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Sample records for ganglion cell neurite

  1. Zebrafish diras1 Promoted Neurite Outgrowth in Neuro-2a Cells and Maintained Trigeminal Ganglion Neurons In Vivo via Rac1-Dependent Pathway.

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    Yeh, Chi-Wei; Hsu, Li-Sung

    2016-12-01

    The small GTPase Ras superfamily regulates several neuronal functions including neurite outgrowth and neuron proliferation. In this study, zebrafish diras1a and diras1b were identified and were found to be mainly expressed in the central nervous system and dorsal neuron ganglion. Overexpression of green fluorescent protein (GFP)-diras1a or GFP-diras1b triggered neurite outgrowth of Neuro-2a cells. The wild types, but not the C terminus truncated forms, of diras1a and diras1b elevated the protein level of Ras-related C3 botulinum toxin substrate 1 (Rac1) and downregulated Ras homologous member A (RhoA) expression. Glutathione S-transferase (GST) pull-down assay also revealed that diras1a and diras1b enhanced Rac1 activity. Interfering with Rac1, Pak1, or cyclin-dependent kinase 5 (CDK5) activity or with the Arp2/3 inhibitor prevented diras1a and diras1b from mediating the neurite outgrowth effects. In the zebrafish model, knockdown of diras1a and/or diras1b by morpholino antisense oligonucleotides not only reduced axon guidance but also caused the loss of trigeminal ganglion without affecting the precursor markers, such as ngn1 and neuroD. Co-injection with messenger RNA (mRNA) derived from mouse diras1 or constitutively active human Rac1 restored the population of trigeminal ganglion. In conclusion, we provided preliminary evidence that diras1 is involved in neurite outgrowth and maintains the number of trigeminal ganglions through the Rac1-dependent pathway.

  2. Decreased intracellular granule movement and glucagon secretion in pancreatic α cells attached to superior cervical ganglion neurites.

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    Watabe, Kiyoto; Yokawa, Satoru; Inoh, Yoshikazu; Suzuki, Takahiro; Furuno, Tadahide

    2018-01-09

    Autonomic neurons innervate pancreatic islets of Langerhans and participate in the maintenance of blood glucose concentrations by controlling hormone levels through attachment with islet cells. We previously found that stimulated superior cervical ganglia (SCG) could induce Ca 2+ oscillation in α cells via neuropeptide substance P using an in vitro co-culture model. In this study, we studied the effect of SCG neurite adhesion on intracellular secretory granule movement and glucagon secretion in α cells stimulated by low glucose concentration. Spinning disk microscopic analysis revealed that the mean velocity of intracellular granules was significantly lower in α cells attached to SCG neurites than that in those without neurites under low (2 mM), middle (10 mM), and high (20 mM) glucose concentrations. Stimulation by a low (2 mM) glucose concentration significantly increased glucagon secretion in α cells lacking neurites but not in those bound to neurites. These results suggest that adhesion to SCG neurites decreases low glucose-induced glucagon secretion in pancreatic α cells by attenuating intracellular granule movement activity.

  3. Synergistic effects of osteonectin and brain-derived neurotrophic factor on axotomized retinal ganglion cells neurite outgrowth via the mitogen-activated protein kinase-extracellular signal-regulated kinase 1/2 pathways.

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    Ma, C H E; Bampton, E T W; Evans, M J; Taylor, J S H

