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PubMed2005

Grating acuity at different luminances in wild-type mice and in mice lacking rod or cone function.

PURPOSE: The mouse eye has become an important model in vision research. However, it is not known how visual acuity changes with luminance. Therefore, grating acuity of mice was measured at different luminances in an automated optomotor paradigm. Furthermore, mutant mice lacking either rods (RHO-/- and CNGB1-/-) or cones (CNGA3-/-), or both, were studied to determine the rod and cone contribution to visual acuity. METHODS: Freely ranging individual mice were automatically tracked at a 25-Hz sampling rate with a self-programmed video system in a large rotating optomotor drum. The drum had a square-wave grating inside with adjustable spatial frequency. The angular speed of the mice with respect to the center of the drum and the angular orientation of the snout-tail body axis were analyzed. In addition, the motor activity of the wild-type mice was recorded at different luminances. RESULTS: The optomotor drum provided reliable data on visual input to the mouse's behavior and was convenient to use, since the experimenter's had only to place the mice individually in a Perspex cylinder. Optomotor grating acuity of the wild-type mice was limited to 0.3 to 0.4 cyc/deg. Maximum optomotor responses were obtained at 0.1 to 0.2 cyc/deg. The importance of visual input declined monotonically with decreasing luminance (30 cd/m2, 100%; 0.1 cd/m2, 76.4%; 0.005 cd/m2, 45.9%; and darkness, -9%). Mice lacking functional rods were able to resolve gratings up to 0.1 cyc/deg at 30 cd/m2. Surprisingly, mice lacking functional cones had an optomotor acuity that was similar to the wild-type. Double-knockout mice without rods and cones had no detectable grating acuity. CONCLUSIONS: Because the visual system of the mouse is more responsive at bright luminances, experiments in which visual input is important should be performed in photopic conditions (30 cd/m2 or even more). Apparently, spatial vision is governed by the rod system, which is not saturated in the mesopic or low photopic range. Mice lacking both rods and cones have no detectable grating acuity, indicating that the retinal melanopsin system does not contribute to spatial vision.

Animals↗
PubMed2005

Leukemia inhibitory factor blocks expression of Crx and Nrl transcription factors to inhibit photoreceptor differentiation.

PURPOSE: Activating ligands of gp130, including leukemia inhibitory factor (LIF), can block differentiation and function of retinal neurons. This study focused on determining whether LIF inhibits differentiation of photoreceptors by altering cell fate or by blocking the expression of essential transcription factors in vivo. METHODS: Transgenic mice were generated that had lens-specific expression of the secreted human LIF protein. Retinal differentiation was assessed by histology and by gene expression analysis, with in situ hybridization, immunohistochemistry, and real-time qRT-PCR. Electroretinograms were used to assess retinal function. RESULTS: LIF did not prevent or alter the timing of outer and inner nuclear layer separation, but it inhibited phototransduction gene expression in both rods and cones, thereby blocking functional maturation of photoreceptors. LIF also reduced the expression of Crx, Nrl, and Nr2e3, and upregulated the expression of transcription inhibitors Baf and Fiz1. CONCLUSIONS: LIF expression did not appear to alter photoreceptor cell fate specification, but it inhibited subsequent differentiation. These results suggest that gp130 ligands can inhibit photoreceptor functional differentiation by reducing Crx- and Nrl-dependent transcription.

Animals↗
PubMed2006

Quantitative analysis of OCT characteristics in patients with achromatopsia and blue-cone monochromatism.

