Preservation of Cone Photoreceptors after a Rapid yet Transient Degeneration and Remodeling in Cone-OnlyNrl^−/−Mouse Retina

Abstract

Cone photoreceptors are the primary initiator of visual transduction in the human retina. Dysfunction or death of rod photoreceptors precedes cone loss in many retinal and macular degenerative diseases, suggesting a rod-dependent trophic support for cone survival. Rod differentiation and homeostasis are dependent on the basic motif leucine zipper transcription factor NRL. The loss of Nrl (Nrl−/−) in mice results in a retina with predominantly S-opsin containing cones that exhibit molecular and functional characteristics of WT cones. Here we report that Nrl−/− retina undergoes a rapid but transient period of degeneration in early adulthood, with cone apoptosis, retinal detachment, alterations in retinal vessel structure, and activation and translocation of retinal microglia. However, cone degeneration stabilizes by four months of age, resulting in a thinner but intact outer nuclear layer with residual cones expressing S- and M-opsins and a preserved photopic ERG. At this stage, microglia translocate back to the inner retina and reacquire a quiescent morphology. Gene profiling analysis during the period of transient degeneration reveals misregulation of genes related to stress response and inflammation, implying their involvement in cone death. The Nrl−/− mouse illustrates the long-term viability of cones in the absence of rods and RPE defects in a rodless retina. We propose that Nrl−/− retina may serve as a model for elucidating mechanisms of cone homeostasis and degeneration that would be relevant to understanding diseases of the cone-dominant human macula.

OTX2 loss causes rod differentiation defect in CRX-associated congenital blindness

“…The long-term preservation of immature and dysfunctional cones in Crx Rip/+ mutant retinas in the absence of rod photoreceptors is consistent with our recent studies (37,50) and provides opportunities for elucidating cone survival pathways and designing treatment paradigms for degenerating retinal diseases. At this stage, it is unclear whether dominant CRX frameshift mutations arrest photoreceptor differentiation in humans, as disease onset is very early and OCT studies are difficult to perform.…”

“…The cone-specific proteins -S-opsin (OPN1SW), M-opsin (OPN1MW), and cone arrestin (ARR3) -were detected in few cells in P21 Crx -/-retinas; however, these markers were completely absent in the Crx Rip/+ and Crx Rip/Rip retinas. Furthermore, Crx Rip/+ retinas revealed very strong and continuous staining for peanut agglutinin (PNA), a cone-specific cell surface marker, similar to that in Nrl -/-retinas (36,37). Immunostaining of recoverin, an early marker of photoreceptor differentiation, was also dramatically reduced in Crx Rip/+ mice compared with that in WT mice, with only a few recoverin-positive cells in Crx Rip/Rip mutants.…”

“…Mice of either sex were used for the study and euthanized by CO2 inhalation. The procedures for tissue preparation for cryopreservation, methacrylate sections, and RNA/protein extraction have been described earlier (37,53), and additional details are provided in the Supplemental Methods.…”

“…Pupils were dilated using topical 0.5% tropicamide and 1% cyclopentolate hydrochloride. The methods for fundus examination, OCT imaging, and ERG have been described previously (37).…”

Reprogramming of adult rod photoreceptors prevents retinal degeneration

“…We reasoned that acute inactivation of Nrl in adult rods might result in direct conversion of these cells into cones. Furthermore, a recent study demonstrated that retinas in which Nrl had been knocked out during development showed longterm survival of cone photoreceptors and preservation of the outer nuclear layer, after a transient initial phase of cell loss (19). This observation suggests that direct conversion of adult rods into cones could also lead to long-term survival of the transdifferentiated cells.…”