Showing papers on "Retinal ganglion cell published in 2004"

Timing and Topography of Cell Genesis in the Rat Retina

Abstract: To understand the mechanisms of cell fate determination in the vertebrate retina, the time course of the generation of the major cell types needs to be established. This will help define and interpret patterns of gene expression, waves of differentiation, timing and extent of competence, and many of the other developmental processes involved in fate acquisition. A thorough retinal cell “birthdating” study has not been performed for the laboratory rat, even though it is the species of choice for many contemporary developmental studies of the vertebrate retina. We investigated the timing and spatial pattern of cell genesis using 3H-thymidine (3H-TdR). A single injection of 3H-TdR was administered to pregnant rats or rat pups between embryonic day (E) 8 and postnatal day (P) 13. The offspring of prenatally injected rats were delivered and all animals survived to maturity. Labeled cells were visualized by autoradiography of retinal sections. Rat retinal cell genesis commenced around E10, 50% of cells were born by approximately P1, and retinogenesis was complete near P12. The first postmitotic cells were found in the retinal ganglion cell layer and were 9–15 μm in diameter. This range includes small to medium diameter retinal ganglion cells and large displaced amacrine cells. The sequence of cell genesis was established by determining the age at which 5, 50, and 95% of the total population of cells of each phenotype became postmitotic. With few exceptions, the cell types reached these developmental landmarks in the following order: retinal ganglion cells, horizontal cells, cones, amacrine cells, rods, bipolar cells, and Muller glia. For each type, the first cells generated were located in the central retina and the last cells in the peripheral retina. Within the sequence of cell genesis, two or three phases could be detected based on differences in timing, kinetics, and topographic gradients of cell production. Our results show that retinal cells in the rat are generated in a sequence similar to that of the primate retina, in which retinogenesis spans more than 100 days. To the extent that sequences reflect underlying mechanisms of cell fate determination, they appear to be conserved. J. Comp. Neurol. 474:304–324, 2004. © 2004 Wiley-Liss, Inc.

Retinal ganglion cells in Alzheimer's disease and aging.

Abstract: Optic nerve and retinal ganglion cell (GC) degeneration are possible explanations for the poor visual function reported in patients with Alzheimer's disease (AD). We investigated whether GC loss could be attributed to AD compared with control subjects by measuring the spatial density of GC (cells/mm2) with methods previously used to analyze the GC distribution of young normal retinas. Retinas from 4 autopsy-confirmed, severely demented patients with AD and 4 age- and sex-matched control subjects (ages, 66-86 yr for both groups) without history of dementing or ocular disease were prepared as unstained whole mounts. There was no evidence for loss of GC within the central 43 degrees of vision in patients with AD. The density of GC subserving the central 11 degrees of vision was reduced by one-fourth in both AD and control eyes compared with retinas from young adults, as was GC density in a wedge of nasal retina. This loss may contribute to deficits in visual function found in aged individuals, whether or not they have dementia.

Cell type-specific effects of Rb deletion in the murine retina

Abstract: Certain cells of the human retina are extremely sensitive to loss of function of the retinoblastoma tumor suppressor gene RB. Retinoblastomas develop early in life and at high frequency in individuals heterozygous for a germ-line RB mutation, and sporadic retinoblastomas invariably have somatic mutation in the RB gene. In contrast, retinoblastomas do not develop in Rb+/- mice. Although retinoblastoma is thought to have developmental origins, the function of Rb in retinal development has not been fully characterized. Here we studied the role of Rb in normal retinal development and in retinoblastoma using conditional Rb mutations in the mouse. In late embryogenesis, Rb-deficient retinas exhibited ectopic S-phase and high levels of p53-independent apoptosis, particularly in the differentiating retinal ganglion cell layer. During postnatal retinal development, loss of Rb led to more widespread retinal apoptosis, and adults showed loss of photoreceptors and bipolar cells. Conditional Rb mutation in the retina did not result in retinoblastoma formation even in a p53-mutant background. However, on a p107- or p130-deficient background, Rb mutation in the retina caused retinal dysplasia or retinoblastoma.