PURPOSE: To quantify optical coherence tomography (OCT) images of the central retina in patients with blue-cone monochromatism (BCM) and achromatopsia (ACH) compared with healthy control individuals. METHODS: The study included 15 patients with ACH, 6 with BCM, and 20 control subjects. Diagnosis of BCM and ACH was established by visual acuity testing, morphologic examination, color vision testing, and Ganzfeld ERG recording. OCT images were acquired with the Stratus OCT 3 (Carl Zeiss Meditec AG, Oberkochen, Germany). Foveal OCT images were analyzed by calculating longitudinal reflectivity profiles (LRPs) from scan lines. Profiles were analyzed quantitatively to determine foveal thickness and distances between reflectivity layers. RESULTS: Patients with ACH and BCM had a mean visual acuity of 20/200 and 20/60, respectively. Color vision testing results were characteristic of the diseases. The LRPs of control subjects yielded four peaks (P1-P4), presumably representing the RPE (P1), the ovoid region of the photoreceptors (P2), the external limiting membrane (ELM) (P3), and the internal limiting membrane (P4). In patients with ACH, P2 was absent, but foveal thickness (P1-P4) did not differ significantly from that in the control subjects (187 +/- 20 vs. 192 +/- 14 microm, respectively). The distance from P1 to P3 did not differ significantly (78 +/- 10 vs. 82 +/- 5 microm) between ACH and controls subjects. In patients with BCM, P3 was lacking, and P2 advanced toward P1 compared with the control subjects (32 +/- 6 vs. 48 +/- 4 microm). Foveal thickness (153 +/- 16 microm) was significantly reduced compared with that in control subjects and patients with ACH. CONCLUSIONS: Quantitative OCT image analysis reveals distinct patterns for controls subjects and patients with ACH and BCM, respectively. Quantitative analysis of OCT imaging can be useful in differentiating retinal diseases affecting photoreceptors. Foveal thickness is similar in both normal subjects and patients with ACH but is decreased in patients with BCM.

Adult↗
PubMed2006

Severity staging by early features of age-related maculopathy exhibits weak relationships with functional deficits on SWS grating acuity.

PURPOSE: To examine the relationship between short-wavelength-sensitive (SWS) resolution acuity and epidemiologically defined stages of early age-related maculopathy (ARM). METHODS: Subjects consisted of 88 adults aged 51 to 87 years. Psychophysical testing was undertaken in only one eye of each subject (the study eye). All study eyes had a LogMAR acuity of 0.30 (20/40 Snellen) or better. SWS and achromatic grating resolution acuity were measured at 6 degrees eccentricity from the fovea. Stereoscopic color fundus photographs centered on the macula were taken on both eyes of each subject and were graded using the Wisconsin Age-Related Maculopathy Grading System (WARMGS). After grading, features of ARM were combined to assign a severity stage from 0 to 5 using the methods described by the Rotterdam Eye Study. Relationships between visual function, study eye ARM stage, and fellow eye status were examined with the use of standard statistical analysis. RESULTS: Although SWS resolution acuity was significantly reduced in eyes classified as having any ARM compared with eyes classified as having no ARM (P = 0.002), there was no relationship between the severity of functional deficits and the morphologic severity from stage 1 to stage 4. On reassigning subject eyes to a revised severity staging (stage 0, stages 1 to 4 combined, and stage 5), SWS acuity was significantly different among these three groups (P < 0.001). No significant relationship was found between achromatic resolution acuity and ARM staging. The status of the fellow eye (advanced macular degeneration present or absent) was not significantly related to visual function in the study eye. CONCLUSIONS: Significant functional deficits in SWS resolution acuity were found in eyes with ARM features, but the severity of functional loss did not correlate well with the currently accepted method of assigning a morphologic severity stage. Longitudinal studies may reveal further information on the relationships between functional deficits, ARM status, disease progression, and outcome.

Aged↗
PubMed2006

Melanopsin-expressing retinal ganglion cells are more injury-resistant in a chronic ocular hypertension model.

