Removal of the circadian clock gene Bmal1 in the RPE abolishes the daily peak of photoreceptor outer segment disc phagocytosis

Abstract

Purpose : Circadian regulation of both the retina and retinal pigment epithelium (RPE) is important to maintain healthy visual function. Previous studies have shown a rhythmic peak in phagocytosis of photoreceptor outer segments (POS) discs by RPE one or two hours after the onset of light and it is believed that the timing of this peak is important to maintain photoreceptor health. While this function of the RPE has been demonstrated to be under circadian control, it is not clear whether a circadian clock in the retina and/or RPE controls the timing in the peak of phagocytosis. In this study we investigated how disruption of the RPE circadian clock affects the daily rhythm of phagocytic activity and photoreceptor health.

Methods : RPE specific disruption of Bmal1 was accomplished using the cre-lox system with a RPE-specific cre recombinase driver (rtTA-VMD2/Cre [RPE^cre]) and the floxed Arntl gene (Bmal1^fl/fl). Following confirmation and validation of doxycycline (dox)-inducible cre recombinase activity in RPE^cre;Bmal1^fl/fl mice (dox diet fed at p60-p74), we investigated the effect of Bmal1 removal on the daily peak of phagocytic activity by counting the number of phagosomes containing POS discs localized in the RPE at three different time points (1 hour before light onset and 1 and 3 hours after light onset). Retinal structure and function were investigated via SD-OCT (spectral domain-optical coherence tomography), ERG (electroretinogram), and OMR (opto-motor response) testing.

Results : RPE^cre;Bmal1^fl/fl mice did not show the typical burst in phagocytic activity by RPE cells 1 hour after the onset of light. Conversely, control mice or retina-(Chx10^cre) Bmal1^fl/fl deficient mice showed a clear increase in phagocytic activity 1 hour after light onset. Interestingly, both retinal structure (SD-OCT) and function (ERG and OMR) were not affected by removal of Bmal1 from the RPE in either young (3 months) or old (12 months) mice.

Conclusions : Our data indicates that the daily peak in POS disc phagocytosis is controlled by the circadian clock located in the RPE and the contribution of the retinal circadian clock seems to be negligible. The apparent lack of effect of RPE Bmal1 disruption on retinal structure and function suggests that the circadian clock in the RPE does not significantly affect photoreceptor survival, up to 10 months after loss of Bmal1 activity in the RPE.

This abstract was presented at the 2019 ARVO Annual Meeting, held in Vancouver, Canada, April 28 - May 2, 2019.