Single and multiple congenic strains for hydrocephalus in the H-Tx rat

Abstract

The H-Tx rat has fetal-onset hydrocephalus with a complex mode of inheritance. Previously, quantitative trait locus mapping using a backcross with Fischer F344 rats demonstrated genetic loci significantly linked to hydrocephalus on Chromosomes 10, 11, and 17. Hydrocephalus was preferentially associated with heterozygous alleles on Chrs 10 and 11 and with homozygous alleles on Chr 17. This study aimed to determine the phenotypic contribution of each locus by constructing single and multiple congenic strains. Single congenic rats were constructed using Fischer F344 as the recipient strain and a marker-assisted protocol. The homozygous strains were maintained for eight generations and the brains examined for dilated ventricles indicative for hydrocephalus. No congenic rats had severe (overt) hydrocephalus. A few pups and a significant number of adults had mild disease. The incidence was significantly higher in the C10 and C17 congenic strains than in the nonhydrocephalic F344 strain. Breeding to F344 to make F.H-Tx C10 or C11 rats heterozygous for the hydrocephalus locus failed to produce progeny with severe disease. Both bicongenic and tricongenic rats of different genotype combinations were constructed by crossing congenic rats. None had severe disease but the frequency of mild hydrocephalus in adults was similar to congenic rats and significantly higher than in the F344 strain. Rats with severe hydrocephalus were recovered in low numbers when single congenic or bicongenic rats were crossed with the parental H-Tx strain. It is concluded that the genetic and epigenetic factors contributing to severe hydrocephalus in the H-Tx strain are more complex than originally anticipated.

Fig. 1

Breeding scheme to make bicongenic and tricongenic rats from single strain rats. Single congenic C10 or C11 rats were crossed to C17 rats to make heterozygous bicongenic rats (Stage I). The heterozygous rats were intercrossed and homozygous progeny were selected to make bicongenic strains (Stage II). To make tricongenic rats, C10, C17 and C11, C17 heterozygous bicongenic rats (Stage I) were crossed and progeny were selected with heterozygous genotypes at all three loci or heterozygous on C10 and C11 but homozygous on C17 (Stage II). These rats were intercrossed and progeny that were homozygous at all three loci were selected to make the tricongenic strains (Stage III). Some were crossed with C17 single congenic rats to make the most favorable genotype for hydrocephalus.

Fig. 2

Chromosome maps of the three congenic strains. The scale on the left is the mapping distance along the chromosome in centimorgans (cM) with the telomeric end at the bottom. The position of the peak LOD score is depicted by the dashed arrow and the confidence limits are depicted by the solid arrow. The microsatellite markers used to map the locus are on the right of the chromosome and the vertical bar indicates the position of the congenic interval. Black fill indicates the H-Tx homozygous region, the checked fill indicates the region where there is recombination between H-Tx and F344, and the white fill is the F344 homozygous region. a. The F.H-TxC10 strain had congenic interval that extended from D10Rat207 to the last marker D10Rat2 and included the hydrocephalus locus between D10Rat 136 and D10Rat135. b. The F.H-TxC11 strain had a congenic interval from D11Rat4 to the last marker mapped and included the locus at D11Arb4. c. F.H-TxC17 strain 1 had a congenic interval from D17Rat 79 to D17Rat42 and included the locus peak at D17Mit4. For strain 2, The congenic interval was longer and extended to the last marker mapped D17Rat154.

Fig. 3

a. The incidence of mild hydrocephalus in adult rats and in pups for the parental and congenic strains. The C10 and C17 congenic strains had significantly more adult rats with mild hydrocephalus than the Fischer F344 strain, p < 0.05 and 0.01, as did the H-Tx strain and the N₁ rats, p < 0.01. Very few congenic pups had mild disease and none occurred in the N₁ or the F344 pups. b. The severity of hydrocephalus for all congenic strain rats. There was no significant difference among the groups. Data are means ±SEM. c. Frequency distribution graph of the hydrocephalus severity for all congenic rats and all H-Tx rats. Most congenic rats have a severity of less than 0.4 and most H-Tx rats have a severity of greater than 0.4. Some H-Tx rats have a severity of less than 0.4 and these tended to be the adult animals.

Fig. 4

Chromosome maps of C10, 11, and 17 for tricongenic rats. The chromosomes are labeled as described for Fig. 3. Note the congenic interval was much longer in tricongenic rats for Chrs 11 and 17. This is because the genetic interval of the founder rats was longer than in those used to found the single congenic strains.