February 26, 2021

Biological Correlations and Confounding Variables for Quantification of Retinal Ganglion Cells Based on Optical Coherence Tomography Using Diversity Outbred Mice

Purpose: Despite popularity of optical coherence tomography (OCT) in glaucoma studies, it is unclear how well OCT-derived metrics compare to traditional measures of retinal ganglion cell (RGC) abundance. Here, Diversity Outbred (J:DO) mice are used to directly compare ganglion cell complex (GCC)-thickness measured by OCT to metrics of retinal anatomy measured ex vivo with retinal wholemounts or optic nerve cross sections. Methods: J:DO mice (n = 48) underwent OCT and fundoscopic exams, with GCC-thickness measured using automated segmentation. Following euthanasia, RGC axons were quantified from para-phenylenediamine-stained optic nerve cross sections and RGC somas from BRN3A-immunolabeled retinal wholemounts with total cellularity assessed by TO-PRO or hematoxylin nuclear staining. Results: J:DO tissues lacked overt disease. GCC-thickness (62.4 {+/-} 3.7 m), RGC abundance (3,097 {+/-} 515 BRN3A+ nuclei/mm2; 45,533 {+/-} 9,077 axons), and total inner retinal cell abundance (6,952 {+/-} 810 nuclei/mm2) varied broadly. GCC-thickness correlated significantly to RGC somal density (r = 0.46) and axon number (r = 0.49), whereas total inner retinal cellularity did not. Retinal area (20.3 {+/-} 2.4 mm2) and optic nerve (0.09 {+/-} 0.02 mm2) cross-sectional area varied widely. Sex did not significantly influence any of these metrics. In bilateral comparisons, GCC-thickness (r = 0.89), inner retinal cellularity (r = 0.47), and RGC axon abundance (r = 0.72) all correlated significantly. Conclusions: Amongst outbred mice with widely variable phenotypes, OCT-derived measurements of GCC thickness significantly correlate to RGC abundance and axon number. The extensive phenotypic variability exhibited by J:DO mice make them a powerful resource for studies of retinal anatomy using quantitative genetics.

 bioRxiv Subject Collection: Neuroscience

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