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Previous quantitative trait loci (QTL) mapping studies document that the distal region of mouse Chromosome (Chr) 1 contains a gene(s) that is in large part responsible for the difference in seizure susceptibility between C57BL/6 (B6) (relatively seizure-resistant) and DBA/2 (D2) (relatively seizure-sensitive) mice. We now confirm this seizure-related QTL ( Szs1) using reciprocal, interval-specific congenic strains and map it to a 6.6-Mb segment between Pbx1 and D1Mit150. Haplotype conservation between strains within this segment suggests that Szs1 may be localized more precisely to a 4.1-Mb critical interval between Fcgr3 and D1Mit150. We compared the coding region sequences of candidate genes between B6 and D2 mice using RT-PCR, amplification from genomic DNA, and database searching and discovered 12 brain-expressed genes with SNPs that predict a protein amino acid variation. Of these, the most compelling seizure susceptibility candidate is Kcnj10. A survey of the Kcnj10 SNP among other inbred mouse strains revealed a significant effect on seizure sensitivity such that most strains possessing a haplotype containing the B6 variant of Kcnj10 have higher seizure thresholds than those strains possessing the D2 variant. The unique role of inward-rectifying potassium ion channels in membrane physiology coupled with previous strong association between ion channel gene mutations and seizure phenotypes puts even greater focus on Kcnj10 in the present model. In summary, we confirmed a seizure-related QTL of large effect on mouse Chr 1 and mapped it to a finely delimited region. The critical interval contains several candidate genes, one of which, Kcnj10, exhibits a potentially important polymorphism with regard to fundamental aspects of seizure susceptibility.

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The mouse mu-opioid receptor gene, Oprm1, is recognized currently to contain 17 alternatively spliced exons that generate 24 splice variants encoding at least 11 morphine-binding isoforms of the receptor. Here, we identify three new MOR splice variants that contain a previously undescribed exon, exon 18, and provide evidence that they are expressed in two mouse strains. The transcripts containing the newly identified exon 18 encode two new putative mu-opioid receptor isoforms, MOR-1V and MOR-1W. In mouse Oprm1, exon 18 is located between the described exons 10 and 6. Exon 18 appears to be conserved in the rat genome between exons 4 and 7. A BLAST search of the non-redundant GenBank database suggests that human OPRM1 may also contain exon 18. Analysis of mouse brain mRNA by RT-PCR suggests that MOR-1Vii transcripts are expressed in all areas of the brain analyzed, whereas expression of MOR-1Vi transcripts was restricted to thalamus and striatum. MOR-1W transcripts are expressed most highly in the hypothalamus, thalamus and striatum. In summary, we have identified three brain expressed, alternatively spliced mouse MOR splice variants containing a novel exon and encoding new putative MOR isoforms, MOR-1V and MOR-1W.

The mouse mu-opioid receptor gene, Oprm1, currently contains 18 recognized alternatively spliced exons [Doyle, G.A., Sheng, X.R., Lin, S.S.J., Press, D.M., Grice, D.E., Buono, R.J., Ferraro, T.N., Berrettini, W.H., 2007. Identification of three mouse mu-opioid receptor (MOR) gene (Oprm1) splice variants containing a newly identified alternatively spliced exon. Gene 388 (1-2) 135-147, in press (doi:10.1016/j.gene.2006.10.017). Electronic publication 2006 November 1] that generate 27 splice variants encoding at least 11 morphine-binding isoforms of the receptor. Here, we identify five MOR variants that contain an as yet undescribed exon (exon 19) of the gene, and we provide evidence that these MOR splice variants are expressed in the C57BL/6 and DBA/2 mouse strains. Three splice variants, MOR-1Eii, MOR-1Eiii and MOR-1Eiv, encode the MOR-1E isoform and contain the newly identified exon 19 in their 3' untranslated regions. The fourth splice variant encodes a novel mu-opioid receptor isoform, MOR-1U, and contains exon 19 in its coding region. The cytoplasmic tail of the putative MOR-1U isoform contains a putative nuclear localization signal encoded by the sequence of exon 19. Exon 19 appears to be conserved in the rat, but not in humans. In mouse and rat Oprm1, exon 19 is located between described exons 7 and 8. We also report the cloning of the "full-length" MOR-1T splice variant [Kvam, T.-M., Baar, C., Rakvag, T.T., Kaasa, S., Krokan, H.E., Skorpen, F., 2004. Genetic analysis of the murine mu-opioid receptor: increased complexity of Oprm1 gene splicing, J. Mol. Med. 82 (4) 250-255] that encodes MOR-1 and contains the newly identified exon in its 3' UTR. RT-PCR analysis suggests that splice variants MOR-1Eii, MOR-1Eiii, MOR-1Eiv, MOR-1T and MOR-1U are expressed in all brain regions analyzed (cortex, cerebellum, hypothalamus, thalamus and striatum). These exon 19-containing splice variants add to the growing complexity of the mouse Oprm1 gene.

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