    2010-01-20

    Our previous study identified osteonectin (ON) in a screen of factors made by Schwann cells (SCs) which promoted peripheral and central neurons survival and neuritogenesis, however, the mechanisms of ON promoting effects are largely unknown. In the present study, we investigated the effects of ON-deficient SC-conditioned medium (SCCM) and molecular mechanisms of ON, in regulating retinal ganglion cells (RGCs) survival and neurite outgrowth. Neonatal rat RGCs and SCs were purified by immunopanning technique. RGC survival and neuritogenesis reduced significantly when treated with either ON-null mice SCCM or ON-immunodepleted (IP) SCCM (Pshowed the synergistic effects of ON and brain-derived neurotrophic factor (BDNF) in promoting RGCs growth and the involvement of ON in two major neurotrophin-mediated signaling pathways, PI-3K-Akt and MAPK-Erk1/2. ON alone activated Akt phosphorylation and increased survival. Blockage of TrkB signalling pathway by TrkB-Fc chimera (BDNF scavenger) or K252a in ON-treated cultures reduced Akt-P level significantly. This suggests that ON induces BDNF synthesis and secretion from RGCs. The enhancement of neuritogenesis and Erk1/2 phosphorylation by ON in BDNF-treated cultures further demonstrate the signaling pathways responsible for the synergistic effect of ON on BDNF-induced neurite outgrowth. To the best of our knowledge, this is the first report showing the synergistic effects of ON on classical neurotrophins which participate in the same signalling pathways in regulating RGC neurite outgrowth.

  4. Chronic neurotrophin delivery promotes ectopic neurite growth from the spiral ganglion of deafened cochleae without compromising the spatial selectivity of cochlear implants.

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    Landry, Thomas G; Fallon, James B; Wise, Andrew K; Shepherd, Robert K

    2013-08-15

    Cochlear implants restore hearing cues in the severe-profoundly deaf by electrically stimulating spiral ganglion neurons (SGNs). However, SGNs degenerate following loss of cochlear hair cells, due at least in part to a reduction in the endogenous neurotrophin (NT) supply, normally provided by hair cells and supporting cells of the organ of Corti. Delivering exogenous NTs to the cochlea can rescue SGNs from degeneration and can also promote the ectopic growth of SGN neurites. This resprouting may disrupt the cochleotopic organization upon which cochlear implants rely to impart pitch cues. Using retrograde labeling and confocal imaging of SGNs, we determined the extent of neurite growth following 28 days of exogenous NT treatment in deafened guinea pigs with and without chronic electrical stimulation (ES). On completion of this treatment, we measured the spread of neural activation to intracochlear ES by recording neural responses across the cochleotopically organized inferior colliculus using multichannel recording techniques. Although NT treatment significantly increased both the length and the lateral extent of growth of neurites along the cochlea compared with deafened controls, these anatomical changes did not affect the spread of neural activation when examined immediately after 28 days of NT treatment. NT treatment did, however, result in lower excitation thresholds compared with deafened controls. These data support the application of NTs for improved clinical outcomes for cochlear implant patients. Copyright © 2013 Wiley Periodicals, Inc.

  5. Inhibitory Activity of Yokukansankachimpihange against Nerve Growth Factor-Induced Neurite Growth in Cultured Rat Dorsal Root Ganglion Neurons

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    Chiaki Murayama

    2015-08-01

    Full Text Available Chronic pruritus is a major and distressing symptom of many cutaneous diseases, however, the treatment remains a challenge in the clinic. The traditional Chinese-Japanese medicine (Kampo medicine is a conservative and increasingly popular approach to treat chronic pruritus for both patients and medical providers. Yokukansankachimpihange (YKH, a Kampo formula has been demonstrated to be effective in the treatment of itching of atopic dermatitis in Japan although its pharmacological mechanism is unknown clearly. In an attempt to clarify its pharmacological actions, in this study, we focused on the inhibitory activity of YKH against neurite growth induced with nerve growth factor (NGF in cultured rat dorsal root ganglion (DRG neurons because epidermal hyperinnervation is deeply related to itch sensitization. YKH showed approximately 200-fold inhibitory activity against NGF-induced neurite growth than that of neurotropin (positive control, a drug used clinically for treatment of chronic pruritus. Moreover, it also found that Uncaria hook, Bupleurum root and their chemical constituents rhynchophylline, hirsutine, and saikosaponin a, d showed inhibitory activities against NGF-induced neurite growth, suggesting they should mainly contribute to the inhibitory activity of YKH. Further study on the effects of YKH against epidermal nerve density in “itch-scratch” animal models is under investigation.