Repulsive guidance molecule (RGM) gene function is required for neural tube closure but not retinal topography in the mouse visual system.

Abstract: The establishment of topographic projections in the developing visual system depends on the spatially and temporally controlled expression of axon guidance molecules. In the developing chick tectum, the graded expression of the repulsive guidance molecule (RGM) has been proposed to be involved in controlling the topography of the retinal ganglion cell (RGC) axon termination zones along the anteroposterior axis of the tectum. We now show that there are three mouse proteins homologous to chick RGM displaying similar proteolytic processing but exhibiting differential cell-surface targeting by glycosyl phosphatidylinositol anchor addition. Two members of this gene family (mRGMa and mRGMb) are expressed in complementary patterns in the nervous system, and mRGMa is expressed prominently in the superior colliculus at the time of anteroposterior targeting of RGC axons. The third member of the family (mRGMc) is expressed almost exclusively in skeletal muscles. Functional studies in the mouse reveal a role for mRGMa in controlling cephalic neural tube closure, thus defining an unexpected role for mRGMa in early embryonic development. In contrast, mRGMa mutant mice did not exhibit defects in anteroposterior targeting of RGC axons to their stereotypic termination zones in the superior colliculus.

Effect of erythropoietin axotomy-induced apoptosis in rat retinal ganglion cells.

Abstract: PURPOSE. Erythropoietin (EPO) modulates erythropoiesis by inhibiting apoptosis in erythrocyte progenitors. Recently, EPO has been shown to be protective in experimental models of mechanical trauma, neuroinflammation, cerebral and retinal ischemia, and even in a human stroke trial. However, little is known about EPO signal transduction in vivo and the usefulness of EPO in the prevention of the chronic, purely apoptotic neuronal cell death that contributes to vision loss in glaucoma and the progression of neurodegenerative diseases. METHODS. EPO's effects and signaling in the retinal ganglion cell axotomy paradigm were studied by Western blot analysis and immunohistochemistry, receptor expression was characterized in the retina before and after lesion. EPO was injected into the vitreous body to investigate neuroprotection of axotomized rat RGCs. Moreover, EPO's effects were studied in cultures of immunopurified retinal ganglion cells. Signal-transduction pathways transmitting neuroprotective EPO effects in vivo were characterized by the use of specific kinase inhibitors, immunohistochemistry, and Western blot analysis. RESULTS. EPO receptors (EPORs) were expressed on RGC somata and dendrites in vivo. EPOR expression did not significantly change after axotomy. Application of EPO prevented death of neurotrophic-factor-deprived immunopurified rat RGCs in vitro, rescued axotomized RGCs in vivo, and prevented caspase-3 activation. EPO-induced Akt phosphorylation and survival-promoting EPO effects were completely abolished by inhibition of PI-3-kinase. EPO neuroprotection followed a bell-shaped dose-response curve in vitro and in vivo, whereas toxic EPO effects were never observed, even at high concentrations. CONCLUSIONS. These data support a potential role for EPO as a therapeutic molecule against predominantly apoptotic neuronal cell death in the context of glaucoma or neurodegenerative diseases and delineate the PI-3-K/Akt pathway as the predominant mediator of EPO neuroprotection in this in vivo paradigm of neuronal cell death.

Convergence and Segregation of the Multiple Rod Pathways in Mammalian Retina

Abstract: Using a multidisciplinary approach, we demonstrate that three different pathways are responsible for the transmission of rod signals across the mouse retina. Each pathway serves a primarily nonoverlapping range of stimulus intensities, with ganglion cells receiving either segregated or convergent inputs. For both on-center (ON) and off-center (OFF) ganglion cells, the primary rod pathway carries signals with the lowest threshold, whereas the secondary rod pathway is less sensitive by approximately 1 log unit. In addition, OFF signaling uses a tertiary rod pathway that is approximately 1 log unit less sensitive than the secondary. Although some ganglion cells received rod inputs exclusively from one of the pathways, others showed convergent inputs. Using pharmacological and genetic approaches, we defined classes of ON and OFF ganglion cells for which the scotopic inputs derive only from the primary pathway or from both primary and secondary pathways. In addition, we observed a class of OFF ganglion cell receiving mixed input from primary and tertiary pathways. Interestingly, OFF ganglion cells receiving convergent inputs from all three rod pathways or from the secondary and tertiary pathways together were never observed. Overall, our data show a complex arrangement of convergence and segregation of rod inputs to ganglion cells in the mammalian retina.

