Chromophore supply modulates cone function and survival in retinitis pigmentosa mouse models

**Background:** Retinitis pigmentosa (RP) is a common inherited retinal degenerative disease characterized by initial rod death followed by secondary cone loss, leading to blindness. The mechanisms underlying cone dysfunction and death after rod loss are unclear. This study investigates whether the supply of visual chromophore (11-cis retinal) contributes to cone toxicity in RP. The authors hypothesized that after rod death, cones are exposed to excessive chromophore, which may be toxic, and that slowing the visual cycle could protect cones.

**Methods:** The study used several RP mouse models: rd1 (Pde6b mutation), Rho-/- (rhodopsin knockout), and Gnat1-/- (transducin knockout, no rod function but rods present). Cone function was assessed by dark-adapted electroretinography (ERG) measuring the cone b-wave. To test the chromophore toxicity hypothesis, the authors crossed RP mice with mice deficient in Rlbp1 (encoding CRALBP, a retinoid carrier) or carrying the Rpe65 L450M mutation (slower visual cycle). They also used AAV vectors to ectopically express RPE65 and LRAT in cones of rd1 mice to increase chromophore supply. Cone survival was quantified by ARR3 (cone arrestin) staining. Additionally, Alms1-/- mice (Alström syndrome model) were studied to examine cone dark adaptation kinetics using ERG and ex vivo transretinal recordings.

**Key Results:** In rd1 mice, cone ERG was detectable at P15 (when rods are present) but nearly absent by P21 (when rods are lost). In Rho-/- mice, cone ERG was present at P60 (3-4 rows of rods) but lost by P90 (rods gone). In Gnat1-/- mice (rods present but non-functional), cone ERG was robust at both P15 and P21, indicating that loss of rod phototransduction alone does not cause cone ERG loss. Rlbp1 deficiency in rd1 mice preserved cone ERG at P20, P30, and P40, whereas controls had flat ERGs. Similarly, Rlbp1 deficiency in Rho-/- mice improved ERG at P90. Cone opsin expression (OPN1MW and OPN1SW) showed preserved outer segment structures in Rlbp1-/-;rd1 mice at P21, while controls had mislocalized opsins. Cone survival (ARR3+ cells) was significantly higher in the central retina of Rlbp1-/-;rd1 mice at P40 compared to controls (p < 0.0001). The Rpe65 L450M mutation also improved cone ERG at P20 (at 10 and 100 cd s/m²) and increased cone survival at P40 (p < 0.0001). Conversely, ectopic expression of RPE65 and LRAT in cones of rd1 mice did not affect ERG at P20 but significantly reduced cone survival at P50 (p < 0.0001). In Alms1-/-;Gnat1-/- mice, cone ERG b-wave amplitude was significantly reduced at most flash intensities compared to controls, and cone dark adaptation was accelerated (faster recovery of sensitivity after bleach) both in vivo and ex vivo, suggesting enhanced chromophore turnover.

**Clinical Implications:** This study provides strong evidence that excessive chromophore supply is toxic to cones in RP, and that slowing the visual cycle can preserve cone function and survival. These findings suggest that pharmacological or genetic interventions to reduce chromophore availability (e.g., by inhibiting RPE65 or RLBP1) could be therapeutic strategies to protect cones in RP patients. The results also offer a potential explanation for cone dysfunction in Alström syndrome, where accelerated chromophore turnover may contribute to cone degeneration. However, the study is limited to mouse models, and translation to humans requires further investigation. The authors note that the Rpe65 L450M mutation is naturally present in C57BL/6 mice and may influence results in studies using this background.