Canine Retina Has a Primate Fovea-Like Bouquet of Cone Photoreceptors Which Is Affected by Inherited Macular Degenerations

Retinal areas of specialization confer vertebrates with the ability to scrutinize corresponding regions of their visual field with greater resolution. A highly specialized area found in haplorhine primates (including humans) is the fovea centralis which is defined by a high density of cone photoreceptors connected individually to interneurons, and retinal ganglion cells (RGCs) that are offset to form a pit lacking retinal capillaries and inner retinal neurons at its center. In dogs, a local increase in RGC density is found in a topographically comparable retinal area defined as the area centralis. While the canine retina is devoid of a foveal pit, no detailed examination of the photoreceptors within the area centralis has been reported. Using both in vivo and ex vivo imaging, we identified a retinal region with a primate fovea-like cone photoreceptor density but without the excavation of the inner retina. Similar anatomical structure observed in rare human subjects has been named fovea-plana. In addition, dogs with mutations in two different genes, that cause macular degeneration in humans, developed earliest disease at the newly-identified canine fovea-like area. Our results challenge the dogma that within the phylogenetic tree of mammals, haplorhine primates with a fovea are the sole lineage in which the retina has a central bouquet of cones. Furthermore, a predilection for naturally-occurring retinal degenerations to alter this cone-enriched area fills the void for a clinically-relevant animal model of human macular degenerations

Photoreceptor layer lamination in wildtype dogs and in naturally-occuring genetic diseases primarily affecting the canine fovea-like area.

<p>(a,d,f) <i>En face</i> infrared view of representative wildtype (a), <i>BEST1</i>-mutant (d) and <i>RPGR</i>-mutant (f) dogs. *, fovea-like area. Arrows, locations of cross-sectional OCT scans shown below each panel. Outer photoreceptor nuclear layer (ONL) and retinal pigment epithelium (RPE) are highlighted for visibility on OCT scans. (b) ONL thickness topography in a 22-wk-old wildtype dog displayed in pseudo-color. There is a distinct localized region of ONL thinning supero-temporal (ST) to the optic nerve head (black circle) corresponding to the fovea-like area. Major blood vessels are overlaid. (c) Diagonal profiles of ONL thickness (along arrow shown in b) in individual wildtype dogs (lines; ages: 7 wks –8 yrs; n = 13). 95% confidence interval shown (gray area). The break in the axis corresponds to the optic nerve head which lacks photoreceptors. F, fovea-like area. (e) Topographic localization of the sites (*) of the early macular lesions in <i>BEST1</i>-mutant dogs (ages: 10–62 wks; n = 7, left) correspond to the localization of the fovea-like area in wildtype dogs (ages: 7 wks –8 yrs; n = 13, right). (g) ONL thickness topography in an 11-wk-old <i>RPGR</i>-mutant dog displayed in pseudo-color. *, fovea-like area. (h) Diagonal profiles of ONL thickness in young <i>RPGR</i>-mutant dogs (lines; age: 11 wks; n = 6 eyes of 3 dogs) shows abnormal thinning corresponding to the fovea-like area and its immediate surrounds compared to WT (gray area). All eyes are shown as left eyes (temporal retina to the right).</p

High density of cone photoreceptors at the wildtype canine fovea-like area.

<p>(a,b) Central areas of specialization (white) are avoided by retinal blood vessels in human and canine eyes. (c) Retinal ganglion cell (RGC) density map across the canine retina and peak density at the center of the area of specialization. Brn3a: brain-specific homeobox/POU domain protein 3A (d) Retinal cross-section (H&E stained) through the fovea-like area of a 6 week-old dog shows a focal elevation on the retinal surface, thickening of the ganglion cell layer (GCL), and thinning of the outer nuclear layer (ONL). Immunohistochemistry at 7 weeks shows focal high density of cones (red), markedly reduced density of rods (green), elongated inner segments (IS), outer segments (OS) and multiple layers of RGCs in GCL. CA: cone arrestin; Rho: Rhodopsin. ONL is stained with DAPI (blue). (e) Abrupt increase of cone density associated with an abrupt decrease of rod density in 4 eyes (at different postnatal ages for later comparison with mutant dogs). Rod and cone nuclei are highlighted in enlarged insets with blue and yellow, respectively, for visibility. (f) Two-photon microscopy imaging of the fovea-like area and immediate surrounding region. <i>En face</i> view (XY) is an overlay of two Z scans taken at different depths (shown as dotted lines on the orthogonal XZ view) to illustrate the cone IS densities at the fovea-like area and surrounding regions. Insets illustrate the abrupt increase in central peak cone IS density within the canine fovea-like area. vWF: von Willebrand factor VIII; (g) Comparison of peak cone densities in dogs to that reported for macaques and humans measured by adaptive optics imaging, or histology Filled symbols are mean±sd.</p

Histolopathology at the fovea-like area in two canine models of inherited macular degeneration.

<p>(a) Fovea-like area in a 112 week-old <i>BEST1</i>-mutant dog. (a₁) Epifluorescence microscopy image (with DIC/Nomarski optics) showing at the fovea-like area increased autofluorescence (yellow) in the retinal pigment epithelium (RPE). (a₂) Immunohistochemistry on the same section as (b₁) shows focal separation (diamond) of cone (red) outer segments (OS) from the underlying RPE. Note: Red fluorescent signal originating from the RPE is endogenous autofluorescence (see a₁). (a₃) Immunohistochemistry shows focal separation (diamond) of rod OS (green) from hypertrophied RPE cells (red; arrowheads), and (a₄) extension of RPE apical processes (arrowheads). (b) Fovea-like areas in <i>RPGR</i> mutant dogs. (b₁) Horizontal retinal cross-section (H&E stained) shows the abrupt ONL thinning and shortened inner segments (IS) at the fovea-like area of a 4 week-old mutant dog while ONL thickness and structure of photoreceptors is preserved in the immediate peri-foveal regions. (b₂) Horizontal retinal cross-section (H&E stained) shows more prominent ONL thinning at the fovea-like area which has now extended peri-foveally in a 22 week-old dog. (b3) Early reduction in the number of cones is seen at the fovea-like area on retinal cross sections (left; cone nuclei are highlighted in yellow for visibility) and quantitative comparison to wildtype results (right). CA: cone arrestin; Rho: rhodopsin.</p