Mechanisms and time course of neuronal degeneration in experimental autoimmune encephalomyelitis.

Abstract: Neuronal and axonal damage is considered to be the main cause for long-term disability in multiple sclerosis. We analyzed the mechanism and kinetics of neuronal cell death in experimental autoimmune encephalomyelitis (EAE) induced by myelin oligodendrocyte glycoprotein (MOG) by combining an electrophysiological in vivo assessment of the optic pathway with the investigation of retinal ganglion cell (RGC) counts. In accordance with our previous findings in this animal model, neuritis of the optic nerve (ON) leads to apoptotic RGC death. By further investigating the time course of RGC apoptosis in the present study, we found that neuronal cell death together with decreased visual acuity values occurred before the onset of clinical symptoms. Simultaneously with the time course of RGC apoptosis, we found a down-regulation of phospho-Akt as well as a shift in the relation of 2 proteins of the Bcl-2 family, Bax and Bcl-2, towards a more proapoptotic ratio in these cells. Comparing the kinetics and mechanisms of RGC death during MOG-EAE with those following complete surgical transection of the ON, we found significant agreement. We hypothesize that the main reason for RGC loss in MOG-EAE is the inflammatory attack but RGC death also occurs independently of histopathological ON changes.

Müller cell response to laser-induced increase in intraocular pressure in rats.

Abstract: The goal of this study was to investigate the reaction of the Muller cells to elevated intraocular pressure (IOP). Elevated IOP is one of the risk factors in glaucomatous retinal ganglion cell (RGC) degeneration. Muller cells play an important role in retinal homeostasis. The reaction of Muller cells was examined by evaluating temporal changes in glutamate aspartate transporter (GLAST), glutamine synthase (GS), glial fibrillary acidic protein (GFAP), and the B-cell lymphoma (Bcl-2) using immunoblotting and immunohistochemical techniques. After IOP was elevated for 4-60 days, there was a time-related decrease in RGC ranging from 6% to 44%. There was also a time-related increase in GLAST protein reaching maximum after 3 weeks of elevated IOP. On the other hand, there was very little change in the expression of GS during the first 2 weeks followed by some increase between 21 and 60 days. An increase in Bcl-2 was biphasic with maximum increase after 4 days followed by decline after 15 and 21 days. GFAP, which is usually not expressed in normal Muller cells, was present at all time points. In all cases, the increase was most intense in the vicinity of the ganglion cells where the astrocytes and endfeet of the Muller cells are located. These results indicate that Muller cells react to the insult of elevated IOP by expressing GFAP and Bcl-2, proteins that are expressed in reactive gliosis and other pathological conditions. The increase in GLAST along with minimum change in GS indicates a disturbance in glutamate homeostasis.

Discrete gene sets depend on POU domain transcription factor Brn3b/Brn-3.2/POU4f2 for their expression in the mouse embryonic retina

Abstract: Brn3b/Brn-3.2/POU4f2 is a POU domain transcription factor that is essential for retinal ganglion cell (RGC) differentiation, axonal outgrowth and survival. Our goal was to establish a link between Brn3b and the downstream events leading to RGC differentiation. We sought to determine both the number and types of genes that depend on Brn3b for their expression. RNA probes from wild-type and Brn3b -/- E14.5, E16.5 and E18.5 mouse retinas were hybridized to a microarray containing 18,816 retina-expressed cDNAs. At E14.5, we identified 87 genes whose expression was significantly altered in the absence of Brn3b and verified the results by real-time PCR and in situ hybridization. These genes fell into discrete sets that encoded transcription factors, proteins associated with neuron integrity and function, and secreted signaling molecules. We found that Brn3b influenced gene expression in non RGCs of the retina by controlling the expression of secreted signaling molecules such as sonic hedgehog and myostatin/Gdf8. At later developmental stages, additional alterations in gene expression were secondary consequences of aberrant RGC differentiation caused by the absence of Brn3b. Our results demonstrate that a small but crucial fraction of the RGC transcriptome is dependent on Brn3b. The Brn3b-dependent gene sets therefore provide a unique molecular signature for the developing retina.