  6. Stratification of alpha ganglion cells and ON/OFF directionally selective ganglion cells in the rabbit retina

    OpenAIRE

    ZHANG, JIAN; LI, WEI; HOSHI, HIDEO; MILLS, STEPHEN L.; MASSEY, STEPHEN C.

    2005-01-01

    The correlation between cholinergic sensitivity and the level of stratification for ganglion cells was examined in the rabbit retina. As examples, we have used ON or OFF α ganglion cells and ON/OFF directionally selective (DS) ganglion cells. Nicotine, a cholinergic agonist, depolarized ON/OFF DS ganglion cells and greatly enhanced their firing rates but it had modest excitatory effects on ON or OFF α ganglion cells. As previously reported, we conclude that DS ganglion cells are the most sens...

  7. The effects of canine bone marrow stromal cells on neuritogenesis from dorsal root ganglion neurons in vitro.

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    Kamishina, Hiroaki; Cheeseman, Jennifer A; Clemmons, Roger M

    2009-10-01

    The present in vitro study was designed to evaluate whether canine bone marrow stromal cells (BMSCs) promote neurite outgrowth from dorsal root ganglion (DRG) neurons. Bone marrow aspirates were collected from iliac crests of three young adult dogs. DRG neurons were cultured on BMSCs, fibroblasts, or laminin substrates. DRG neurons were also cultured in BMSC- or fibroblast-conditioned media. DRG neurons grown on BMSCs extended longer neurites and developed a much more elaborate conformation of branching neurites compared to those on fibroblasts or laminin. Quantitative analysis revealed that these effects were associated with the emergence of increased numbers of primary and branching neurites. The effect appears to be dependent upon cell-cell interactions rather than by elaboration of diffusible molecules. With more extensive investigations into the basic biology of canine BMSCs, their ability for promoting neurite outgrowth may be translated into a novel therapeutic strategy for dogs with a variety of neurological disorders.

  8. Mechanosensitivity of Embryonic Neurites Promotes Their Directional Extension and Schwann Cells Progenitors Migration

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    Gonzalo Rosso

    2017-11-01

    Full Text Available Background/Aims: Migration of Schwann cells (SCs progenitors and neurite outgrowth from embryonic dorsal root ganglions (DRGs are two central events during the development of the peripheral nervous system (PNS. How these two enthralling events preceding myelination are promoted is of great relevance from basic research and clinical aspects alike. Recent evidence demonstrates that biophysical cues (extracellular matrix stiffness and biochemical signaling act in concert to regulate PNS myelination. Microenvironment stiffness of SCs progenitors and embryonic neurites dynamically changes during development. Methods: DRG explants were isolated from day 12.5 to 13.5 mice embryos and plated on laminin-coated substrates with varied stiffness values. After 4 days in culture and immunostaining with specific markers, neurite outgrowth pattern, SCs progenitors migration, and growth cone shape and advance were analyzed with confocal fluorescence microscopy. Results: We found out that growing substrate stiffness promotes directional neurite outgrowth, SCs progenitors migration, growth cone advance and presumably axons fasciculation. Conclusions: DRG explants are in vitro models for the research of PNS development, myelination and regeneration. Consequently, we conclude the following: Our observations point out the importance of mechanosensitivity for the PNS. At the same time, they prompt the investigation of the important yet unclear links between PNS biomechanics and inherited neuropathies with myelination disorders such as Charcot-Marie-Tooth 1A and hereditary neuropathy with liability to pressure palsies. Finally, they encourage the consideration of mechanosensitivity in bioengineering of scaffolds to aid nerve regeneration after injury.