PURPOSE: To investigate the survival of melanopsin-expressing retinal ganglion cells (mRGCs) after the induction of chronic ocular hypertension. METHODS: Intraocular pressure (IOP) was elevated in adult Sprague-Dawley rats using an argon laser to photocoagulate the episcleral and limbal veins. IOP was measured with a calibrated tonometer and monitored for a period. Seven days before the animals were killed, a piece of sterile foam soaked with gold fluorescent dye was placed onto the superior colliculus (SC) to label the SC-projecting retinal ganglion cells (scRGCs) retrogradely. mRGCs were visualized by free floating immunohistochemistry on whole-mounted retinas. The number of surviving scRGCs and mRGCs were counted on flatmounted retinas. The branching pattern of dendrites and soma size of mRGCs were examined. RESULTS: An approximately 1.7-fold increase of IOP and a significant loss of scRGCs were found in experimental eyes after laser photocoagulation. However, no significant cell loss or morphologic changes on mRGCs and their dendrites after the induction of chronic ocular hypertension are noticed over a 12-week period. CONCLUSIONS: Although the degeneration of retinal ganglion cells (RGCs) is a major concern in glaucomatous damage, the findings show that mRGCs are less susceptible to death after the induction of chronic ocular hypertension. This result indicates that mRGCs carry some unique properties that are different from those of other subpopulations of RGCs. The immunohistochemistry approach can be used to distinguish easily these mRGCs from other subtypes. This method provides a useful tool to investigate their injury-resistant properties that are informative for the development of effective neuroprotective treatment for glaucoma.

Animals↗
PubMed2003

Melanopsin, ganglion-cell photoreceptors, and mammalian photoentrainment.

An understanding of the retinal mechanisms in mammalian photoentrainment will greatly facilitate optimization of the wavelength, intensity, and duration of phototherapeutic treatments designed to phase shift endogenous biological rhythms. A small population of widely dispersed retinal ganglion cells projecting to the suprachiasmatic nucleus in the hypothalamus is the source of the critical photic input. Recent evidence has shown that many of these ganglion cells are directly photosensitive and serve as photoreceptors. Melanopsin, a presumptive photopigment, is an essential component in the phototransduction cascade within these intrinsically photosensitive ganglion cells and plays an important role in the retinal photoentrainment pathway. This review summarizes recent findings related to melanopsin and melanopsin ganglion cells and lists other retinal proteins that might serve as photopigments in the mammalian photoentrainment input pathway.

Animals↗
PubMedDate not supplied

Optical coherence tomography in enhanced S-cone syndrome: large macular retinoschisis with disorganized retinal lamination.

PURPOSE: The authors previously reported clinical and molecular features of a boy (at age 17) with enhanced S-cone syndrome (ESCS) who had bilateral cystoid maculopathies. The purpose of the current study was to describe the patient's optical coherence tomography (OCT) findings. METHODS: OCT was performed when the patient was 23 years old. RESULTS: OCT images revealed formations of single large hyporeflective cystic spaces in the cystoid maculopathies of both eyes. The cystic spaces were much larger than those of previously reported cases. In the temporal region of each cystic space, symmetric disorganized retinal lamination was observed with the retina lacking well-defined and hyporeflective bands of the inner nuclear layer and the outer nuclear layer, which are seen in normal retina. Splitting in the retinal thickness was at the level of the outer retinal layer rather than at the inner retinal layer in both eyes. CONCLUSIONS: Splitting is likely to occur close to the outer plexiform layer in which the cleavage plane of familial juvenile retinoschisis is identified. The unique OCT manifestation of symmetric large macular retinoschisis with disorganized retinal lamination may indicate a severe form of ESCS.

Adult↗
PubMed1984

Preembedding labeling with biotinylated antibodies and subsequent visualization of the biotin groups exposed on thin sections.

The feasibility of labeling cell membranes with biotinylated ligands and detecting the biotin groups on thin sections was investigated. Fixed retinal tissue was incubated with biotinyl- antiopsin . Half of the biotinyl-antibody labeled retinal tissue was incubated with avidin-ferritin (AvF) and embedded in Epon (preembedding reaction). The second half was embedded in glutaraldehyde cross-linked bovine serum albumin (BSA). Thin sections of this preparation were incubated with AvF to detect biotinyl-antibodies exposed by the sectioning (postembedding reaction). Biotin groups on the thin section surface could be readily visualized with AvF. Stereoscopic images demonstrated that the ferritin particles were localized only on the exposed surface of the thin section. The labeling was highly specific, with a very low background. Quantitative analysis was employed in order to determine the optimal reaction conditions for maximizing the labeling density with minimizing nonspecific binding. The possibility of using biotinylated molecules in the study of dynamic cellular events and for the subsequent intracellular localization of biotin on thin sections is suggested.

