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[A study on transplantation of rat photoreceptor cell layer].

OBJECTIVE: To study the relationship between photoreceptors and other layers of the retina in the aspects of anatomical structure, physiology, pathology, etc. METHODS: RCS rats were used as host animals, and the pure cone and rod layer of the donor retina was prepared from Wistar rat and was transplanted under the host retina where the cones and rods were degenerated. RESULTS: After 2 weeks of transplantation, the retinas of the host rats were reattached, and the transplanted cells were survival. The transplanted cells were in the subretinal space between the retinal pigment epithelium (RPE) and the inner unclear layer, and they had the staining characteristics similar to that of the normal control photoreceptors. CONCLUSION: In the animal model of photoreceptor cell layer transplantation, the transplanted cells possess normal anatomical location and are connected with RPE and inner nuclear layer. The results provide a new route to investigate the re-establishment of anatomical association with the optic nerve and the physiological activity of the nerve.

Animals↗

Immunocytochemical localization of taurine in the developing retina of the lefteye flounder Paralichthys olivaceus.

Light microscopic immunolocalization of taurine, a sulfur-containing free amino acid, was investigated in the developing retina of a lefteye flounder, Paralichthys olivaceus, which exhibits metamorphic changes with rod cell addition for 3-5 weeks after hatching. This immunocytochemical study of the developing retina revealed: 1) From 3 to 13 days after hatching, intense immunostaining was shifted from the surroundings of neural cells to the neural somata and processes in the inner retina. 2) Intense immunoreactivity appeared also in the outer and inner segments and basal processes (pedicles) of cone cells within 6 days or 13 days after hatching. 3) Lack of immunoreactivity was found in the outer segment of rod cells from their appearance during metamorphosis. These findings are discussed with the possible functional roles of taurine in the fish retina: 1) involvement in cell differentiation and/or development; 2) protection of the outer segments against light stimuli; and 3) regulation of neural transmission.

Age Factors↗

Photoreceptor transplants increase host cone survival in the retinal degeneration (rd) mouse.

Retinal transplants offer a potentially interesting approach to treating human retinal degenerations, but so far little quantitative data are available on possible beneficial effects. We isolated photoreceptor layers from normal-sighted mice and grafted them into the subretinal space of retinal degeneration (rd) mice lacking rod photoreceptors. At 2 weeks after surgery, the numbers of residual host cone photoreceptors outside the graft zone were quantified following specific labelling. Examination of operated retinas revealed highly significantly greater numbers of surviving cones (mean of 38% more at 2 weeks) within the central field compared to sham-operated paired control retinas (p < 0.01). These are the first quantified data indicating a trophic effect of transplanted photoreceptors upon host cone cells. As cone cells are responsible for high acuity and colour vision, such data could have important implications not only for eventual therapeutic approaches to human retinal degenerations but also to understanding underlying interactions between retinal photoreceptors.

Animals↗

Oxidative damage is a potential cause of cone cell death in retinitis pigmentosa.

Retinitis pigmentosa (RP) is a prevalent cause of blindness caused by a large number of different mutations in many different genes. The mutations result in rod photoreceptor cell death, but it is unknown why cones die. In this study, we tested the hypothesis that cones die from oxidative damage by performing immunohistochemical staining for biomarkers of oxidative damage in a transgenic pig model of RP. The presence of acrolein- and 4-hydroxynonenal-adducts on proteins is a specific indicator that lipid peroxidation has occurred, and there was strong immunofluorescent staining for both in cone inner segments (IS) of two 10-month-old transgenic pigs in which almost all rods had died, compared to faint staining in two 10-month-old control pig retinas. In 22- and 24-month-old transgenic pigs in which all rods and many cones had died, staining was strong in cone axons and some cell bodies as well as IS indicating progression in oxidative damage between 10 and 22 months. Biomarkers for oxidative damage to proteins and DNA also showed progressive oxidative damage to those macromolecules in cones during the course of RP. These data support the hypothesis that the death of rods results in decreased oxygen consumption and hyperoxia in the outer retina resulting in gradual cone cell death from oxidative damage. This hypothesis has important therapeutic implications and deserves rapid evaluation.

Acrolein↗

Morphogenesis of the different types of photoreceptors of the chicken (Gallus domesticus) retina and the effect of amblyopia in neonatal chicken.