  9. Somatic and neuritic spines on tyrosine hydroxylase-immunopositive cells of rat retina.

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    Fasoli, Anna; Dang, James; Johnson, Jeffrey S; Gouw, Aaron H; Fogli Iseppe, Alex; Ishida, Andrew T

    2017-05-01

    Dopamine- and tyrosine hydroxylase-immunopositive cells (TH cells) modulate visually driven signals as they flow through retinal photoreceptor, bipolar, and ganglion cells. Previous studies suggested that TH cells release dopamine from varicose axons arborizing in the inner and outer plexiform layers after glutamatergic synapses depolarize TH cell dendrites in the inner plexiform layer and these depolarizations propagate to the varicosities. Although it has been proposed that these excitatory synapses are formed onto appendages resembling dendritic spines, spines have not been found on TH cells of most species examined to date or on TH cell somata that release dopamine when exposed to glutamate receptor agonists. By use of protocols that preserve proximal retinal neuron morphology, we have examined the shape, distribution, and synapse-related immunoreactivity of adult rat TH cells. We report here that TH cell somata, tapering and varicose inner plexiform layer neurites, and varicose outer plexiform layer neurites all bear spines, that some of these spines are immunopositive for glutamate receptor and postsynaptic density proteins (viz., GluR1, GluR4, NR1, PSD-95, and PSD-93), that TH cell somata and tapering neurites are also immunopositive for a γ-aminobutyric acid (GABA) receptor subunit (GABA A R α1 ), and that a synaptic ribbon-specific protein (RIBEYE) is found adjacent to some colocalizations of GluR1 and TH in the inner plexiform layer. These results identify previously undescribed sites at which glutamatergic and GABAergic inputs may stimulate and inhibit dopamine release, especially at somata and along varicose neurites that emerge from these somata and arborize in various levels of the retina. J. Comp. Neurol. 525:1707-1730, 2017. © 2016 Wiley Periodicals, Inc. © 2017 Wiley Periodicals, Inc.

  10. Changes in ganglion cells during retinal degeneration.

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    Saha, Susmita; Greferath, Ursula; Vessey, Kirstan A; Grayden, David B; Burkitt, Anthony N; Fletcher, Erica L

    2016-08-04

    Inherited retinal degeneration such as retinitis pigmentosa (RP) is associated with photoreceptor loss and concomitant morphological and functional changes in the inner retina. It is not known whether these changes are associated with changes in the density and distribution of synaptic inputs to retinal ganglion cells (RGCs). We quantified changes in ganglion cell density in rd1 and age-matched C57BL/6J-(wildtype, WT) mice using the immunocytochemical marker, RBPMS. Our data revealed that following complete loss of photoreceptors, (∼3months of age), there was a reduction in ganglion cell density in the peripheral retina. We next examined changes in synaptic inputs to A type ganglion cells by performing double labeling experiments in mice with the ganglion cell reporter lines, rd1-Thy1 and age-matched wildtype-Thy1. Ribbon synapses were identified by co-labelling with CtBP2 (RIBEYE) and conventional synapses with the clustering molecule, gephyrin. ON RGCs showed a significant reduction in RIBEYE-immunoreactive synapse density while OFF RGCs showed a significant reduction in the gephyrin-immmunoreactive synapse density. Distribution patterns of both synaptic markers across the dendritic trees of RGCs were unchanged. The change in synaptic inputs to RGCs was associated with a reduction in the number of immunolabeled rod bipolar and ON cone bipolar cells. These results suggest that functional changes reported in ganglion cells during retinal degeneration could be attributed to loss of synaptic inputs. Copyright © 2016 IBRO. Published by Elsevier Ltd. All rights reserved.

  11. Melanopsin Ganglion Cells Are the Most Resistant Retinal Ganglion Cell Type to Axonal Injury in the Rat Retina

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    Pérez de Sevilla Müller, Luis; Sargoy, Allison; Rodriguez, Allen R.; Brecha, Nicholas C.

