A gold loop electrode was placed on the surface of the cornea and referenced to a gold wire placed in the mouth. 24 Briefly, anesthesia was induced by intraperitoneal injection of a mixture of ketamine (70 mg/kg) and xylazine (15 mg/kg), and body temperature (36.5☌-37☌) was maintained with a temperature-controlled heating pad. 22, 23įull-field ERG recordings were performed as previously described. 21 Photoreceptor death in these Mertk mutants is associated with an inability of the RPE to properly phagocytize shed photoreceptor outer segment tips. 20 This murine allele closely recapitulates the RCS rat phenotype of rapid photoreceptor degeneration. 18, 19 Additionally, a targeted mutation of an exon encoding a portion of the Mer tyrosine kinase domain in Mertk tm1Gkm mice has been reported. 15 – 17 Mapping efforts determined that a deletion within the Mertk gene ( Mertk rdy) led to a frameshift and a termination signal 20 codons after the start codon in RCS rats. 10 – 14 The Royal College of Surgeons (RCS) rat is a classic model for assessing photoreceptor-RPE interaction. 9 Multiple instances of retinitis pigmentosa or allied disorders caused by mutations in the MERTK gene have been reported. Its importance is underscored by the large number of diseases associated with improper function of the RPE, such as macular edema, 1 Best's vitelliform macular degeneration, 2 – 4 autosomal recessive macular degeneration, 5 Leber congenital amaurosis, 6 – 8 and Stargardt disease. Proper function of cells in the retinal pigment epithelium (RPE) is required for normal vision and for the survival of photoreceptor cells. These results demonstrate that TNF family members play a role in protecting photoreceptors of Mertk nmf12 homozygotes from cell death. These findings illustrate that a mutation in the Mertk gene leads to a significantly slower progressive retinal degeneration compared with other alleles of Mertk. Surprisingly, mice homozygous for both the Mertk nmf12 and the Ltb/Tnf/Lta tm1Dvk allele ( Tnfabc −/−) demonstrated an increase in the rate of retinal degeneration. 2 B6.129P2- Ltb/Tnf/Lta tm1Dvk/J homozygotes did not exhibit a retinal degeneration phenotype and will, hereafter, be referred to as Tnfabc −/− mice. Mertk nmf12 homozygous mice were mated to mice lacking the entire Tnf gene and partial coding sequences of the Lta ( Tnfb) and Ltb ( Tnfc) genes. We detected elevated levels of tumor necrosis factor ( Tnf, previously Tnfa) in retinas of Mertk nmf12 homozygotes relative to wild-type controls and investigated whether the increase of TNF, an inflammatory cytokine produced by macrophages/monocytes that signals intracellularly to cause necrosis or apoptosis, could underlie the retinal degeneration observed in Mertk nmf12 homozygotes. Sequencing of nmf12 DNA revealed a point mutation in the c-mer tyrosine kinase gene, designated Mertk nmf12. The mutation was named neuroscience mutagenesis facility 12 ( nmf12), and mapping localized the critical region to Chromosome 2. Longitudinal studies of retinal histologic sections showed photoreceptors in the peripheral retina undergoing slow, progressive degeneration. Screening by indirect ophthalmoscopy identified a line of N-ethyl- N-nitrosourea (ENU) mutagenized mice demonstrating retinal patches. To determine the basis and to characterize the phenotype of a chemically induced mutation in a mouse model of retinal degeneration.
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