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A. Tsin, A. Muniz, A. Trevino, R. Roman, B. S. Betts, E. Villazana-Espinoza; A Study on the Isomerase Activity of the Cone-Dominated Chicken Retina. Invest. Ophthalmol. Vis. Sci. 2008;49(13):3529.
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© ARVO (1962-2015); The Authors (2016-present)
Recently a novel cone visual cycle has been reported in the cone -dominated chicken retina. The regeneration of visual chromophore involves the action of an isomerase enzyme to produce 11-cis retinoids. Past studies have suggested that chicken retina has a retinol isomerase (Isomerase II) which isomerizes retinol at the alcohol oxidation. The purpose of this research is to study the nature of the isomerase activity in support of the cone cycle in the chicken retina.
Retinas were dissected under dim light from dark-adapted chicken heads. Retinas were placed in Tyrode’s buffer at 25 °C and exposed to light (700 lux) for 10 min. and then dark-adapted for 10 min. Isomerase assays were conducted in retina and RPE microsomes (200 µg) incubated with 20 µM ATOL, 20 µM ATRP or 20 µM ATOL and 100 µM palmitoyl CoA (palm CoA), in the presence of CRALBP. Retinoids were extracted by hexane (or by formaldehyde/hexane, for retinals) and analyzed by HPLC.
Dark adapted chicken retinas contained 0.2 nmol/mg of 11-cis retinal. Light adaptation for 10 min. completely depleted this retinal chromophore. Subsequent 10 min of dark adaptation resulted in 70% regeneration (i.e. 11-cis retinal, 0.14 nmol/mg). Biochemical assays indicate that bovine and chicken RPE microsomes had similar Isomerase I activity (16.6 and 11.8 pmol/min/mg respectively) and this isomerase I activity is absent in bovine and chicken retina microsomes. Isomerase II activity was assayed in the presence of palm CoA and all-trans retinol in chicken retina microsomes. 11-cis RE was formed and this isomerase activity was 2.0 pmol/min/mg. In the absence of palm CoA no isomerase activity was detected.
Isolated chicken retina regenerated 70% of its visual chromophores after 10 min. of dark-adaptation, indicating chromophore regeneration. This pigment regeneration must be supported by the activity of an isomerase enzyme in the retina. Biochemical studies show that chicken retina does not have Isomerase I activity. An palm-CoA dependent isomerase activity, using all-trans retinol as the substrate, was observed in the chicken retina. Our results suggest that the cone cycle in the chicken retina is not supported by the activity of the isomerase I (hydroisomerase or RPE65). Rather, it is support by activity of another isomerase (isomerase II?) which remains to be further elucidated.
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