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Replacement of dysfunctional or dying photoreceptors offers a promising approach for retinal neurodegenerative diseases, including age-related macular degeneration and retinitis pigmentosa. Several studies have demonstrated the integration and differentiation of developing rod photoreceptors when transplanted in wild-type or degenerating retina; however, the physiology and function of the donor cells are not adequately defined. Here, we describe the physiological properties of developing rod photoreceptors that are tagged with green fluorescent protein (GFP) driven by the promoter of rod differentiation factor, Nrl. GFP-tagged developing rods show Ca2 + responses and rectifier outward currents that are smaller than those observed in fully developed photoreceptors, suggesting their immature developmental state. These immature rods also exhibit hyperpolarization-activated current ( Ih) induced by the activation of hyperpolarization-activated cyclic nucleotide-gated (HCN) channels. When transplanted into the subretinal space of wild-type or retinal degeneration mice, GFP-tagged developing rods can integrate into the photoreceptor outer nuclear layer in wild-type mouse retina and exhibit Ca2 + responses and membrane current comparable to native rod photoreceptors. A proportion of grafted rods develop rhodopsin-positive outer segment-like structures within 2 weeks after transplantation into the retina of Crx-knockout mice and produce rectifier outward current and Ih upon membrane depolarization and hyperpolarization. GFP-positive rods derived from induced pluripotent stem (iPS) cells also display similar membrane current Ih as native developing rod photoreceptors, express rod-specific phototransduction genes, and HCN-1 channels. We conclude that Nrl-promoter-driven GFP-tagged donor photoreceptors exhibit physiological characteristics of rods and that iPS cell-derived rods in vitro may provide a renewable source for cell-replacement therapy. S TEM C ells 2013;31:1149-1159 [ABSTRACT FROM AUTHOR]

Experimental autoimmune uveitis (EAU) is an inflammatory disease of the immune privileged inner eye, mediated by CD4+ Th1 cells specific for retinal autoantigens. To elucidate the fate of the T cells in the eye we adoptively transferred green fluorescent protein-positive (GFP+) T cells with specificity for R14, a peptide from interphotoreceptor retinoid-binding protein (IRBP) or OVA as foreign control antigen to naive Lewis rats. We also used the model of immunogenic uveitis, an inflammatory eye disease induced by intraocular injection of soluble OVA 1 day post transfer of OVA-specific GFP+ cells. We investigated the timing of ocular T cell infiltration and their immunological activation state by intravital fluorescence microscopy (IVFM) of the iris until onset of intraocular inflammation. Within 30 min of injection, GFP+ cells invaded the iris tissue, irrespective of their antigen specificity, whereas intraocular inflammation was only observed 3 days later, if cells recognized their respective antigen (R14-specific cells in EAU, OVA-specific cells in immunogenic uveitis). Using FACS analysis we found that activation markers were upregulated only on cells from uveitic eyes, but not from other sources, suggesting that intraocularly presented specific antigen is a prerequisite for T cell reactivation and subsequent recruitment of inflammatory cells. [ABSTRACT FROM AUTHOR]

Many tissues in non-mammalian vertebrates contain both photoreceptors and circadian clock systems. Among these photosensitive clock structures, the chick pineal gland has been characterized in detail from cellular and molecular aspects of the clock oscillation and entrainment. Analyses of the pineal photic-input pathway revealed a phase-shifting mechanism mediated by activation of G11, one of the Gq-type G-proteins. A major photoreceptive molecule, pinopsin, likely triggers this pathway by transmitting the light signal to the circadian oscillator. In the chick pineal oscillator, the transcription/translation-based autoregulatory feedback loop is composed of positive and negative elements (clock gene products) that are homologous to those identified in mammals. In the molecular cycling, a CACGTG E-box located in the promoter region of the negative element genes plays a central role in the transcriptional regulation. The phase of the molecular cycling is modulated by many regulatory components, among which E4BP4 and extracellular signal-regulated kinase (ERK) are closely associated with the photic entrainment. A light-responsive element was found in the promoter region of the Pinopsin gene, and the element included a CACGTG E-box, suggesting a novel role of the E-box as a point of convergence of light and circadian signals. These observations together point to general and unique features of the chick pineal circadian system among animal clocks. [ABSTRACT FROM PUBLISHER]

