Function of mammalian M-cones depends on the level of CRALBP in Müller cells

Cone photoreceptors mediate daytime vision in vertebrates. The rapid and efficient regeneration of their visual pigments following photoactivation is critical for the cones to remain photoresponsive in bright and rapidly changing light conditions. Cone pigment regeneration depends on the recycling of visual chromophore, which takes place via the canonical visual cycle in the retinal pigment epithelium (RPE) and the Müller cell-driven intraretinal visual cycle. The molecular mechanisms that enable the neural retina to regenerate visual chromophore for cones have not been fully elucidated. However, one known component of the two visual cycles is the cellular retinaldehyde-binding protein (CRALBP), which is expressed both in the RPE and in Müller cells. To understand the significance of CRALBP in cone pigment regeneration, we examined the function of cones in mice heterozygous for Rlbp1, the gene encoding CRALBP. We found that CRALBP expression was reduced by ∼50% in both the RPE and retina of Rlbp1+/- mice. Electroretinography (ERG) showed that the dark adaptation of rods and cones is unaltered in Rlbp1+/- mice, indicating a normal RPE visual cycle. However, pharmacologic blockade of the RPE visual cycle revealed suppressed cone dark adaptation in Rlbp1+/- mice in comparison with controls. We conclude that the expression level of CRALPB specifically in the Müller cells modulates the efficiency of the retina visual cycle. Finally, blocking the RPE visual cycle also suppressed further cone dark adaptation in Rlbp1-/- mice, revealing a shunt in the classical RPE visual cycle that bypasses CRALBP and allows partial but unexpectedly rapid cone dark adaptation

Retinoid isomerase inhibitors impair but do not block mammalian cone photoreceptor function

Visual function in vertebrates critically depends on the continuous regeneration of visual pigments in rod and cone photoreceptors. RPE65 is a well-established retinoid isomerase in the pigment epithelium that regenerates rhodopsin during the rod visual cycle; however, its contribution to the regeneration of cone pigments remains obscure. In this study, we use potent and selective RPE65 inhibitors in rod- and cone-dominant animal models to discern the role of this enzyme in cone-mediated vision. We confirm that retinylamine and emixustat-family compounds selectively inhibit RPE65 over DES1, the putative retinoid isomerase of the intraretinal visual cycle. In vivo and ex vivo electroretinography experiments in Gnat1-/- mice demonstrate that acute administration of RPE65 inhibitors after a bleach suppresses the late, slow phase of cone dark adaptation without affecting the initial rapid portion, which reflects intraretinal visual cycle function. Acute administration of these compounds does not affect the light sensitivity of cone photoreceptors in mice during extended exposure to background light, but does slow all phases of subsequent dark recovery. We also show that cone function is only partially suppressed in cone-dominant ground squirrels and wild-type mice by multiday administration of an RPE65 inhibitor despite profound blockade of RPE65 activity. Complementary experiments in these animal models using the DES1 inhibitor fenretinide show more modest effects on cone recovery. Collectively, these studies demonstrate a role for continuous RPE65 activity in mammalian cone pigment regeneration and provide further evidence for RPE65-independent regeneration mechanisms

Conditional deletion of Des1 in the mouse retina does not impair the visual cycle in cones

Cone photoreceptors are essential for vision under moderate to high illuminance and allow color discrimination. Their fast dark adaptation rate and resistance to saturation are believed to depend in part on an intraretinal visual cycle that supplies 11- cis-retinaldehyde to cone opsins. Candidate enzymes of this pathway have been reported, but their physiologic contribution to cone photoresponses remains unknown. Here, we evaluate the role of a candidate retinol isomerase of this pathway, sphingolipid δ4 desaturase 1 (Des1). Single-cell RNA sequencing analysis revealed Des1 expression not only in Müller glia but also throughout the retina and in the retinal pigment epithelium. We assessed cone functional dependence on Müller cell-expressed Des1 through a conditional knockout approach. Floxed Des1 mice, on a guanine nucleotide-binding protein subunit α transducin 1 knockout ( Gnat

A combination treatment based on drug repurposing demonstrates mutation-agnostic efficacy in pre-clinical retinopathy models