Leber hereditary optic neuropathy mtDNA mutations disrupt glutamate transport in cybrid cell lines

Abstract: Leber hereditary optic neuropathy (LHON) is a maternally inherited form of retinal ganglion cell degeneration leading to optic atrophy which is caused by point mutations in the mitochondrial genome (mtDNA). Three pathogenic mutations (positions 11778/ND4, 3460/ND1 and 14484/ND6) account for the majority of LHON cases and they affect genes that encode for different subunits of mitochondrial complex I. Excitotoxic injury to retinal ganglion cells and the optic nerve has been previously hypothesized, especially given the high susceptibility of this neural cell type to glutamate toxicity. Osteosarcoma-derived cytoplasmic hybrids (cybrids) generated from six unrelated LHON patients, two cell lines for each pathogenic mutation, were compared with cybrids obtained from three healthy controls. Molecular and biochemical analyses showed that excitatory amino acid transporter 1 (EAAT1)/GLAST is the most active glutamate transporter in this cellular model. The glutamate uptake maximal velocity was significantly reduced in all LHON cybrids compared with control cybrids. This reduction was correlated in a mutation-specific fashion with the degree of mitochondrial production of reactive oxygen species, which is enhanced in LHON cybrids. Our findings support the hypothesis that the genetically determined mitochondrial dysfunction in LHON patients leads to impaired activity of the EAAT1 glutamate transporter. This observation is particularly relevant since EAAT1 is the major means of glutamate removal in the inner retina and this prevents retinal ganglion cells being damaged as a result of excitotoxicity.

A relative signalling model for the formation of a topographic neural map

Abstract: The highly ordered wiring of retinal ganglion cell (RGC) neurons in the eye to their synaptic targets in the superior colliculus of the midbrain has long served as the dominant experimental system for the analysis of topographic neural maps1,2,3. Here we describe a quantitative model for the development of one arm of this map—the wiring of the nasal–temporal axis of the retina to the caudal–rostral axis of the superior colliculus. The model is based on RGC–RGC competition that is governed by comparisons of EphA receptor signalling intensity, which are made using ratios of, rather than absolute differences in, EphA signalling between RGCs4. Molecular genetic experiments, exploiting a combinatorial series of EphA receptor knock-in and knockout mice, confirm the salient predictions of the model, and show that it both describes and predicts topographic mapping.

Changes in retinal expression of neurotrophins and neurotrophin receptors induced by ocular hypertension.

Abstract: Open angle glaucoma is defined as a progressive and time-dependent death of retinal ganglion cells concomitant with high intraocular pressure, leading to loss of visual field. Because neurotrophins are a family of growth factors that support neuronal survival, we hypothesized that quantitative and qualitative changes in neurotrophins or their receptors may take place early in ocular hypertension, preceding extensive cell death and clinical features of glaucoma. We present molecular, biochemical, and phenotypic evidence that significant neurotrophic changes occur in retina, which correlate temporally with retinal ganglion cell death. After 7 days of ocular hypertension there is a transient up-regulation of retinal NGF, while its receptor TrkA is up-regulated in a sustained fashion in retinal neurons. After 28 days of ocular hypertension there is sustained up-regulation of retinal BDNF, but its receptor TrkB remains unchanged. Throughout, NT-3 levels remain unchanged but there is an early and sustained increase of its receptor TrkC in Muller cells but not in retinal ganglion cells. These newly synthesized glial TrkC receptors are truncated, kinase-dead isoforms. Expression of retinal p75 also increases late at day 28. Asymmetric up-regulation of neurotrophins and neurotrophin receptors may preclude efficient neurotrophic rescue of RGCs from apoptosis. A possible rationale for therapeutic intervention with Trk receptor agonists and p75 receptor antagonists is proposed.