    2014-01-01

    We report that the most common retinal ganglion cell type that remains after optic nerve transection is the M1 melanopsin ganglion cell. M1 ganglion cells are members of the intrinsically photosensitive retinal ganglion cell population that mediates non-image-forming vision, comprising ∼2.5% of all ganglion cells in the rat retina. In the present study, M1 ganglion cells comprised 1.7±1%, 28±14%, 55±13% and 82±8% of the surviving ganglion cells 7, 14, 21 and 60 days after optic nerve transect...

  12. 7, 8, 3′-Trihydroxyflavone Promotes Neurite Outgrowth and Protects Against Bupivacaine-Induced Neurotoxicity in Mouse Dorsal Root Ganglion Neurons

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    Shi, Haohong; Luo, Xingjing

    2016-01-01

    Background 7, 8, 3′-trihydroxyflavone (THF) is a novel pro-neuronal small molecule that acts as a TrkB agonist. In this study, we examined the effect of THF on promoting neuronal growth and protecting anesthetics-induced neurotoxicity in dorsal root ganglion (DRG) neurons in vitro. Material/Methods Neonatal mouse DRG neurons were cultured in vitro and treated with various concentrations of THF. The effect of THF on neuronal growth was investigated by neurite outgrowth assay and Western blot. In addition, the protective effects of THF on bupivacaine-induced neurotoxicity were investigated by apoptosis TUNEL assay, neurite outgrowth assay, and Western blot, respectively. Results THF promoted neurite outgrowth of DRG neurons in dose-dependent manner, with an EC50 concentration of 67.4 nM. Western blot analysis showed THF activated TrkB signaling pathway by inducing TrkB phosphorylation. THF also rescued bupivacaine-induced neurotoxicity by reducing apoptosis and protecting neurite retraction in DRG neurons. Furthermore, the protection of THF in bupivacaine-injured neurotoxicity was directly associated with TrkB phosphorylation in a concentration-dependent manner in DRG neurons. Conclusions THF has pro-neuronal effect on DRG neurons by promoting neurite growth and protecting against bupivacaine-induced neurotoxicity, likely through TrkB activation. PMID:27371503

  13. Ganglion cell neurites in human idiopathic epiretinal membranes

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    Lesnik Oberstein, S. Y.; Lewis, G. P.; Chapin, E. A.; Fisher, S. K.

    2008-01-01

    AIM: To identify and confirm the presence of neural elements in idiopathic epiretinal membranes removed from patients' eyes during vitrectomy with epiretinal membrane peeling. METHODS: Human epiretinal membranes from patients with no other known eye disease and of varying durations were labelled

  14. Retinal ganglion cell topography in elasmobranchs.

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    Bozzano, A; Collin, S P

    2000-04-01

    Retinal wholemounts are used to examine the topographic distribution of retinal cells within the ganglion cell layer in a range of elasmobranchs from different depths. The retina is examined for regional specializations for acute vision in six species of selachians, Galeocerdo cuvieri, Hemiscyllium ocellatum, Scyliorhinus canicula, Galeus melastomus, Etmopterus spinax, Isistius brasiliensis, one species of batoid, Raja bigelowi and one species of chimaera, Hydrolagus mirabilis. These species represent a range of lifestyles including pelagic, mesopelagic and benthic habitats, living from shallow water to the sea bottom at a depth of more than 3000 m. The topography of cells within the ganglion cell layer is non-uniform and changes markedly across the retina. Most species possess an increased density of cells across the horizontal (dorsal) meridian or visual streak, with a density range of 500 to 2,500 cells per mm(2) with one or more regional increases in density lying within this specialized horizontal area. It is proposed that the higher spatial resolving power provided by the horizontal streak in these species mediates panoramic vision in the lower frontal visual field. Only I. brasiliensis possesses a concentric arrangement of retinal iso-density contours in temporal retina or an area centralis, thereby increasing spatial resolving power in a more specialized part of the visual field, an adaptation for its unusual feeding behavior. In Nissl-stained material, amacrine and ganglion cell populations could be distinguished on the criteria of soma size, soma shape and nuclear staining. Quantitative analyses show that the proportion of amacrine cells lying within the ganglion cell layer is non-uniform and ranges between 0.4 and 12.3% in specialized retinal areas and between 8.2 and 48.1% in the peripheral non-specialized regions. Analyses of soma area of the total population of cells in the ganglion cell layer also show that the pelagic species possess significantly

  15. Learning LM Specificity for Ganglion Cells

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    Ahumada, Albert J.

