Quantitative in vivo Analysis of Cone Degeneration Secondary to a Rhodopsin Mutation

Abstract

Purpose: : To characterise in detail cone photoreceptor degeneration in the presence of a rod-specific mutation, and correlate in vivo findings made with the scanning laser ophthalmoscope with histological and functional assessments.

Methods: : A double transgenic mouse was generated which was homozygous for a rhodopsin knockout mutation and heterozygous for the eGFP gene under control of the human long-wavelength sensitive cone promoter. The time course of the cone degeneration was characterised in vivo through functional ERG assessments and repeated counts of cone cell numbers with a scanning laser ophthalmoscope; retinal flatmounts and cryosections were used to corroborate in vivo findings.

Results: : Individual cone photoreceptors can be imaged in vivo with the scanning laser ophthalmoscope and monitored over repeated assessments. In vivo quantification detected 67% (sd 12.3) of the GFP expressing cones seen histologically, and 52% (sd 1.9) of all cones in the posterior pole as detected with anti-cone transducin staining of retinal flat mounts. Cone photoreceptor loss in this murine model of retinitis pigmentosa (RP) follows an inferior to superior gradient, analogous to that in patients with selected rhodopsin mutations. In the rhodopsin knockout mouse cone photoreceptors survive in the posterior pole for up to 7 weeks after they have lost function, but have abnormal morphology.

Conclusions: : The use of the Scanning Laser Ophthalmoscope allows repeated in vivo assessments of cone photoreceptors in this murine model of RP. The secondary loss of cone photoreceptors follows an inferior to superior gradient and is delayed relative to the loss of photopic function, which has implications for treatments attempting to preserve vision through neuro-protective mechanisms.

Keywords: retinal degenerations: hereditary • imaging/image analysis: non-clinical • transgenics/knock-outs