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Beatrice M Tam, Orson L Moritz; Repeated synchronous photoactivation of T4K rhodopsin induces outer segment shedding and retinal degeneration in a Xenopus laevis model of retinitis pigmentosa. Invest. Ophthalmol. Vis. Sci. 2016;57(12):2252.
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© ARVO (1962-2015); The Authors (2016-present)
The rhodopsin mutation T4K is associated with sector RP in humans and causes light dependent retinal degeneration (RD) in transgenic X. laevis. Our objective is to elucidate the pathogenic mechanisms underlying the light dependence of this RP model.
Transgenic lines of X. laevis were mated to produce tadpoles expressing human T4K opsin in their rods. Transgenic tadpoles and their nontransgenic siblings were reared in varying light regimens in which we altered the light cycle frequency and/or light intensity. Following light exposure, animals were killed and eyes were processed for confocal or electron microscopy or quantitative protein analysis.
In transgenic X. laevis expressing mutant T4K opsin, severe RD is induced by 12h:12h cyclic light. Both constant dark and constant light were protective relative to cyclic light. Increasing the frequency of cycling increased the rate of RD. In animals exposed to 1h:1h cyclic light, RD was largely complete within 48 hours. Further titration of the duration of light and dark periods demonstrated that a minimum of 20 min. of light exposure and 60 min. of darkness were required to induce maximum RD. Under the 12h:12h cyclic light paradigm, instantaneous light onset at the start of each light cycle was more damaging than gradual light onset (from 0 to 1700 lux over 2hrs). Pre-treatment with constant light prior to exposure to 1h:1h cyclic light was also protective. Transgenic eyes exposed to 1h:1h cyclic light exhibited extensive shedding of rod outer segments (OS) beginning approximately 13 hours after the first light exposure. This wave of shedding preceded any gross abnormalities in the inner segments. OS disk structure remained well organized, although some subtle perturbations of disk structure were noted.
Our results indicate that T4K rhodopsin-induced RD requires repeated synchronized activation of the mutant rhodopsin present in rod OS. Reducing synchronous activation (by gradual light onset, or constant light) resulted in less severe RD. The protective effect of prolonged light exposure may indicate instability of T4K rhodopsin to multiple cycles of photobleaching. Our results also indicate that T4K rhodopsin activation causes significantly increased shedding of rod OS, which may be linked to the mechanism of cell death in this model.
This is an abstract that was submitted for the 2016 ARVO Annual Meeting, held in Seattle, Wash., May 1-5, 2016.
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