    2015-01-01

    Unsupervised learning models have been proposed based on experience (Ahumada and Mulligan, 1990;Wachtler, Doi, Lee and Sejnowski, 2007) that allow the cortex to develop units with LM specific color opponent receptive fields like the blob cells reported by Hubel and Wiesel on the basis of visual experience. These models used ganglion cells with LM indiscriminate wiring as inputs to the learning mechanism, which was presumed to occur at the cortical level.

  16. Interplay between cell migration and neurite outgrowth determines SH2B1β-enhanced neurite regeneration of differentiated PC12 cells.

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    Chia-Ling Wu

    Full Text Available The regulation of neurite outgrowth is crucial in developing strategies to promote neurite regeneration after nerve injury and in degenerative diseases. In this study, we demonstrate that overexpression of an adaptor/scaffolding protein SH2B1β promotes neurite re-growth of differentiated PC12 cells, an established neuronal model, using wound healing (scraping assays. Cell migration and the subsequent remodeling are crucial determinants during neurite regeneration. We provide evidence suggesting that overexpressing SH2B1β enhances protein kinase C (PKC-dependent cell migration and phosphatidylinositol 3-kinase (PI3K-AKT-, mitogen activated protein kinase (MAPK/extracellular signal-regulated protein kinase (ERK kinase (MEK-ERK-dependent neurite re-growth. Our results further reveal a cross-talk between pathways involving PKC and ERK1/2 in regulating neurite re-growth and cell migration. We conclude that temporal regulation of cell migration and neurite outgrowth by SH2B1β contributes to the enhanced regeneration of differentiated PC12 cells.

  17. Interplay between Cell Migration and Neurite Outgrowth Determines SH2B1β-Enhanced Neurite Regeneration of Differentiated PC12 Cells

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    Teng, Nan-Yuan; Hsu, Hsin-Ling; Chen, Linyi

    2012-01-01

    The regulation of neurite outgrowth is crucial in developing strategies to promote neurite regeneration after nerve injury and in degenerative diseases. In this study, we demonstrate that overexpression of an adaptor/scaffolding protein SH2B1β promotes neurite re-growth of differentiated PC12 cells, an established neuronal model, using wound healing (scraping) assays. Cell migration and the subsequent remodeling are crucial determinants during neurite regeneration. We provide evidence suggesting that overexpressing SH2B1β enhances protein kinase C (PKC)-dependent cell migration and phosphatidylinositol 3-kinase (PI3K)-AKT-, mitogen activated protein kinase (MAPK)/extracellular signal-regulated protein kinase (ERK) kinase (MEK)-ERK-dependent neurite re-growth. Our results further reveal a cross-talk between pathways involving PKC and ERK1/2 in regulating neurite re-growth and cell migration. We conclude that temporal regulation of cell migration and neurite outgrowth by SH2B1β contributes to the enhanced regeneration of differentiated PC12 cells. PMID:22539954

  18. Staurosporine induces ganglion cell differentiation in part by stimulating urokinase-type plasminogen activator expression and activation in the developing chick retina

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    Kim, Yeoun-Hee; Chang, Yongmin; Jung, Jae-Chang

