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G. Tosini, K. Baba, S. Contreras-Alcantara; Circadian Rhythms in the Retina and Retinal Pigmented Epithelium of Per2:luc Mice. Invest. Ophthalmol. Vis. Sci. 2009;50(13):3232.
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Daily phagocytosis by the retinal pigmented epithelium (RPE) is essential for the function and viability of the photoreceptors. Previous in vivo studies have shown that this phenomenon is under control of a circadian clock. The location of the circadian pacemaker controlling this important circadian rhythm is still unknown. The goal of this study was to investigate whether cultured retina and RPE-choroid obtained from mPer2:Luc mice exhibited circadian rhythms in bioluminescence levels .
Eyes were obtained from mPer2:Luc mice and the retina and choroid cup containing RPE were dissected in Hank’s salt solution. The retina and RPE- choroid cup were carefully placed on the culture membrane (Millicell-CM pore size 0.4µm) in a 35 mm Petri dish with 1.2ml of 199 medium containing with 0.1mM D-Luciferin K salt, penicillin/streptomycin cocktail and B27. The bioluminescence emitted by the cultured retina and RPE-choroid was measured using a luminometer (Lumicycle, Actimetrics) for 10 days. Then, to test the effect of light on the circadian rhythm the retina and RPE-choroid explants were subjected to a pulse of light at different time of day.
Both the retina and RPE-choroid explants showed robust circadian rhythms in the bioluminescence levels (mean free-running period 24.3±0.03 SEM and 23.7±0.02 SEM, respectively). In the retina exposure to 10 minutes of light (1000 lux) during the subjective day or early night did not induce a phase-shift of the circadian rhythm whereas exposure to light in the late night induced significant phase-shifts (advance and delay). No significant phase-shifts were observed in RPE-choroid explants exposed to light at any time of the day.
Our data indicate that the retina and the RPE-choroid exhibited robust circadian rhythms, thus suggesting that the RPE contains an autonomous circadian clock. However light was able to phase-shift (i.e., synchronize) the circadian rhythm only in the retina.
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