Characterization of Retinal Chemistry and Function of Fat–1 Transgenic Mice Maintained on an N–6 Polyunsaturated Fatty Acid Enriched Diet

Abstract

Purpose: : Enzymatic expression of the fat–1 transgene results in the conversion of n–6 polyunsaturated fatty acids to n–3 polyunsaturated fatty acids. We sought to examine the effects of fat–1 enzymatic activity on the retinal chemistry and function of fat–1 transgenic mice.

Methods: : Fat–1 C57BL/6J mice were bred onto a Balb/c background and utilized alongside their fat–1 negative siblings (controls). Prior to breeding, dams were fed a semi–synthetic diet consisting of 10% (wt/wt) safflower oil (n–6/n–3 ratio of 273). Pups were maintained on this diet in <30 lux cyclic light (12L:12D) until tissues were harvested at six weeks of age. Four days prior to harvesting tissues, electroretinograms (ERG) were obtained to evaluate retinal function. Photoreceptor outer segments were prepared by discontinuous sucrose density centrifugation and evaluated by polyacrylamide gel electrophoresis (PAGE). Fatty acid profiles of outer segments were determined by gas–liquid chromatography. Rhodopsin measurements of whole eye were obtained on dark–adapted animals. Multivariant ANOVA with post–hoc Neuman–Keuls tests determined statistical significance (p<0.05).

Results: : Fatty acid analysis revealed fat–1 outer segments to be significantly enriched in 20:5n3, 22:5n3 and 22:6n3 in comparison to control animals, which were enriched in 20:4n6, 22:4n6 and 22:5n6. Differences in 18:2n6 (the primary fat constituent of the diet) were insignificant. The n6/n3 ratios of fat–1 and control outer segments were 0.2 and 1.5 respectively. The protein profiles determined by PAGE showed no observable differences and there were no significant differences in ERG amplitudes or rhodopsin concentrations.

Conclusions: : Fat–1 enzymatic activity does significantly affect the retinal fatty acid content of mice fed an n–6 polyunsaturated fatty acid enriched diet. However, retinal function, as measured by ERG, was not significantly altered.

Keywords: lipids • gene/expression • photoreceptors