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L. R. Blakeley, C. Chen, R. K. Crouch, Y. Koutalos; Rapid Formation of All-Trans Retinol After Bleaching in Photoreceptors of Nrl-/- Mice. Invest. Ophthalmol. Vis. Sci. 2008;49(13):151.
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© ARVO (1962-2015); The Authors (2016-present)
To examine the formation of all-trans retinol after bleaching in mammalian photoreceptors that contain cone pigments. The photoreceptors in the retinas of Nrl-/- mice are cone-like and contain only cone pigments. Most of the light sensitivity of the Nrl-/- retina is due to a cone pigment with max ~ 360 nm.
Experiments were carried out with dark-adapted isolated retinas and isolated photoreceptor cells from Nrl-/- mice. Animals were 2-3 months old. All-trans retinol formation after pigment bleaching (10 sec of 360 nm light) was measured by quantitative HPLC of retinoids extracted from isolated retinas and by imaging the fluorescence of retinol (Ex: 360 nm; Em: >420 nm) in the outer segments of isolated photoreceptors. Experiments were carried out at 37 0C.
Bleaching of dark-adapted isolated retinas converted approximately half of the 11-cis retinyl chromophore to all-trans. Afterwards, the fraction of retinol in the all-trans retinoid pool increased with a time constant of 0.35 min and reached a value of 0.75 that remained stable for at least one hour. Bleaching of dark-adapted isolated cells resulted in an increase in retinol fluorescence in the outer segment. Subsequently, outer segment fluorescence declined with a time constant of approximately 5 min, indicating the elimination of retinol.
The formation and elimination of all-trans retinol in the outer segments of Nrl-/- mouse photoreceptors, which contain cone pigments, proceed much more rapidly than in rods, consistent with a faster regeneration of the visual pigment after bleaching. The fraction of all-trans retinal converted to retinol at equilibrium is similar to that in rod photoreceptors. This suggests that the Nrl-/- photoreceptors contain a comparable fraction of reduced NADPH as rods to drive the reduction of all-trans retinal.Support: NIH/NEI grants EY04939, EY14850, and an unrestricted grant to MUSC Storm Eye Institute from Research to Prevent Blindness, Inc., New York, NY. RKC is a RPB Senior Scientific Investigator.
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