Despite the great variety in chicken photoreceptors, existing morphogenetic studies only deal with two types: rods and cones. We have therefore examined by scanning electron microscopy the first appearance and maturation of different retinal photoreceptors in 36 chicken embryos (Gallus domesticus), aged 5-19 days prehatching. On day 5 of incubation, chicken retinae were only composed of proliferating ventricular cells devoid of photoreceptors. On day 8, outer mitotic cells were separated from inner differentiating photoreceptors, by the transient layer of Chievitz. Ball-like protrusions appeared at the ventricular surface, representing the first signs of photoreceptor inner segment formation. From day 10 onward, double cones, single cones, and rods could be clearly distinguished, and occasional cilia were detected at their tip. On day 12, inner segments had increased in length and diameter, and frequently carried a cilium representing the beginning of outer segment formation. On day 14, most photoreceptors displayed a distinct outer segment. On day 19, photoreceptors had essentially assumed adult morphology. Based on the shape of their outer segments, two subtypes of cones and three subtypes of double cones could be distinguished. Throughout development, we observed microvilli close to maturing photoreceptors, either originating from their lateral sides, from their tip, or from Müller cells. Microvillus density peaked between day 12 and 14, indicating an important role in photoreceptor morphogenesis. Unilateral occlusion of the eyes of posthatching chicken reduced the proportion of double cones to single cones in the retina, indicating dependence of retinal morphogenesis upon functional activity of visual cells.

Amblyopia↗

Time course and development of light adaptation processes in the outer zebrafish retina.

Retinomotor movements are morphological changes in the outer retina in response to changing light conditions. They can be separated into two components: Migration of pigment granules within the microvilli of the retinal pigment epithelium (RPE) and positional changes in photoreceptor cells. These positional changes optimize exposure of the cone and rod photoreceptors to light. The aim of this study was to analyze both the time course of retinomotor movements in the adult zebrafish and the maturation of these processes in the developing fish. We show that retinomotor movements are used as a dark/light adaptation mechanism in zebrafish. In adult zebrafish, melanin granules of the RPE migrate with constant speed and reach the fully light adapted (LA) state approximately after 1 h. In contrast, about two thirds of double cone outer segment movements are finished in 5 min, and are fully completed in 10 to 20 min. During development there are three crucial stages leading to mature retinomotor movements in response to light: at 5 dpf (days post fertilization) the migration of pigment granules begins, at 20 dpf the pigment granules condense in the apical part of the RPE microvilli, and at 28 dpf, concomitant with the functional maturation of rods, the double cones contract as in adult retinas.

Adaptation, Ocular↗

Like night and day: rods and cones have different pigment regeneration pathways.

Sustained vision requires continuous regeneration of visual pigments in rod and cone photoreceptors by the 11-cis-retinal chromophore. In this issue of Neuron, Mata et al. report a novel enzymatic pathway uniquely designed to keep up with the high demand for cone pigment regeneration in bright light and to preclude rods from utilizing chromophore produced in daylight, when rods are not very useful for vision.

Animals↗

Chromatic sensitivity of ganglion cells in the peripheral primate retina.

Visual abilities change over the visual field. For example, our ability to detect movement is better in peripheral vision than in foveal vision, but colour discrimination is markedly worse. The deterioration of colour vision has been attributed to reduced colour specificity in cells of the midget, parvocellular (PC) visual pathway in the peripheral retina. We have measured the colour specificity (red-green chromatic modulation sensitivity) of PC cells at eccentricities between 20 and 50 degrees in the macaque retina. Here we show that most peripheral PC cells have red-green modulation sensitivity close to that of foveal PC cells. This result is incompatible with the view that PC pathway cells in peripheral retina make indiscriminate connections ('random wiring') with retinal circuits devoted to different spectral types of cone photoreceptors. We show that selective cone connections can be maintained by dendritic field anisotropy, consistent with the morphology of PC cell dendritic fields in peripheral retina. Our results also imply that postretinal mechanisms contribute to the psychophysically demonstrated deterioration of colour discrimination in the peripheral visual field.

Animals↗

Light-induced photoreceptor damage triggers DNA repair: differential fate of rods and cones.

