Monitoring Progressive Cone Cell Loss in Cpfl1 Mutant Mice in vivo

Abstract

Purpose:: To follow progressive cone cell loss in cpfl1 mice in vivo functionally by ERG (Ganzfeld electroretinography) and SLO imaging (scanning-laser ophthalmoscopy). For imaging,cpfl1 mutant mice expressing GFP allows to follow the time course of retinal degeneration in double mutants by SLO. We show the loss of cones in cpfl1 mutant mice by SLO and loss of cone-function by ERG analysis.

Methods:: Cpfl1 mice, bearing a natural mutation (116 bp insertion) in the gene encoding the cone cGMP-phosphodiesterase α subunit (PDE6C), were examined between 3 and 8 weeks of age. For imaging,cpfl1-GFP mice were generated by a crossbreed with a transgenic line expressing GFP under the control of RG cone opsin promotor, allowing to identify single photoreceptors in vivo by means of SLO autofluorescence (HRA I, Heidelberg Engineering). The results were compared to those of functional tests performed with the ERG (Multiliner Vision, Viasys), and to histological data. The study was performed in accordance with the ARVO Statement for the Use of Animals in Ophthalmic and Visual Research.

Results:: At 3 weeks of age, there was a dense distribution of GFP-expressing cones across the retina in both control mice expressing GFP without a cpfl1 mutation and cpfl1-GFP mice. Whereas in mice expressing GFP alone the amount of fluorescence and the number of visible photoreceptors remained approximately constant during the observation period, both of these indicators decreased with age in cpfl1-GFP mice. In these animals, a progressive reduction ofGFP expression was observed between 4 and 8 weeks. These data paralleled the reduction in the number of cells stainable with the cone-specific peanut agglutinin (PNA) in retinal whole mounts. Functionally, the cone-specific responses in the cpfl1-GFP mice were already severely reduced at 4 weeks of age and deteriorated further during the course of the study, whereas the responses from the rod system showed no comparable alterations, and also the ERG of the GFP expressing control mice remained completely unchanged.

Conclusions:: The cpfl1 mice undergo a progressive, selective degeneration of cone photoreceptors with age, which, when cones express GFP, can be very well monitored in vivo with SLO imaging. Interestingly, the functional loss precedes the morphological cell death by a considerable amount of time.

Keywords: imaging methods (CT, FA, ICG, MRI, OCT, RTA, SLO, ultrasound) • retina: distal (photoreceptors, horizontal cells, bipolar cells)