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J. J. Alexander, J. R. O'rourke, C. L. Cepko, M. T. McManus, A. P. Jadhav, W. W. Hauswirth, B. D. Harfe; Investigating the Role of MicroRNA’s in the Mouse Retina: I. Electrophysiology. Invest. Ophthalmol. Vis. Sci. 2007;48(13):2918.
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Many fundamental processes in development are regulated by microRNA’s. The enzyme Dicer is responsible for processing microRNA to their mature form and removal of dicer has been shown to disrupt many developmental processes. In order to study the implications of removing microRNA’s in the retina we generated mice that have Dicer conditionally knocked out within cells that express Chx10.
We crossed mice expressing Cre recombinase under the Chx10 promoter (Chx10cre) with mice containing a floxed dicer allele (dicerflox) to generate homozygous dicer knockout (dicerflox/flox;Chx10cre), heterozygous (dicerflox/+;Chx10cre), and wildtype (dicer +/+;Chx10cre) mice. The mice were backcrossed for at least 3 generations to C57/B6. The resulting knockout mouse has Dicer removed in the descendents of Chx10 expressing cells, which includes all retinal cell types. We measured the electroretinograms (ERG’s) of these mice at 1 and 5 months under scotopic and photopic conditions. To assay for microRNA reduction, we examined the expression levels of the microRNA miR-183 in P16 whole mouse retinas by microRNA northern blot, as miR-183 has been shown to be expressed in the retina.
We found that the knockout mice had a decrease in both scotopic and photopic ERG amplitudes when compared to heterozygous and/or wild type mice, amounting to a loss of 70% or greater. We also found that the levels of miR-183 were decreased by 27% in the knockout mice when compared to wild type.
To our knowledge, this is the first study to show the importance of microRNA’s for the development of normal electrophysiological functioning in the retina. Furthermore, the reduction in miR-183 expression demonstrates that microRNA’s are reduced upon removal of Dicer in the mouse retina.
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