    2012-01-01

    Highlights: ► Staurosporine mediates stimulation of RGC differentiation in vitro cultured retinal neuroblasts. ► Staurosporine mediates uPA activation during RGC differentiation in vitro. ► Inhibition of uPA blocks the staurosporine mediated RGC differentiation both in vitro and in ovo. ► Thus, uPA may play a role in the staurosporine-mediated stimulation of RGC differentiation. -- Abstract: Here, we investigated whether staurosporine-mediated urokinase-type plasminogen activator (uPA) activation is involved in retinal ganglion cell (RGC) differentiation. Retinal cells were isolated from developing chick retinas at embryonic day 6 (E6). Relatively few control cells grown in serum-free medium started to form processes by 12 h. In contrast, staurosporine-treated cells had processes within 3 h, and processes were evident at 8 h. Immunofluorescence staining showed that Tuj-1-positive cells with shorter neurites could be detected in control cultures at 18 h, whereas numerous Tuj-1 positive ganglion cells with longer neuritic extensions were seen in staurosporine-treated cultures. BrdU-positive proliferating cells were more numerous in control cultures than in staurosporine-treated cultures, and the BrdU staining was not detected in post-mitotic Tuj-1 positive ganglion cells. Western blotting of cell lysates showed that staurosporine induced high levels of the active form of uPA. The staurosporine-induced uPA signal was localized predominantly in the soma, neurites and axons of Tuj-1-positive ganglion cells. Amiloride, an inhibitor of uPA, markedly reduced staurosporine-induced Tuj-1 staining, neurite length, neurite number, and uPA staining versus controls. In developing retinas in ovo, amiloride administration remarkably reduced the staurosporine-induced uPA staining and RGC differentiation. Taken together, our in vitro and in vivo data collectively indicate that uPA plays a role in the staurosporine-mediated stimulation of RGC differentiation.

  19. Stimulation of autophagy prevents amyloid-β peptide-induced neuritic degeneration in PC12 cells.

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    Yang, Yi; Chen, Sicong; Zhang, Jiafeng; Li, Chentan; Sun, Yonghong; Zhang, Lihui; Zheng, Xiaoxiang

    2014-01-01

    Autophagy is a lysosomal degradative process essential for neuronal homeostasis, whereas autophagic failure has been linked to accumulating neurodegenerative disorders. However, the precise role of autophagy in axonal and dendritic degeneration in Alzheimer's disease (AD) remains unclear. In this study, we aim to investigate the precise effect of autophagy in amyloid-β peptide (Aβ)25-35-mediated neurite degeneration. Aβ35-25, the non-neurotoxic reverse sequence analogue of Aβ25-35, was used as a negative control. Our results showed that Aβ25-35 dose-dependently suppressed PC12 proliferation and induced autophagy induction in neurites (axons and dendrites). A high proportion of autophagic structures in PC12 neurites were autolysosomes after 24 h of Aβ25-35 treatment. Autophagy inhibition by 3-methyladenine (3MA), LY294002, and chloroquine (CQ) could not relieve the Aβ25-35-induced neurite degeneration, while administration of autophagy stimulator rapamycin or AR-12 efficiently suppressed neurite degeneration. Autophagosomes colocalized with fragmented mitochondria after Aβ25-35 treatment. Similar results were obtained using in vitro cultured superior cervical ganglion neurons. These findings demonstrate that autophagy stimulation may prevent neuritic degeneration following Aβ25-35 treatment. Upregulation of autophagic activity may provide a valuable approach for the treatment of axonal and dendritic dystrophy in AD patients.

  20. Topography of ganglion cell production in the cat's retina

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    Walsh, C.; Polley, E.H.