The circadian clock forms one of the most fascinating adaptations to life on earth. Organisms can not only anticipate the day/night cycle but can make use of an internal clock to measure daylength as an indicator of the changing of the seasons. The innate period of the clock is not exactly equal to 24 h, but is reset each day by environmental signals at dawn and dusk, most notably by changes in light and temperature. This ability to re‐entrain also ensures that the clock is synchronized with the day/night cycle which in turn is crucial for anticipation of dawn and dusk. Recent advances in the field have identified the photoreceptors involved in resetting the clock in several systems. This has revealed surprising similarities, but also key differences in the circadian systems of plants, fungi, insects, and mammals. One recurring feature emerging from this research is that the photoreceptors themselves are under the control of the clock with transcript abundance being tightly regulated. Furthermore, elements of a feedback pathway whereby the clock modulates the activity of the light input pathway are now being identified. [ABSTRACT FROM PUBLISHER]

Pinopsin is a chicken pineal photoreceptive molecule with a possible role in photoen-trainment of the circadian clock. Sequence comparison among members of the rhodop-sin family has suggested that pinopsin might have properties more similar to cone visual pigments than to rhodopsin, but the lifetime of the physiologically active intermediate (meta II) of pinopsin is rather similar to that of metarhodopsin II, which is far more stable than meta II intermediates of cone visual pigments [Nakamura, A. et al., (1999) Biochemistry 38, 14738–14745]. In the present study, we investigated the amino acid residue(s) contributing to this unique property of pinopsin by using site-directed mutagenesis to pinopsin-specific structural features, (i) Serl71, (ii) Asn184, and (iii) the second extracellular loop two-amino acids shorter than that of cone visual pigments. The meta II stability of the 171/184 double mutant of pinopsin (S171R/N184D) is almost the same as that of wild-type pinopsin. In contrast, the meta II lifetime is markedly shortened (one third) by introduction of the third mutation (replacement of a six-amino acid stretch, 188–193, by the corresponding eight residues of chicken green-sensitive cone pigment) to the 171/184 double mutant of pinopsin. Consistently, meta II of the green-sensitive pigment mutant, in which the eight-amino acid stretch is inversely replaced by the corresponding six residues of pinopsin, is more stable than meta II of the wild-type pigment. These results strongly suggest that the specific sequence and/or the number of residues at amino acids 188–193 in pinopsin play an important role in the stabilization of the meta II intermediate. [ABSTRACT FROM AUTHOR]

Glycoconjugates are likely to be of fundamental importance in the complex interactions between photoreceptors and the retinal pigment epithelium, but few have been characterized, especially in human tissue. As a preliminary step towards determining their biological functions in health and disease, a lectin-based histochemical study of the glycan expression of human outer retina was performed on glutaraldehyde-fixed, semi-thin, resin-embedded sections. The interphotoreceptor matrix and photoreceptor plasma-lemmata expressed complex bisected and non-bisected biantennary and/or triantennary -glycans. In addition, both the rod and cone outer segments bound strongly agglutinin (which binds terminal Manα1, 3Man-) and the rod outer segments bound selectively the isolectin II of (which binds terminal GlcNAc-). The cilia of the rods and cones were labelled selectively with agglutinin after sialidase pretreatment. Four putative glycan outer sequences were identified within the interphotoreceptor matrix: (i) sialylated glycans with subterminal GaINAcα1, 3GaINAc-sequences; (ii) a sialylated type with subterminal -acetyl-lactosamine residues; (iii) Galβ1, 3GalNAcα1- residues which were substituted with sialic acid except in the cone matrix sheath; (iv) GalNAcα1, 6Galβ1- residues which were substituted in part with sialic acid. The sialic acid expression throughout was predominantly of the 2, 3-linked form with lesser amounts of 2, 6-linkage, and rod-associated structures (including the surrounding interphotoreceptor matrix) were labelled more strongly with the sialic acid-binding lectins than cone-associated structures (including the cone matrix sheath). [ABSTRACT FROM PUBLISHER]

Looks at a study which proposed a molecular mechanism for the transport of retinoids between the retinal pigment epithelium and the rod outer segment, by the usage of interphotoreceptor retinoid-binding proteins. Function of the interphotoreceptor matrix; Discussion on the delivery of vitamin A by blood; Methodology used to conduct the study; Results of the study.