Intense light triggers cell death in cones and rods by two distinct mechanisms: rapid, necrosis-like cell death occurs in cones; prolonged, apoptotic death is displayed by rods. However, the areas of cone and rod loss coincide, suggesting some interrelationship. DNA fragmentation in rods occurs in two waves. Our evidence suggests a repair mechanism that triggers up-regulation of the enzyme DNA polymerase beta, implying a rod-specific mechanism that initially attempts DNA repair, but which subsequently fails, resulting in a second wave of damage leading to apoptotic rod cell death. This study suggests two different but coincidental pathways lead to photoreceptor death. Definition of these two pathways could suggest useful pharmacologic sites toward which compounds could be targeted to promote cell survival in damaging retinal degenerative diseases. In addition, maintenance or enhancement of existing in-house repair mechanisms could provide additional approaches for the rescue of neurons with stress-induced nuclear damage.

Animals↗

Subsets of retinal progenitors display temporally regulated and distinct biases in the fates of their progeny.

Cell fate determination in the developing vertebrate retina is characterized by the sequential generation of seven classes of cells by multipotent progenitor cells. Despite this order of genesis, more than one cell type is generated at any time; for example, in the rat, several cell types are born during the prenatal period, while others are born postnatally. In order to examine whether there are classes of progenitor cells with distinct developmental properties contributing to this developmental progression, we examined antigen expression in progenitor cells during rat retinal development. Two markers of amacrine and horizontal cells, the VC1.1 epitope and syntaxin, were found to be expressed on a subset of progenitors in a temporally regulated manner that closely paralleled the birthdays of these cell types. In order to investigate which cell types were produced by the progenitors expressing these markers, fluorescent latex microspheres covalently coupled to VC1.1 antibodies were used to indelibly label VC1.1+ progenitor cells and their progeny. Early in retinal development, VC1.1+ progenitors generated a high percentage of amacrine and horizontal cells, but no cone photoreceptors. During this same period, a comparable number of cone photoreceptors were generated by VC1.1- progenitors. In the late embryonic and early postnatal period, VC1.1+ progenitors continued to generate predominantly amacrine cells, but also gave rise to an increasing number of rod photoreceptors. These findings demonstrate that expression of these two markers by progenitors is highly correlated with a bias towards the production of amacrine and horizontal cells. The fact that subsets of progenitors with temporally regulated and distinct biases are intermingled within the retinal neuroepithelium provides a basis for understanding how different cell types are generated both simultaneously and in a particular order by multipotent progenitors during retinal development.

Aging↗

Melanopsin-expressing ganglion cells in primate retina signal colour and irradiance and project to the LGN.

Human vision starts with the activation of rod photoreceptors in dim light and short (S)-, medium (M)-, and long (L)- wavelength-sensitive cone photoreceptors in daylight. Recently a parallel, non-rod, non-cone photoreceptive pathway, arising from a population of retinal ganglion cells, was discovered in nocturnal rodents. These ganglion cells express the putative photopigment melanopsin and by signalling gross changes in light intensity serve the subconscious, 'non-image-forming' functions of circadian photoentrainment and pupil constriction. Here we show an anatomically distinct population of 'giant', melanopsin-expressing ganglion cells in the primate retina that, in addition to being intrinsically photosensitive, are strongly activated by rods and cones, and display a rare, S-Off, (L + M)-On type of colour-opponent receptive field. The intrinsic, rod and (L + M) cone-derived light responses combine in these giant cells to signal irradiance over the full dynamic range of human vision. In accordance with cone-based colour opponency, the giant cells project to the lateral geniculate nucleus, the thalamic relay to primary visual cortex. Thus, in the diurnal trichromatic primate, 'non-image-forming' and conventional 'image-forming' retinal pathways are merged, and the melanopsin-based signal might contribute to conscious visual perception.

Animals↗

Bipolar cell diversity in the primate retina: morphologic and immunocytochemical analysis of a new world monkey, the marmoset Callithrix jacchus.

