Quantifying cones in outer retinal tubulation (ORT) in age-related macular degeneration (AMD) from spectral domain optical coherence tomography (SD-OCT)

Abstract

 

Purpose

 

By SD-OCT, ORT is a hyper-reflective border surrounding a hypo-reflective area in the outer nuclear layer (Figure A) {1}. By histology, ORT is a formation of cone photoreceptors and Müller cells with a lumen delimited by the external limiting membrane (ELM) (Figure B) {2}. To inform future adaptive optics assisted retinal imaging of ORT, we use SD-OCT volume scans from patients with AMD to assess ORT dimensions and donor eye histology to assess density and number of cones.

 

Methods

 

Ten SD-OCT volumes (Heidelberg Spectralis) from 9 patients (82.1 ± 8.2 years, 7 women, 2 men) with advanced AMD and ORT were analyzed with custom acquisition and reconstruction software. The inner and outer aspect of the hyper-reflective ORT border in cross-section was delineated with polygons. To approximate a surface at the ELM, a surface 5 µm abluminal from the inner polygon was calculated. To visualize the ELM cytoskeleton meshwork through which ORT cones point into the lumen, a flat mount retina from an 86-year-old woman with neovascular AMD was labeled with phalloidin {3} (Figure C). From cones in hexagonal cross-section, area and center-to-center spacing was measured with ImageJ. ORT cone density is estimated by ORT surface area divided by mean area of a single cone.

 

Results

 

The surface areas of partial ORT assessed by SD-OCT volumes range from 0.16 to 1.76 mm². From the flat mount, the average cross-sectional area of a cone is 49.1 ± 7.9 µm². The average cone center-to-center spacing is 7.5 ± 0.6 µm, assuming triangular packing. ORT cone density is 20,351 cones/mm². The number of cones in these partial ORT ranges from 3,212 to 35,743.

 

Conclusions

 

We provide the first estimates for the density and number of cones surviving in ORT. Compared to cones in the healthy macula {4}, cone density in ORT is decreased relative to the fovea and increased relative to perifovea, suggesting considerable distortion of the photoreceptor mosaic. Longitudinal monitoring of ORT change may provide deep insight into cone degeneration and disease progression. 1. Zweifel PMID 20008714. 2. Schaal Retina in press. 3. Ach PMID 25034602 4. Curcio PMID 2324310.  

 

Figure. Imaging of ORT (yellow arrows) in SD-OCT (A), histology (B), and flat mount retina (C). ONL, outer nuclear layer. IS, inner segment, BrM, Bruch’s membrane. Phalloidin, red. Lipofuscin by 488 nm autofluorescence, green.