Characterization of a Canine Model of Autosomal Recessive Retinitis Pigmentosa due to aPDE6AMutation

Abstract

Purpose To characterize a canine model of autosomal recessive RP due to a PDE6A gene mutation. Methods Affected and breed- and age-matched control puppies were studied by electroretinography (ERG), light and electron microscopy, immunohistochemistry and by assay for retinal PDE6 levels and enzymatic activity. Results The mutant puppies failed to develop normal rod-mediated ERG responses and had reduced light-adapted a-wave amplitudes from an early age. The residual ERG waveforms originated primarily from cone-driven responses. Development of photoreceptor outer segments was halted and rod cells were lost by apoptosis. Immunohistochemistry demonstrated a marked reduction in rod-opsin immunostaining outer segments and relative preservation of cones early in the disease process. With exception of rod bipolar cells that appeared to be reduced in number relatively early in the disease process other inner retinal cells were preserved in the early stages of the disease although there was marked and early activation of Müller glia. Western blotting showed that the PDE6A mutation not only resulted in a lack of PDE6A protein but the affected retinas also lacked the other PDE6 subunits, suggesting expression of PDE6A is required for normal expression of PDE6B and PDE6G. Affected retinas lacked PDE6 enzymatic activity. Conclusions This represents the first characterization of a PDE6A model of autosomal recessive retinitis pigmentosa and the PDE6A mutant dog shows promise as a large animal model for investigation of therapies to rescue mutant rod photoreceptors and to preserve cone photoreceptors in the face a rapid loss of rod cells.

Increased Plasma cGMP in a Family With Autosomal Recessive Retinitis Pigmentosa Due to Homozygous Mutations in the PDE6A Gene

“…One may argue that patients with a loss of measurable retinal functions as in this case should not have any photoreceptors left to produce cGMP. However, results from a PDE6A mutant canine model suggests that ERGs may be severely affected well in advance to structural loss of photoreceptors, 67 and the present clinical finding does not as such exclude the presence of rod photoreceptors, albeit in reduced numbers. Moreover, our results agree with those of Martinez-Fernandez de la Camara et al 15 where the serum cGMP levels of a group of RP patients are increased by approximately 65% compared with a control group.…”

Gene therapy in the second eye of RPE65-deficient dogs improves retinal function

“…ERGs were recorded under inhalant isoflurane anesthesia as previously described, except ERG-Jet corneal contact lens electrodes were used. 37 Briefly, globes were positioned in primary gaze using stay sutures of 4-0 silk (Ethicon, Inc., Piscataway, NJ, USA), and the pupils were dilated with 1% tropicamide (Mydriacyl, Alcon Laboratories, Honolulu, HI, USA) and 10% phenylephrine hydrochloride (AK-Dilate, Akorn Inc., Buffalo Grove, IL, USA). Full-field flash ERGs were recorded using ERGJet lenses (Microponent, Le Cret-du-Locie, Switzerland) and the UTAS-E 3000 electrophysiology unit with a Ganzfeld (LKC Technologies Inc.; Gaithersburg, MD, USA).…”

Electroretinography in dogs: a review

“…These differences concern individual features of ERG wavelets -implicit times and amplitudes. In the early stage of disease during which the rod system is first inactivated, such as rd, rcd and erd, roddriven responses are absent, while cone-driven responses may be intact or slightly reduced (Aguirre and Rubin 1971;Aguirre and Rubin 1975a;Aguirre 1978;Buyukmihci et al 1980;Petersen-Jones et al 2003;Tuntivanich et al 2009). Conversely, in cases when the cone system is inactivated first (for instance, in the course of crd1 in Dachshunds), cone-specific responses are significantly reduced, although the response of the rods remains within the normal range until 40 weeks of age (Curtis and Barnett 1993;Turney et al 2007;Busse et al 2011).…”