Phenotypic characterization of P23H and S334ter rhodopsin transgenic rat models of inherited retinal degeneration

Abstract

We produced 8 lines of transgenic (Tg) rats expressing one of two different rhodopsin mutations in albino Sprague-Dawley (SD) rats. Three lines were generated with a proline to histidine substitution at codon 23 (P23H), the most common autosomal dominant form of retinitis pigmentosa in the United States. Five lines were generated with a termination codon at position 334 (S334ter), resulting in a C-terminal truncated opsin protein lacking the last 15 amino acid residues and containing all of the phosphorylation sites involved in rhodopsin deactivation, as well as the terminal QVAPA residues important for rhodopsin deactivation and trafficking. The rates of photoreceptor (PR) degeneration in these models vary in proportion to the ratio of mutant to wild-type rhodopsin. The models have been widely studied, but many aspects of their phenotypes have not been described. Here we present a comprehensive study of the 8 Tg lines, including the time course of PR degeneration from the onset to one year of age, retinal structure by light and electron microscopy (EM), hemispheric asymmetry and gradients of rod and cone degeneration, rhodopsin content, gene dosage effect, rapid activation and invasion of the outer retina by presumptive microglia, rod outer segment disc shedding and phagocytosis by the retinal pigmented epithelium (RPE), and retinal function by the electroretinogram (ERG). The biphasic nature of PR cell death was noted, as was the lack of an injury-induced protective response in the rat models. EM analysis revealed the accumulation of submicron vesicular structures in the interphotoreceptor space during the peak period of PR outer segment degeneration in the S334ter lines. This is likely due to the elimination of the trafficking consensus domain as seen before as with other rhodopsin mutants lacking the C-terminal QVAPA. The 8 rhodopsin Tg lines have been, and will continue to be, extremely useful models for the experimental study of inherited retinal degenerations.

PRCD is essential for high-fidelity photoreceptor disc formation

“…Although not ubiquitously observed across animal models of inherited photoreceptor degeneration, Prcd −/− and rds mice are not the only animals where extracellular vesicles are observed between photoreceptors or in the subretinal space. Other models include Tulp1 −/− mice (33), tubby mice (34), BBS8 −/− mice (35), Tg737/IFT88 mice (36), rhodopsin P347S transgenic mice (37), rhodopsin Q344ter transgenic mice (38), pcd mice (30,39), rhodopsin P347L transgenic rabbits (40), and rhodopsin S334ter transgenic rats (41). However, unlike in Prcd −/− and rds mice, the location of these vesicles typically extends to the space between photoreceptor inner segments.…”

“…As conceptualized by Lavail et al in ref. 41: "In all cases, the origin of the vesicles appears to be from budding or blebbing from the [inner segments], or in some cases, protrusions from the [inner segments]." Therefore, the phenotypes observed in PRCD and peripherin knockouts may be rather unique and potentially foretelling for addressing the fine mechanisms of photoreceptor disc morphogenesis.…”

Detailed Visual Cortical Responses Generated by Retinal Sheet Transplants in Rats with Severe Retinal Degeneration

“…Spikes were sorted online in Trellis while performing visual stimulation. Visual stimuli were generated in MATLAB (The MathWorks) using Psychophysics Toolbox (Brainard, 1997;Pelli, 1997;Kleiner et al, 2007) and displayed on a gamma-corrected LCD monitor (55 inches, 60 Hz) at resolution 1920 ϫ 1080 pixels and 52 cd/m 2 mean luminance. Stimulus onset times were corrected for LCD monitor delay using a photoresistor and microcontroller (in-house design).…”

“…It is not until after LGN afferents converge onto neurons in primary visual cortex (V1) that key features, such as orientation selectivity, fully emerge, although in mouse LGN direction selectivity is already present (Hubel and Wiesel, 1962;Reid and Alonso, 1996;Niell and Stryker, 2008;Marshel et al, 2012). Many additional stimulus features, such as temporal and spatial frequency, contrast, size, and direction, also elicit highly selective responses in V1 and are considered key building blocks for the perception of complex shapes and motion (Livingstone and Hubel, 1988;Kobatake and Tanaka, 1994;Marshel et al, 2011;Glickfeld et al, 2014).…”

Coupling of Human Rhodopsin to a Yeast Signaling Pathway Enables Characterization of Mutations Associated with Retinal Disease

“…P23H is the most common RP-associated rhodopsin mutation in North America (Dryja et al 1990;Mendes et al 2005), and has been characterized in a number of cell and animal models. P23H rhodopsin consistently displays poor stability (Krebs et al 2010;Chen et al 2014), aggregation in the ER (Chiang et al 2012b), and disrupted transducin activation (Opefi et al 2013;Chen et al 2014), leading to severe retinal degeneration (Cideciyan et al 1998;Athanasiou et al 2017;LaVail et al 2018). The less severe M39R mutation, which is also associated with RP, has been studied using both bovine and human rhodopsin genes displaying a more severe cytosolic aggregation phenotype in the human gene background (Davies et al 2012;Ramon et al 2014).…”