Inherited retinopathies are devastating diseases that in most cases lack treatment options. Disease-modifying therapies that mitigate pathophysiology regardless of the underlying genetic lesion are desirable due to the diversity of mutations found in such diseases. We tested a systems pharmacology-based strategy that suppresses intracellular cAMP and Ca2+ activity via G protein-coupled receptor (GPCR) modulation using tamsulosin, metoprolol, and bromocriptine coadministration. The treatment improves cone photoreceptor function and slows degeneration in Pde6βrd10 and RhoP23H/WT retinitis pigmentosa mice. Cone degeneration is modestly mitigated after a 7-month-long drug infusion in PDE6A-/- dogs. The treatment also improves rod pathway function in an Rpe65-/- mouse model of Leber congenital amaurosis but does not protect from cone degeneration. RNA-sequencing analyses indicate improved metabolic function in drug-treated Rpe65-/- and rd10 mice. Our data show that catecholaminergic GPCR drug combinations that modify second messenger levels via multiple receptor actions provide a potential disease-modifying therapy against retinal degeneration

Different paths to the modern state in Europe: the interaction between domestic political economy and interstate competition

Theoretical work on state formation and capacity has focused mostly on early modern Europe and on the experience of western European states during this period. While a number of European states monopolized domestic tax collection and achieved gains in state capacity during the early modern era, for others revenues stagnated or even declined, and these variations motivated alternative hypotheses for determinants of fiscal and state capacity. In this study we test the basic hypotheses in the existing literature making use of the large date set we have compiled for all of the leading states across the continent. We find strong empirical support for two prevailing threads in the literature, arguing respectively that interstate wars and changes in economic structure towards an urbanized economy had positive fiscal impact. Regarding the main point of contention in the theoretical literature, whether it was representative or authoritarian political regimes that facilitated the gains in fiscal capacity, we do not find conclusive evidence that one performed better than the other. Instead, the empirical evidence we have gathered lends supports to the hypothesis that when under pressure of war, the fiscal performance of representative regimes was better in the more urbanized-commercial economies and the fiscal performance of authoritarian regimes was better in rural-agrarian economie

Retinal pigment epithelium 65 kDa protein (RPE65): An update

Vertebrate vision critically depends on an 11-cis-retinoid renewal system known as the visual cycle. At the heart of this metabolic pathway is an enzyme known as retinal pigment epithelium 65 kDa protein (RPE65), which catalyzes an unusual, possibly biochemically unique, reaction consisting of a coupled all-trans-retinyl ester hydrolysis and alkene geometric isomerization to produce 11-cis-retinol. Early work on this isomerohydrolase demonstrated its membership to the carotenoid cleavage dioxygenase superfamily and its essentiality for 11-cis-retinal production in the vertebrate retina. Three independent studies published in 2005 established RPE65 as the actual isomerohydrolase instead of a retinoid-binding protein as previously believed. Since the last devoted review of RPE65 enzymology appeared in this journal, major advances have been made in a number of areas including our understanding of the mechanistic details of RPE65 isomerohydrolase activity, its phylogenetic origins, the relationship of its membrane binding affinity to its catalytic activity, its role in visual chromophore production for rods and cones, its modulation by macromolecules and small molecules, and the involvement of RPE65 mutations in the development of retinal diseases. In this article, I will review these areas of progress with the goal of integrating results from the varied experimental approaches to provide a comprehensive picture of RPE65 biochemistry. Key outstanding questions that may prove to be fruitful future research pursuits will also be highlighted

Pathways and disease-causing alterations in visual chromophore production for vertebrate vision.

All that we view of the world begins with an ultrafast cis to trans photoisomerization of the retinylidene chromophore associated with the visual pigments of rod and cone photoreceptors. The continual responsiveness of these photoreceptors is then sustained by regeneration processes that convert the trans-retinoid back to an 11-cis configuration. Recent biochemical and electrophysiological analyses of the retinal G-protein-coupled receptor (RGR) suggest that it could sustain the responsiveness of photoreceptor cells, particularly cones, even under bright light conditions. Thus, two mechanisms have evolved to accomplish the reisomerization: one involving the well-studied retinoid isomerase (RPE65) and a second photoisomerase reaction mediated by the RGR. Impairments to the pathways that transform all-trans-retinal back to 11-cis-retinal are associated with mild to severe forms of retinal dystrophy. Moreover, with age there also is a decline in the rate of chromophore regeneration. Both pharmacological and genetic approaches are being used to bypass visual cycle defects and consequently mitigate blinding diseases. Rapid progress in the use of genome editing also is paving the way for the treatment of disparate retinal diseases. In this review, we provide an update on visual cycle biochemistry and then discuss visual-cycle-related diseases and emerging therapeutics for these disorders. There is hope that these advances will be helpful in treating more complex diseases of the eye, including age-related macular degeneration (AMD)