Abstract
Purpose :
Rodent models of Retinitis Pigmentosa (RP) typically manifest a phenotype during late retinal development, differing from what observed in the human pathology. Tvrm4 rhodopsin mice bear a photoinducible mutation that can be triggered in adulthood, better mimicking human RP. Here we exploit Tvrm4 mice to gain insight into the nature, time-course and severity of remodelling of bipolar cells, key neurons for retinal repair strategies.
Methods :
Tvrm4 mice aged 3-5 months and wild type (WT) littermates (n≥3/strain/group) were photo-induced with 12k LUX white-neon light for 2 mins. Eyes were explanted 3, 6 or 9 weeks post-induction (PI) and processed for immunohistochemistry or TEM for imaging bipolar cells and IPL connections. Quantitative image analysis was performed using Methamorph software, followed by statistical analysis in SigmaPlot.
Results :
Rod bipolar cells (RBCs) stained for anti-PKCα were counted in retinal whole mounts 3 and 6 weeks PI. No statistical difference was found in their number compared to WT controls (p=0,295). A significant shrinkage of axonal arborizations of these cells (p≥0,001) appeared 3 and 6 weeks PI. Anti-CtBP2/RIBEYE staining of ribbons in frozen, vertical sections 3, 6 and 9 weeks PI revealed a significant increase in the density of RIBEYE+ puncta in the IPL (p=0,010), 3 weeks PI, with respect to controls. A similar analysis of anti-Cx36 positive gap-junctions showed significant increase of puncta density in all experimental groups compared to WT controls (p≥0,001).
Cone bipolar cells were studied on vertical sections stained for anti-secretagogin (SCGN) 3 and 6 weeks PI. No statistical difference appeared either in the number of cell bodies, or in the area covered by axonal endings of SCGN+ cells. Yet, we found a gradual, significant decrease in the thickness of SCGN+ profiles in the IPL (p≥0,001). TEM studies confirmed shrinkage of individual RBC axonal endings but preservation of IPL synapses, although 50% of ribbons displayed an abnormally globular shape 6 weeks PI.
Conclusions :
High bipolar cell survival despite regressive dendritic and axonal remodelling found here coincide with previously described features of RP. Hence, we conclude that these events are independent of age of disease onset and underlying mutation. Increased density of ribbon synapses and gap-junctions may indicate retinal plasticity attempting to maintain the flow of remnant visual information and should be investigated further.
This is an abstract that was submitted for the 2018 ARVO Annual Meeting, held in Honolulu, Hawaii, April 29 - May 3, 2018.