Assessment of Rod, Cone, and Intrinsically Photosensitive Retinal Ganglion Cell Contributions to the Canine Chromatic Pupillary Response

Abstract

PurposeThe purpose of this study was to evaluate a chromatic pupillometry protocol for specific functional assessment of rods, cones, and intrinsically photosensitive retinal ganglion cells (ipRGCs) in dogs.MethodsChromatic pupillometry was tested and compared in 37 dogs in different stages of primary loss of rod, cone, and combined rod/cone and optic nerve function, and in 5 wild-type (WT) dogs. Eyes were stimulated with 1-s flashes of dim (1 cd/m2) and bright (400 cd/m2) blue light (for scotopic conditions) or bright red (400 cd/m2) light with 25-cd/m2 blue background (for photopic conditions). Canine retinal melanopsin/Opn4 was cloned, and its expression was evaluated using real-time quantitative reverse transcription-PCR and immunohistochemistry.ResultsMean ± SD percentage of pupil constriction amplitudes induced by scotopic dim blue (scDB), scotopic bright blue (scBB), and photopic bright red (phBR) lights in WT dogs were 21.3% ± 10.6%, 50.0% ± 17.5%, and 19.4% ± 7.4%, respectively. Melanopsin-mediated responses to scBB persisted for several minutes (7.7 ± 4.6 min) after stimulus offset. In dogs with inherited retinal degeneration, loss of rod function resulted in absent scDB responses, followed by decreased phBR responses with disease progression and loss of cone function. Primary loss of cone function abolished phBR responses but preserved those responses to blue light (scDB and scBB). Although melanopsin/Opn4 expression was diminished with retinal degeneration, melanopsin-expressing ipRGCs were identified for the first time in both WT and degenerated canine retinas.ConclusionsPupil responses elicited by light stimuli of different colors and intensities allowed differential functional assessment of canine rods, cones, and ipRGCs. Chromatic pupillometry offers an effective tool for diagnosing retinal and optic nerve diseases.

Protocol for assessment of the pupillary light reflex in dogs without chemical restraint: preliminary investigation

“…In doing so, we identified a measurable reduction in pupil response to white and red light stimuli in eyes with SARDS, and decreased pupillary response in eyes predisposed to primary angle-closure glaucoma in response to high-intensity blue light stimuli when compared other groups in this study. Pupil responses in SARDS cases were consistent with existing reports (Grozdanic et al 2007, Komáromy et al 2016, Young et al 2018 and other retinal diseases (Yeh et al 2017), providing a form of external validation for our protocol. We suggest quantifying pupil responses with technology that is affordable to veterinarians may facilitate better assessment and monitoring of neuroretinal function in veterinary practice.…”

“…With the addition of a total 30 minutes dark adaptation (in 10 minute intervals), this testing procedure does take a considerable amount of time, although actual testing is not prolonged and was well tolerated by the dogs. By attempting to develop a protocol practical in routine veterinary assessments, conditions in this study were not optimised for specific and isolated testing of individual photoreceptor layers and ipRGCs as they have been in previous reports describing optimal conditions for these assessments (Yeh et al 2017). For example, the low-intensity blue light in our protocol was the equivalent of 1.6 cd which is higher than that described (1 cd/m 2 ) (Yeh et al 2017) to specifically assess rod-mediated responses.…”

“…By attempting to develop a protocol practical in routine veterinary assessments, conditions in this study were not optimised for specific and isolated testing of individual photoreceptor layers and ipRGCs as they have been in previous reports describing optimal conditions for these assessments (Yeh et al 2017). For example, the low-intensity blue light in our protocol was the equivalent of 1.6 cd which is higher than that described (1 cd/m 2 ) (Yeh et al 2017) to specifically assess rod-mediated responses. Despite these limitations, our results show there is merit in further investigations using our modified protocol.…”

Sudden acquired retinal degeneration syndrome (SARDS) – a review and proposed strategies toward a better understanding of pathogenesis, early diagnosis, and therapy

“…and resulted in the recommendation that chromatic pupillary light reflexes (PLRs) may serve as a practical diagnostic tool to assess the canine visual pathways. In addition to electroretinography, loss of photoreceptor function can be verified clinically by observation of the lack of a PLR following stimulation with a bright (200 kcd/m 2 ) red light of 630 nm wavelength which primarily stimulates the cone photoreceptors . Because retinal ganglion cells (RGCs), including the melanopsin‐expressing intrinsically photosensitive RGCs (ipRGCs), are initially not affected by SARDS, PLRs are retained when eyes are stimulated with a bright (200 kcd/m 2 ) blue light of 480 nm wavelength that corresponds to the peak spectral sensitivity of melanopsin, whereas red light with a wavelength of 630 nm does not overlap with the melanopsin absorption spectrum .…”

Pupillary responses in non-proliferative diabetic retinopathy

“…As such, the response of the pupil to a flash of light can be complex, with potential contributions from rods, cones, and ipRGCs212223. However, by altering the adaptation conditions and stimulus characteristics, both inner- and outer-retinal contributions to the pupil response can be examined14152425. The pupil response driven by inner-retina neurons (melanopsin-mediated response) is characterized by a prolonged constriction following the offset of the stimulus, whereas the rod- and cone-mediated responses are characterized by rapid, transient constrictions15262728.…”