Animals↗
PubMed1996

Retinal dysplasia and degeneration in RARbeta2/RARgamma2 compound mutant mice.

The eye is the organ whose development is the most frequently altered in response to maternal vitamin A deficiency [VAD; Warkany, J. and Schraffenberger, S. (1946). Archs Ophthalmol. 35, 150-169]. With the exception of prenatal retinal dysplasia, all the ocular abnormalities of the fetal VAD syndrome are recapitulated in mouse mutants lacking either RARalpha and RARbeta2, RARalpha and RARgamma, RARgamma and RARbeta2, or RXRalpha [Lohnes, D., Mark, M., Mendelsohn, C., Dolle, P., Dierich, A., Gorry, P., Gansmuller, A. and Chambon, P. (1994) Development 120, 2723-2748; Mendelsohn, C., Lohnes, D., Décimo, D., Lufkin, T., LeMeur, M., Chambon, P. and Mark, M. (1994) Development 120, 2749-2771; Kastner, P., Grondona, J. Mark, M., Gansmuller, A., LeMeur, M., Décimo, D., Vonesch, J.L., Dollé, P. and Chambon, P. (1994) Cell 78, 987-1003], thus demonstrating that retinoic acid (RA) is the active vitamin A metabolite during prenatal eye morphogenesis. Whether retinoids are also involved in postnatal eye development could not be investigated, as VAD newborns are not viable and the above RAR double null mutants and RXRalpha null mutants died in utero or at birth. We report here the generation of viable RARbeta2/RARgamma2 double null mutant mice, which exhibit several eye defects. The neural retina of newborn RARbeta2gamma2 mutants is thinner than normal due to a reduced rate of cell proliferation, and from day 4 shows multiple foci of disorganization of its layers. These RARbeta2gamma2 mutants represent the first genetically characterized model of retinal dysplasia and their phenotype demonstrates that RARs, and therefore RA, are required for retinal histogenesis. The RARbeta2gamma2 retinal pigment epithelium (RPE) cells display histological and/or ultrastructural alterations and/or fail to express cellular retinol binding protein I (CRBPI). Taken altogether, the early onset of the RPE histological defects and their striking colocalisation with areas of the neural retina displaying a faulty laminar organization, a reduced neuroblastic proliferation, and a lack of photoreceptor differentiation and/or increased apoptosis, make the RPE a likely target tissue of the RARbeta2gamma2 double null mutation. A degeneration of the adult neural retina, which may similarly be secondary to a defective RPE, is also observed in these mutants, thus demonstrating an essential role of RA in the survival of retinal cells. Moreover, all RARbeta2gamma2 mutants display defects in structures derived from the periocular mesenchyme including local agenesis of the choroid and of the sclera, small eyelids, and a persistence of the primary mesenchymal vitreous body. A majority of the RARbeta2 single null mutants also exhibit this latter defect, thus demonstrating that the RARbeta2 isoform plays a unique role in the formation of the definitive vitreous body.

Animals↗
PubMed2002

Three vitamins are involved in regulation of the circadian rhythm.

Photoreceptors in the eyes of mammals synchronize the innate circadian rhythm to the solar light-dark cycle. They differ from the photoreceptors serving vision in rods and cones of the retina and are located in the ganglion cells of the retina. They consist of melanopsin, a protein homologous to rhodopsin, with retinal as the light-sensitive prosthetic group; and cryptochromes 1 and 2, proteins combined with methenyltetrahydrofolate and flavin adenine dinucleotide, which function as blue light-sensitive photoreceptors.

Animals↗
PubMed2006

Evolution of melanopsin photoreceptors: discovery and characterization of a new melanopsin in nonmammalian vertebrates.

