SUMMARY Phagocytosis and degradation of photoreceptor outer segments (POS) by the retinal pigment epithelium (RPE) is fundamental to vision. Autophagy is also responsible for bulk degradation of cellular components but its role in POS degradation is not well understood. We report that the morning burst of RPE phagocytosis coincided with the enzymatic conversion of autophagy protein LC3 to its lipidated form. LC3 then associated with single membrane phagosomes containing engulfed POS in an Atg5 dependent manner that required Beclin1 but not the autophagy pre-initiation complex. The importance of this process was verified in mice with Atg5-deficient RPE cells that showed evidence of disrupted lysosomal processing. These mice also exhibited decreased photoreceptor responses to light stimuli and decreased chromophore levels that were restored with exogenous retinoid supplementation. These results establish that the interplay of phagocytosis and autophagy within the RPE are required for both POS degradation and the maintenance of retinoid levels to support vision.

Purpose Polymorphisms in factor H (fH), an inhibitor of the alternative pathway (AP) of complement activation, are associated with increased risk for age-related macular degeneration (AMD). The authors investigated the therapeutic use of a novel recombinant form of fH, CR2-fH, which is targeted to sites of complement activation, in mouse choroidal neovascularization (CNV). CR2-fH consists of the N terminus of mouse fH, which contains the AP-inhibitory domain, linked to a complement receptor 2 (CR2) targeting fragment that binds complement activation products. Methods Laser-induced CNV was analyzed in factor-B–deficient mice or in mice treated with CR2-fH, soluble CR2 (targeting domain), or PBS. CNV progression was analyzed by molecular, histologic, and electrophysiological readouts. Results Intravenously administered CR2-fH reduced CNV size, preserved retina function, and abrogated the injury-associated expression of C3 and VEGF mRNA. CR2 and PBS treatment was without effect. In therapeutically relevant paradigms involving delayed treatment after injury, CR2-fH was effective in reducing CNV and provided approximately 60% of the amount of protection of that seen in factor B–deficient mice that lacked functional AP. After intravenous injection, CR2-fH localized to sites of C3 deposition in RPE-choroid. Conclusions Specific inhibition of the AP reduces angiogenesis in mouse CNV. Of note, intravenous injection of C3d-targeted CR2-fH is protective even though endogenous fH is present in serum at a higher relative concentration, and serum fH contains native C3d and cell surface binding domains that target it to cell surfaces. The most common AMD-associated variant of fH resides within a native cell-binding region of fH (Tyr402His). These data may open new avenues for AMD treatment strategies.

Properties of fhRPE cells align with a functionally normal RPE in vivo, while ARPE-19 cells resemble a pathologic or aged RPE. These results suggest a utility for both cell types in understanding distinct, particular aspects of RPE function.

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The retinoid cycle is a series of biochemical reactions within the eye that is responsible for synthesizing the chromophore, 11-cis retinal, for visual function. The chromophore is bound to G-protein coupled receptors, opsins, within rod and cone photoreceptor cells forming the photosensitive visual pigments. Integral to the sustained function of photoreceptors is the continuous generation of chromophore by the retinoid cycle through two separate processes, one that supplies both rods and cones and another that exclusively supplies cones. Recent findings such as RPE65 localization within cones and the pattern of distribution of retinoid metabolites within mouse and human retinas have challenged previous proposed schemes. This review will focus on recent findings regarding the transport of retinoids, the mechanisms by which chromophore is supplied to both rods and cones, and the metabolism of retinoids within the posterior segment of the eye.

Rapid regeneration of the visual pigment following its photoactivation is critical for the function of cone photoreceptors throughout the day. Though the reactions of the visual cycle in the retinal pigment epithelium (RPE) that recycle chromophore for rod pigment regeneration are well characterized, the corresponding mechanisms that enable rapid regeneration of cone pigment are poorly understood. A key remaining question is the relative contribution of the recently discovered cone-specific retina visual cycle and the classic RPE-dependent visual cycle to mammalian cone pigment regeneration. In addition, it is not clear what role, if any, the abundant interphotoreceptor matrix protein, IRBP, presumed to facilitate the traffic of chromophore, plays in accelerating mammalian cone pigment regeneration. To address these issues we used transretinal recordings to evaluate M/L-cone pigment regeneration in isolated retinas and eyecups from control and IRBP-deficient mice. Remarkably, the mouse retina promoted M/L-cone dark adaptation 8-fold faster than the RPE. However, complete cone recovery required both visual cycles. We conclude that the retina visual cycle is critical for the initial rapid regeneration of mouse M/L-cone pigment during dark adaptation whereas the slower RPE visual cycle is required to complete the process. While the deletion of IRBP reduced the amplitude and slowed the kinetics of mouse M/L-cone photoresponses, cone adaptation in bright steady light and the kinetics of cone dark adaptation were not affected in isolated retina or in intact eyecup. Thus, IRBP does not accelerate cone pigment regeneration and is not critical for the function of mouse M/L-cones in bright light.

In five experiments we measured the amplitude of the haemodynamic response to visual patterns using near infrared spectroscopy of the visual cortex. The patterns were gratings with bars that differed in chromaticity but not in luminance. In all experiments, with a wide range of chromaticities of the grating bars, the amplitude of the haemodynamic response increased with the separation of the chromaticities in the CIE 1976 UCS diagram. The amplitude did not vary consistently with the cone activation, or with the signal in colour difference channels. In four further experiments, again with a wide range of chromaticities, the gratings were rated for visual comfort. Discomfort increased consistently with the separation of the chromaticities. Given that a large haemodynamic response to patterns is generally associated with headache, we suggest that the discomfort may be a homeostatic signal to reduce sustained metabolic load on the visual cortex.

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