Foxd1 is required for proper formation of the optic chiasm

Abstract: In animals with binocular vision, retinal ganglion cell (RGC) axons from each eye sort in the developing ventral diencephalon to project to ipsi- or contralateral targets, thereby forming the optic chiasm. Ipsilaterally projecting axons arise from the ventrotemporal (VT) retina and contralaterally projecting axons primarily from the other retinal quadrants. The winged helix transcription factor Foxd1 (previously known as BF-2, Brain Factor 2) is expressed in VT retina, as well as in the ventral diencephalon during the formation of the optic chiasm. We report here that in embryos lacking Foxd1, both retinal development and chiasm morphogenesis are disrupted. In the Foxd1 deficient retina, proteins designating the ipsilateral projection, such as Zic2 and EphB1, are missing, and the domain of Foxg1 (BF-1) expands from nasal retina into the VT crescent. In retina-chiasm co-cultures, VT RGCs from Foxd1 deficient retina are not repulsed by chiasm cells, and in vivo many VT RGCs aberrantly project contralaterally. However, even though the ipsilateral program is lost in the retina, a larger than normal uncrossed component develops in Foxd1 deficient embryos. Chiasm defects include axon stalling in the chiasm and a reduction in the total number of RGCs projecting to the optic tract. In addition, in the Foxd1 deficient ventral diencephalon, Foxg1 invades the Foxd1 domain, Zic2 and Islet1 expression are minimized, and Slit2 prematurely expands, changes that could contribute to axon projection errors. Thus, Foxd1 plays a dual role in the establishment of the binocular visual pathways: first, in specification of the VT retina, acting upstream of proteins directing the ipsilateral pathway; and second, in the patterning of the developing ventral diencephalon where the optic chiasm forms.

Neurotrophic regulation of retinal ganglion cell synaptic connectivity: from axons and dendrites to synapses

Abstract: This review highlights important events during the morphological development of retinal ganglion cells (RGCs), focusing on mechanisms that control axon and dendritic arborization as a means to understand synaptic connectivity with special emphasis on the role of neurotrophins during structural and functional development of RGCs. Neurotrophins and their receptors participate in the development of visual connectivity at multiple levels. In the visual system, neurotrophins have been shown to exert various developmental influences, from guiding the morphological differentiation of neurons to controlling the functional plasticity of visual circuits. This review article examines the role of neurotrophins, and in particular of BDNF, during the morphological development of RGCs, and discusses potential interactions between activity and neurotrophins during development of neuronal connectivity.

The Ginkgo biloba extract (EGb 761) provides a neuroprotective effect on retinal ganglion cells in a rat model of chronic glaucoma

Abstract: Purpose To investigate the effect of Ginkgo biloba extract (EGb 761) against neurotoxicity of retinal ganglion cells of rats with chronic moderately elevated intraocular pressure (IOP) Methods Unilateral chronic moderately elevated IOP was produced in rats by cautery of three episcleral vessels Secondary degeneration was measured with and without EGb 761 for 5 months At 5 months, retinal ganglion cells were labeled with a fast blue tracer applied to both superior colliculi Densities of surviving retinal ganglion cells were estimated by counting fast blue labeled cells in whole mounted retinas Results When compared with their contralateral control eyes with normal IOP, in the peripheral retina, retinal ganglion cell loss in eyes with chronic, moderately elevated IOP was 298 ± 15% (n = 5) at 5 months in untreated animals and 46 ± 45% (n = 5) at 5 months in treated animals with EGb 761 Conclusions Pretreatment and early posttreatment with EGb 761 is an effective neuroprotectant in a rat model o