The aim of this study was to identify the bipolar cell types in the retina of a New World monkey, the common marmoset, and compare them with those found in the Old World macaque monkey. Retinal whole-mounts, sections, or both, were stained by using DiI labeling and immunohistochemical methods. Semithin sections were analyzed by using quantitative methods. We show that the same morphologic types of bipolar cell as described for the Old World macaque monkey by Boycott and Wässle (Boycott and Wässle [1991] Eur. J. Neurosci. 3:1069-1088) are present in marmoset retina: two types of midget bipolar cells, six type of diffuse bipolar cells, a blue cone bipolar cell, and one type of rod bipolar cell. The pattern of staining with different immunohistochemical markers ("fingerprint") of each bipolar cell type in marmoset was also the same as described for macaque, with one exception: the flat midget bipolar cell (FMB) class is labeled by antibodies to recoverin in macaque but is labeled by antibodies to CD15 in marmoset. The labeled FMB cells in marmoset make contact with multiple cone photoreceptors throughout most of the extrafoveal retina. The spatial density of bipolar cells in marmoset is shown to be sufficient to support one-to-one connectivity of midget bipolar and ganglion cells in the fovea and to allow for parallel pathways to ganglion cells throughout the retina. Quantitative differences in the morphology and receptor connectivity between marmoset and macaque can be related to differences in cone and rod photoreceptor density between the species. We conclude that bipolar cell diversity is a preserved feature of the primate retina.

ATP-Binding Cassette Transporters↗

Distribution of two splice variants of the glutamate transporter GLT-1 in the developing rat retina.

The distributions of a carboxyl terminal splice variant of the glutamate transporter GLT-1, referred to as GLT-1B, and the carboxyl terminus of the originally described variant of GLT-1, referred to hereafter as GLT-1 alpha, were examined using specific antisera. GLT-1B was present in the retina at very early developmental stages. Labelling was demonstrable at embryonic day 14, and strong labelling was evident by embryonic day 18. Such labelling was initially restricted to populations of cone photoreceptors, the processes of which extended through the entire thickness of the retina and appeared to make contact with the retinal ganglion cells. During postnatal development the GLT-1B-positive photoreceptor processes retracted to form the outer plexiform layer, and around postnatal day 7, GLT-1B-immunoreactive bipolar cells appeared. The pattern of labelling of bipolar cell processes within the inner plexiform layer changed during postnatal development. Two strata of strongly immunoreactive terminals were initially evident in the inner plexiform layer, but by adulthood these two bands were no longer evident and labelling was restricted to the somata and processes (but not synaptic terminals) of the bipolar cells, as well as the somata, processes, and terminals of cone photoreceptors. By contrast, GLT-1 alpha appeared late in postnatal development and was restricted mainly to a population of amacrine cells, although transient labelling was also associated with punctate elements in the outer plexiform layer, which may represent photoreceptor terminals.

Aging↗

Quantitative estimations of foveal and extra-foveal retinal circuitry in humans.

For an understanding of the basis for psychophysical measurement of visual resolution, quantitative morphological studies of retinal neuronal architecture are needed. Here we report on cell densities and retinal ganglion cell:cone ratio (RGC:C) from the foveal border to the peripheral retina (34 degrees eccentricity). Quantitative estimates of RGC and C densities were made using a modified disector method in three vertically sectioned human retinae and were adjusted for RGC displacement. In agreement with our previous data on humans, we found an RGC:C ratio close to 3 at 2-3 degrees eccentricity. Outside the foveal border, the ratio declined to 1.0 at 7.5 degrees eccentricity and to 0.5 at eccentricities larger than 19 degrees. Center-to-center separation of C and RGC in addition to center-to-center separation of estimated 'receptive fields' was calculated at corresponding locations along the superior and inferior hemimeridians. The center-to-center separation of estimated 'receptive fields' was found to be more closely related to resolution thresholds from the fovea to 19 degrees eccentricity than was the separation of RGC and C. On the basis of these quantitative estimates, models for neural circuitry involved in central and peripheral spatial vision can be discussed.

Cell Count↗

Multivesicular release and saturation of glutamatergic signalling at retinal ribbon synapses.

Pronounced multivesicular release (MVR) occurs at the ribbon synapses of sensory neurones that signal via graded potential changes. As MVR increases the likelihood of postsynaptic receptor saturation, it is of interest to consider how sensory synapses overcome this problem and use MVR to encode signals of widely varying intensities. Here, I discuss three postsynaptic mechanisms that permit three different retinal synapses to utilize MVR.

Amacrine Cells↗

Connections of two types of flat cone bipolars in the rabbit retina.

