These mice carry a conditional knock-in allele of the mouse Glra1 gene. When crossed with a cre strain, expression of the wildtype gene is replaced with that of a KK385-386AA modulatory site mutation in cre-expressing tissues of the offspring. The modified sequence has been associated with the sedative effects produced by intoxicating doses of ethanol.
Gregg E Homanics, University of Pittsburgh
Glycine receptors are linked to the actions of alcohol and anesthetics and appear to play prominent roles in disorders such as tremor, seizures, and hyperekplexia. The contribution of G protein regulation of glycine receptor function to these processes/disorders is largely unknown.
These mice carry a conditional knock-in allele of the mouse Glra1 (glycine receptor, alpha 1 subunit) gene. When crossed with a cre strain, expression of the wildtype gene can be replaced with that of a KK385-386AA (lysine to alanine) modulatory site mutation in tissues of interest.
After crossing with a global cre deleter (EIIa-cre mice, Stock No. 003724) to completely delete the floxed wild type exon, global knock-in mice display largely normal behavior and expression as determined by Western blot. Immunocytochemistry studies also indicate that expression is similar to that of wildtype. Knock-in mice exhibit a lower sensitivity to ethanol, as demonstrated by shorter times in loss of righting reflex (LORR) in response to a sedative dose of ethanol (3.5g/kg). Their glycinergic synaptic transmission has normal properties, but their sensitivity to ethanol is significantly reduced. Data links Glra1 amino acids 385-386 with the sedative effects produced by intoxicating doses of ethanol.
A targeting construct was used to introduce a conditional knock-in to an exon 9 modulatory site in the mouse Glra1 gene: loxP-exon 9-FRT-Neo-FRT-loxP-(KK385-386AA mutant exon 9). The KK385-386AA (lysine to alanine) mutation represents an AAGAAG to GCCGCC sequence modification. The mutation was created through homologous recombination in AB2.2 129S7/SvEvBrd-Hprt1+-derived embryonic stem (ES) cells. Resulting chimeric males were crossed with C57BL/6 females and offspring were crossed to Actin-Flp mice on a C57BL/6 genetic background
(see Stock No. 003800 backcrossed to C57BL/6 for 15 generations) to remove the neomycin cassette. Animals harboring the targeted locus but lacking the neomycin cassette were backcrossed to C57BL/6J for 5 generations by the donating laboratory.
|Allele Name||targeted mutation 3.1, Gregg E Homanics|
|Allele Type||Targeted (Inserted expressed sequence)|
|Gene Symbol and Name||Glra1, glycine receptor, alpha 1 subunit|
|Site of Expression||When crossed with a cre strain, expression of the wildtype gene is replaced with the mutated gene in cre-expressing tissues of the offspring|
|Strain of Origin||129S7/SvEvBrd-Hprtb-m2|
|Molecular Note||A targeting construct was used to introduce a conditional knock-in to an exon 9 modulatory site (loxP-exon 9-FRT-Neo-FRT-loxP). The KK385-386AA (lysine to alanine) mutation represents an AAGAAG to GCCGCC sequence modification. Flp-mediated recombination removed the neomycin resistance cassette.|
Heterozygous and homozygous mice are viable and fertile.
When using the B6.129S7(SJL)-Glra1tm3.1Geh/J mouse strain in a publication, please cite the originating article(s) and include JAX stock #023516 in your Materials and Methods section.