Circulation and axonal transport in the optic nerve

Abstract: Retinal ganglion cells are the output cells of the retina whose axons are under considerable metabolic stress in both health and disease states. They are highly polarised to ensure that mitochondria and enzymes involved in the generation of ATP are strategically concentrated to meet the local energy demands of the cell. In passing from the eye to the brain, axons are protected and supported by glial tissues and the blood supply of the optic nerve head is regulated to maintain the supply of oxygen and nutrients to the axons. In spite of this, the optic nerve head remains the point at which retinal ganglion cell axons are most vulnerable to the effects of increased intraocular pressure or ischaemia. Considerable work has been undertaken in this area to advance our understanding on the pathophysiology of axon damage and to develop new strategies for the prevention of retinal ganglion cell death.

The winged helix transcription factor Foxg1 facilitates retinal ganglion cell axon crossing of the ventral midline in the mouse.

Abstract: During normal development, retinal ganglion cells (RGCs) project axons along the optic nerve to the optic chiasm on the ventral surface of the hypothalamus. In rodents, most RGC growth cones then cross the ventral midline to join the contralateral optic tract; those that do not cross join the ipsilateral optic tract. Contralaterally projecting RGCs are distributed across the retina whereas ipsilaterally projecting RGCs are concentrated in temporal retina. The transcription factor Foxg1 (also known as BF1) is expressed at several key locations along this pathway. Analysis of Foxg1 expression using lacZ reporter transgenes shows that Foxg1 is normally expressed in most, if not all, nasal RGCs but not in most temporal RGCs, neither at the time they project nor earlier in their lineage. Foxg1 is also expressed at the optic chiasm. Mice that lack Foxg1 die at birth and, although the shape of their eyes is abnormal, their retinas still project axons to the brain via the optic chiasm. Using anterograde and retrograde tract tracing, we show that there is an eightfold increase in the ipsilateral projection in Foxg1-/- embryos. The distributions of cells expressing the transcription factors Foxg1 and Nkx2.2, and cell-surface molecules Ephb2, ephrin B2 and SSEA-1 (Fut4) have been correlated to the normally developing retinothalamic projection and we show they are not much altered in the developing Foxg1-/- retina and optic chiasm. As much of the increased ipsilateral projection in Foxg1-/- embryos arises from temporal RGCs that are unlikely to have an autonomous requirement for Foxg1, we propose that the phenotype reflects at least in part a requirement for Foxg1 outwith the RGCs themselves, most likely at the optic chiasm.

Expression of P2Y1, P2Y2, P2Y4, and P2Y6 receptor subtypes in the rat retina.

Abstract: Purpose To elucidate the expression pattern of different types of metabotropic P2Y receptors in the adult rat retina. Methods Qualitative RT-PCR was used to investigate the expression profile of different P2Y receptor subtypes (P2Y1, P2Y2, P2Y4, and P2Y6), and in situ hybridization studies were performed to show their cellular localization within the retina. Immunohistochemical staining was used to detect the corresponding P2Y proteins (P2Y1, P2Y2, and P2Y4) and their cellular localization. Southern blot analysis and sequencing verified the identity of the P2Y PCR products. Results RT-PCR revealed the presence of P2Y1, -2, -4, and -6 mRNA in the neural retina and the retinal pigment epithelium (RPE) and choroid. In situ hybridization showed labeling in the retinal ganglion cell layer for all four P2Y receptor subtypes, although the intensity varied. In addition, staining for P2Y1, -4, and -6 mRNA was shown in the inner nuclear layer, but was absent for the P2Y2 receptor subtype. Immunohistochemistry showed intense staining for P2Y1, -2, and -4 in the ganglion cell layer and the outer plexiform layer. There was also a specific subtype staining in the inner plexiform layer (P2Y2, -4), the inner (P2Y1, -4) and outer (P2Y1) nuclear layers and the inner segments of the photoreceptors (P2Y1, -2). discussion. The data suggest that extracellular nucleotides may play complex roles as autocrine-paracrine mediators and may have neuromodulatory effects in the retina through metabotropic P2Y receptors.