In mammals, the melanopsin gene (Opn4) encodes a sensory photopigment that underpins newly discovered inner retinal photoreceptors. Since its first discovery in Xenopus laevis and subsequent description in humans and mice, melanopsin genes have been described in all vertebrate classes. Until now, all of these sequences have been considered representatives of a single orthologous gene (albeit with duplications in the teleost fish). Here, we describe the discovery and functional characterisation of a new melanopsin gene in fish, bird, and amphibian genomes, demonstrating that, in fact, the vertebrates have evolved two quite separate melanopsins. On the basis of sequence similarity, chromosomal localisation, and phylogeny, we identify our new melanopsins as the true orthologs of the melanopsin gene previously described in mammals and term this grouping Opn4m. By contrast, the previously published melanopsin genes in nonmammalian vertebrates represent a separate branch of the melanopsin family which we term Opn4x. RT-PCR analysis in chicken, zebrafish, and Xenopus identifies expression of both Opn4m and Opn4x genes in tissues known to be photosensitive (eye, brain, and skin). In the day-14 chicken eye, Opn4m mRNA is found in a subset of cells in the outer nuclear, inner nuclear, and ganglion cell layers, the vast majority of which also express Opn4x. Importantly, we show that a representative of the new melanopsins (chicken Opn4m) encodes a photosensory pigment capable of activating G protein signalling cascades in a light- and retinaldehyde-dependent manner under heterologous expression in Neuro-2a cells. A comprehensive in silico analysis of vertebrate genomes indicates that while most vertebrate species have both Opn4m and Opn4x genes, the latter is absent from eutherian and, possibly, marsupial mammals, lost in the course of their evolution as a result of chromosomal reorganisation. Thus, our findings show for the first time that nonmammalian vertebrates retain two quite separate melanopsin genes, while mammals have just one. These data raise important questions regarding the functional differences between Opn4x and Opn4m pigments, the associated adaptive advantages for most vertebrate species in retaining both melanopsins, and the implications for mammalian biology of lacking Opn4x.

Amino Acid Sequence↗
PubMed2003

Melanopsin in the circadian timing system.

In mammals, circadian rhythms are generated by a light-entrainable oscillator located in the hypothalamic suprachiasmatic nucleus (SCN). Light signals reach the SCN via a dedicated retinal pathway, the retinohypothalamic tract (RHT). One question that continues to elude scientists is whether the circadian system has its own dedicated photoreceptor or photoreceptors. It is well established that conventional photoreceptors, rods and cones, are not required for circadian photoreception, suggesting that the inner retinal layer might contribute to circadian photoreception. Melanopsin, a novel photo pigment expressed in retinal ganglion cells (RGCs), has been proposed recently as a candidate circadian photoreceptor. Melanopsin-containing RGCs are intrinsically photosensitive, form part of the RHT, and contain neurotransmitters known to play a critical role in the circadian response to light. Furthermore, melanopsin-containing RGCs do not depend on inputs from rods and cones to transmit light signals to the SCN. However, based on a review of the available information about melanopsin and on new data from our laboratory, we propose that melanopsin, in itself, is not necessary for circadian photoreception. In fact, it appears that of the known photoreceptor systems, none, in and of itself, is necessary for circadian photoreception. Instead, it appears that within the photoreceptive systems there is some degree of redundancy, each contributing in some way to photic entrainment.

Animals↗
PubMed2005

Does the chromophore's ring move after photoexcitation of rhodopsin?

By comparing the shift of the absorption maxima when a visual pigment is converted to its lumirhodopsin photointermediate for two classes of pigments, we can infer whether or not the pigment's beta-ionone ring has left its binding site. We compare this shift for the long-wavelength sensitive visual pigment of chicken iodopsin (lambdamax = 571 nm), which has polar residues in the ring binding site that interact with the ring, with that for three pigments, which do not. We conclude that by the time the Lumi product of the pigment is formed, the ring has moved away from the ring binding site.

Animals↗
PubMed1998

Extraretinal photoreceptors and their regulation of temporal physiology.

