilepsy later in life and are known to have a strong genetic component. Experimental FS (eFS) can be elicited in mice by warm‐air induced hyperthermia. We used this model to screen the chromosome substitution strain (CSS) panel derived from C57BL/6J and A/J for FS susceptibility and identified C57BL/6J‐Chr2^A/NaJ (CSS2), as the strain with the strongest FS susceptibility phenotype. The aim of this study was to map FS susceptibility loci and select candidate genes on mouse chromosome 2. We generated an F₂ population by intercrossing the hybrids (F₁) that were derived from CSS2 and C57BL/6J mice. All CSS2‐F₂ individuals were genotyped and phenotyped for eFS susceptibility, and QTL analysis was performed. Candidate gene selection was based on bioinformatics analyses and differential brain expression between CSS2 and C57BL/6J strains determined by microarray analysis. Genetic mapping of the eFS susceptibility trait identified two significant loci: FS‐QTL2a (LOD‐score 3.6) and FS‐QTL2b (LOD‐score 6.2). FS‐QTL2a contained 44 genes expressed in the brain at post natal day 14. Four of these (Arl6ip6, Cytip, Fmnl2 Ifih1) contained a non‐synonymous SNP comparing CSS2 and C57BL/6J, six genes (March7, Nr4a2, Gpd2, Grb14, Scn1a, Scn3a) were differentially expressed between these strains. A region within FS‐QTL2a is homologous to the human FEB3 locus. The fact that we identify mouse FS‐QTL2a with high FEB3 homology is strong support for the validity of the eFS mouse model to study genetics of human FS.     

Protection provided by cyclosporin A against excitotoxic neuronal death is genotype dependent

PURPOSE: Previous studies have shown that the immunosuppressant cyclosporin A (CsA), a specific blocker of the mitochondrial permeability transition (MPT) pore, can dramatically ameliorate the selective neuronal necrosis resulting from ischemia-reperfusion, traumatic brain injury, and N-methyl-d-aspartate (NMDA)-evoked neurotoxicity. The purpose of this study was to determine whether two different immunosuppressants, CsA and FK-506, could ameliorate the neuronal damage observed after kainate-induced seizures in strains that are differentially susceptible to excitotoxin-induced cell death. METHODS: Excitotoxin-resistant (C57BL/6) or -susceptible (FVB/N) mice were administered kainate alone (30 mg/kg), CsA alone (5, 10, or 20 mg/kg), or one of the immunosuppressants (CsA, 5 mg/kg or 10 mg/kg; FK-506, 0.5 mg/kg) followed by kainate. After drug administration, mice were monitored continuously for the onset and extent of seizure activity. After a survival of 7 days, animals were assessed for hippocampal damage. RESULTS: Whereas CsA alone induced no epileptogenic effects and both immunosuppressants were without effect on the induction of kainate-induced seizures, administration of CsA to excitotoxin-susceptible mice (FVB/N) virtually eliminated neuronal cell death. In contrast, induction of neuronal cell death was evident when CsA was administered to excitotoxin-resistant mice (C57BL/6). Administration of FK-506, another commonly used immunosuppressant, which lacks an effect on the MPT, had no effect on modification of susceptibility to kainate-induced cell death in either strain. CONCLUSIONS: As our data show differential protection of hippocampal neurons against excitotoxic cell death by pretreatment with CsA, these results suggest that strain-dependent differences in mitochondrial integrity and function may exist.     

Mapping a mouse limbic seizure susceptibility locus on chromosome 10

Purpose: Mapping seizure susceptibility loci in mice provides a framework for identifying potentially novel candidate genes for human epilepsy. Using C57BL/6J × A/J chromosome substitution strains (CSS), we previously identified a locus on mouse chromosome 10 (Ch10) conferring susceptibility to pilocarpine, a muscarinic cholinergic agonist that models human temporal lobe epilepsy by inducing initial limbic seizures and status epilepticus (status), followed by hippocampal cell loss and delayed‐onset chronic spontaneous limbic seizures. Herein we report further genetic mapping of pilocarpine quantitative trait loci (QTLs) on Ch10. Methods: Seventy‐nine Ch10 F₂ mice were used to map QTLs for duration of partial status epilepticus and the highest stage reached in response to pilocarpine. Based on those results we created interval‐specific congenic lines to confirm and extend the results, using sequential rounds of breeding selectively by genotype to isolate segments of A/J Ch10 genome on a B6 background. Key Findings: Analysis of Ch10 F₂ genotypes and seizure susceptibility phenotypes identified significant, overlapping QTLs for duration of partial status and severity of pilocarpine‐induced seizures on distal Ch10. Interval‐specific Ch10 congenics containing the susceptibility locus on distal Ch10 also demonstrated susceptibility to pilocarpine‐induced seizures, confirming results from the F₂ mapping population and strongly supporting the presence of a QTL between rs13480781 (117.6 Mb) and rs13480832 (127.7 Mb). Significance: QTL mapping can identify loci that make a quantitative contribution to a trait, and eventually identify the causative DNA‐sequence polymorphisms. We have mapped a locus on mouse Ch10 for pilocarpine‐induced limbic seizures. Novel candidate genes identified in mice can be investigated in functional studies and tested for their role in human epilepsy.     

Strain differences in seizure-induced cell death following pilocarpine-induced status epilepticus

Mouse strains differ from one another in their susceptibility to seizure-induced excitotoxic cell death. Previously, we have demonstrated that mature inbred strains of mice show remarkable genetic differences in susceptibility to the neuropathological consequences of seizures in the kainate model of status epilepticus. At present, while the cellular mechanisms underlying strain-dependent differences in susceptibility remain unclear, some of this variation is assumed to have a genetic basis. However, it remains unclear whether strain differences in susceptibility to seizure-induced cell death observed following kainate administration are observed following systemic administration of other chemoconvulsants. In rodents, the cholinomimetic convulsant pilocarpine is widely used to induce status epilepticus (SE), followed by hippocampal damage and spontaneous recurrent seizures, resembling temporal lobe epilepsy. This model has initially been described in rats, but is increasingly used in mice. We characterized neuronal pathologies after pilocarpine-induced status epilepticus (SE) in eight inbred strains of mice focusing on the hippocampus. A ramping-up dose protocol for pilocarpine was used and behavior was monitored for 4-5 h. While we did not observe any significant differences in seizure latency or duration to pilocarpine among the inbred strains, we did observe a significant difference in susceptibility to the neuropathological consequences of pilocarpine-induced SE. Of the eight genetically diverse mouse strains screened for pilocarpine-induced status, BALB/cJ and BALB/cByJ were the only two strains that were resistant to the neuropathological consequences of seizure-induced cell death. Additional studies of these murine strains may be useful for investigating genetic influences on pilocarpine-induced status epilepticus.