The synaptic connections of two types of cone bipolar cells in the rabbit retina were studied with the electron microscope after labeling in vitro with 4',6-diamidino-2-phenylindole (DAPI), intracellular injection with Lucifer Yellow, and photooxidation (Mills and Massey [1992] J. Comp. Neurol. 321:133). Both types of bipolars belong to the flat variety, because they make basal junctions with a group of four to ten neighboring cone pedicles. One cell type has an axonal arborization that occupies strata 1 through 3 of the inner plexiform layer (IPL). At ribbon synaptic junctions, it is presynaptic to ganglion cell dendrites and to reciprocal dendrites belonging to narrow-field bistratified (AII) amacrine cells. In addition, it contacts and is contacted by other amacrine cell processes of unknown origin. The other cell type has an axonal arborization entirely confined to stratum 2 of the IPL; it is pre- or postsynaptic to a pleomorphic population of amacrine cell processes, and, in particular, it receives input from the lobular appendages of AII. Thus, these two bipolar types probably belong to the off-variety because they make basal junctions with cone photoreceptors and send their axon to sublamina a of the IPL, which is occupied by the dendrites of off-ganglion cells. They are also part of the rod pathway because they receive input from AII amacrine cells.

Animals↗

Age-related distribution of cones and ON-bipolar cells in the rd mouse retina.

PURPOSE: Retinal dystrophic (rd) mice lose most of their rod photoreceptors within the first three weeks after birth. We determined the age-related distribution of peanut agglutinin lectin (PNA)-labeled cones during the first 12 months of age. We also investigated whether the density of ON-bipolar cells expressing L7 protein was affected by their loss of photoreceptor inputs. METHODS: rd mice were selected from a transgenic strain which expresses an L7-beta-galactosidase fusion gene localized to ON-bipolar cells. Cones were stained with PNA and ON-bipolar cells with bluo-gal (halogenated indolyl-beta-D-galactoside). Retinas were flat-mounted and observed at 1, 2, 3, 6 and 12 months of age. RESULTS: PNA-labeled cones are distributed unevenly across the retina at 1 month postnatal. Their concentration decreases first in the central and far peripheral retina, leaving a ring of labeled cells in the midperipheral region. At 3 months, a larger patch of cones remains in the supero-temporal midperipheral region and a smaller patch in the infero-nasal retina. By 6 months, few cones remain in the infero-nasal retina; by 1 year approximately 100 cones remain in the entire retina, localized to the superior midperipheral region. ON-bipolar cells appear evenly distributed at 1 month. By 2-3 months, relatively more bluo-gal staining is seen in the midperipheral regions underlying dense cone populations. At 6-12 months, bluo-gal label is distributed in a spotty pattern with little or no staining seen in areas of apparent neovascularization. CONCLUSIONS: (1) PNA-labeling of cones in the rd retina deteriorates in a distinct spatial pattern with the longest cone survival in the midperipheral superior retina. (2) ON-bipolar cells are more densely labeled in regions of high cone density during the early months of cone degeneration and, in later stages, show relative decreases in regions of apparent neovascularization.

Age Factors↗

Disease sequence from mutant rhodopsin allele to rod and cone photoreceptor degeneration in man.

Mutations in the gene encoding rhodopsin, the visual pigment in rod photoreceptors, lead to retinal degeneration in species from Drosophila to man. The pathogenic sequence from rod cell-specific mutation to degeneration of rods and cones remains unclear. To understand the disease process in man, we studied heterozygotes with 18 different rhodopsin gene mutations by using noninvasive tests of rod and cone function and retinal histopathology. Two classes of disease expression were found, and there was allele-specificity. Class A mutants lead to severely abnormal rod function across the retina early in life; topography of residual cone function parallels cone cell density. Class B mutants are compatible with normal rods in adult life in some retinal regions or throughout the retina, and there is a slow stereotypical disease sequence. Disease manifests as a loss of rod photoreceptor outer segments, not singly but in microscopic patches that coalesce into larger irregular areas of degeneration. Cone outer segment function remains normal until >75% of rod outer segments are lost. The topography of cone loss coincides with that of rod loss. Most class B mutants show an inferior-nasal to superior-temporal retinal gradient of disease vulnerability associated with visual cycle abnormalities. Class A mutant alleles behave as if cytotoxic; class B mutants can be relatively innocuous and epigenetic factors may play a major role in the retinal degeneration.

Adult↗