The extraretinal photoreceptors of non-mammalian vertebrates play an important role in the regulation of temporal physiology. Both the regulation of circadian clocks and the photoperiodic response of many animals depend upon the photic information provided by these receptors. Since their discovery at the beginning of this century, and despite their importance, extraretinal photoreceptors have remained poorly understood. Until recently, their cellular location within the central nervous system, and the nature of the photopigments they use, remained a mystery. Antibodies directed against rod or cone photopigment proteins have been used in immunocytochemical procedures to localize extraretinal photoreceptors. However, findings have been confusing. The use of molecular approaches has led to the identification of several new photopigment gene families. Significantly, these genes are not expressed in the rods and cones of the retina, but in many sites within the central nervous system. Moreover, molecular approaches have proved useful in clarifying some of the earlier immunocytochemical results. Collectively, the recent findings show that non-mammalian vertebrates possess multiple extraocular photoreceptors that may express novel, rod or even cone photopigments. The future challenge is to link these photoreceptors with circadian and photoperiodic physiology.

Animals↗
PubMed2005

Elephants and human color-blind deuteranopes have identical sets of visual pigments.

Being the largest land mammals, elephants have very few natural enemies and are active during both day and night. Compared with those of diurnal and nocturnal animals, the eyes of elephants and other arrhythmic species, such as many ungulates and large carnivores, must function in both the bright light of day and dim light of night. Despite their fundamental importance, the roles of photosensitive molecules, visual pigments, in arrhythmic vision are not well understood. Here we report that elephants (Loxodonta africana and Elephas maximus) use RH1, SWS1, and LWS pigments, which are maximally sensitive to 496, 419, and 552 nm, respectively. These light sensitivities are virtually identical to those of certain "color-blind" people who lack MWS pigments, which are maximally sensitive to 530 nm. During the day, therefore, elephants seem to have the dichromatic color vision of deuteranopes. During the night, however, they are likely to use RH1 and SWS1 pigments and detect light at 420-490 nm.

Animals↗
PubMedDate not supplied

Melanopsin triggers the release of internal calcium stores in response to light.

Melanopsin is the photopigment that confers photosensitivity upon intrinsically photosensitive retinal ganglion cells (ipRGCs). This subset of retinal ganglion cells comprises less than 2% of all RGCs in the mammalian retina. The paucity of melanopsin-positive cells has made studies on melanopsin signaling difficult to pursue in ipRGCs. To address this issue, we have established several cell lines consisting of a transformed human embryonic kidney cell line (HEK293) stably expressing human melanopsin. With these cell lines, we have investigated the intracellular rise in calcium triggered upon light activation of melanopsin. Our human melanopsin-expressing cells exhibit an irradiance-dependent increase in intracellular calcium. Control cells expressing human melanopsin, where the Schiff-base lysine has been mutated to alanine, show no responses to light. Chelating extracellular calcium has no effect on the light-induced increase in intracellular calcium suggesting that calcium is mobilized from intracellular stores. This involvement of intracellular stores has been confirmed through their depletion by thapsigargin, which inhibits a subsequent light-induced increase in intracellular calcium. Addition of the nonselective cation channel blocker lanthanum does not alter light-induced rises in intracellular calcium, further supporting that melanopsin triggers a release of internal calcium from internal stores. HEK293 cells stably expressing melanopsin have proven to be a useful tool to study melanopsin-initiated signaling.

Base Sequence↗
PubMed1990

Comparison of clinical and experimental uveitis.

Uveitis is a term which encompasses many clinical syndromes which would appear to be discrete entities. Both clinically and experimentally, the separation of uveitis affecting only the anterior segment from that affecting the posterior segment has a sound pathogenetic basis. However, clear distinctions among the various forms of endogenous posterior uveitis are more difficult to maintain in the light of evidence from experimental models of autoimmune uveitis (EAU). EAU can be induced by a variety of retinal antigens and each antigen has been shown to induce somewhat different forms of EAU, depending on such factors as dose of antigen, species and strains of animal model, and the type(s) of adjuvant used. However, within each model a similar spectrum of uveoretinal responses can be induced by each antigen suggesting that the pathogenetic mechanisms are probably similar also. In addition, if these models are analogous to human disease, then each clinical entity within this apparently heterogeneous group of clinical posterior uveitis syndromes may represent one aspect of a general organ-specific uveoretinal response to autoantigens.

Adjuvants, Immunologic↗