    1985-01-01

    The ganglion cells of the cat's retina form several classes distinguishable in terms of soma size, axon diameter, dendritic morphology, physiological properties, and central connections. Labeling with [ 3 H]thymidine shows that the ganglion cells which survive in the adult are produced as several temporally shifted, overlapping waves: medium-sized cells are produced before large cells, whereas the smallest ganglion cells are produced throughout the period of ganglion cell generation. Large cells and medium-sized cells show the same distinctive pattern of production, forming rough spirals around the area centralis. The oldest cells tend to lie superior and nasal to the area centralis, whereas cells in the inferior nasal retina and inferior temporal retina are, in general, progressively younger. Within each retinal quadrant, cells nearer the area centralis tend to be older than cells in the periphery, but there is substantial overlap. The retinal raphe divides the superior temporal quadrant into two zones with different patterns of cell addition. Superior temporal retina near the vertical meridian adds cells only slightly later than superior nasal retina, whereas superior temporal retina near the horizontal meridian adds cells very late, contemporaneously with inferior temporal retina. The broader wave of production of smaller ganglion cells seems to follow this same spiral pattern at its beginning and end. The presence of the area centralis as a nodal point about which ganglion cell production in the retinal quadrants pivots suggests that the area centralis is already an important retinal landmark even at the earliest stages of retinal development

  1. Frequency Responses of Rat Retinal Ganglion Cells.

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    Alex E Hadjinicolaou

    Full Text Available There are 15-20 different types of retinal ganglion cells (RGC in the mammalian retina, each encoding different aspects of the visual scene. The mechanism by which post-synaptic signals from the retinal network generate spikes is determined by each cell's intrinsic electrical properties. Here we investigate the frequency responses of morphologically identified rat RGCs using intracellular injection of sinusoidal current waveforms, to assess their intrinsic capabilities with minimal contributions from the retinal network. Recorded cells were classified according to their morphological characteristics (A, B, C or D-type and their stratification (inner (i, outer (o or bistratified in the inner plexiform layer (IPL. Most cell types had low- or band-pass frequency responses. A2, C1 and C4o cells were band-pass with peaks of 15-30 Hz and low-pass cutoffs above 56 Hz (A2 cells and ~42 Hz (C1 and C4o cells. A1 and C2i/o cells were low-pass with peaks of 10-15 Hz (cutoffs 19-25 Hz. Bistratified D1 and D2 cells were also low-pass with peaks of 5-10 Hz (cutoffs ~16 Hz. The least responsive cells were the B2 and C3 types (peaks: 2-5 Hz, cutoffs: 8-11 Hz. We found no difference between cells stratifying in the inner and outer IPL (i.e., ON and OFF cells or between cells with large and small somas or dendritic fields. Intrinsic physiological properties (input resistance, spike width and sag had little impact on frequency response at low frequencies, but account for 30-40% of response variability at frequencies >30 Hz.

  2. Neurite development in PC12 cells on flexible micro-textured substrates under cyclic stretch.

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    Haq, Furqan; Keith, Charles; Zhang, Guigen

    2006-01-01

    We investigated the combined effect of micro-texture and mechanical strain on neuronal cell development such as neurite length and neurite density in a rat pheochromocytoma cell line (PC12 cells). Cells were seeded on flexible silicone substrates with micro-texture or no texture (smooth) and cultured under static and dynamic conditions. In the static condition substrates were not stretched and in the dynamic conditions substrates were subjected to cyclic uniaxial stretching at three different strain levels of 4%, 8%, and 16% with each at three different strain rates at 0.1, 0.5, and 1.0 Hz. Results showed that of all cell cultures there was no significant difference in neurite development between cells on smooth and textured substrates, except in the static and 4% at 0.1 Hz conditions, where micro-texture induced significantly longer neurites. With both types of substrates, a lower mechanical condition (4% at 1.0 Hz or 16% at 0.1 Hz) resulted in more and longer neurites and lower cell density, and a higher mechanical condition (16% at 1.0 Hz) resulted in fewer and shorter neurites and lower cell density as compared to the static condition. These findings suggest that the effect of the micro-texture on neurite development is more prominent in low mechanical conditions than in high mechanical conditions and that the strain level and strain rate have an interrelated effect on neurite development: a higher strain level at a lower strain rate has a similar effect as a lower strain level at a higher strain rate in terms of promoting neurite development.

  3. Ionic channel changes in glaucomatous retinal ganglion cells: multicompartment modeling.

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    Maturana, Matias I; Turpin, Andrew; McKendrick, Allison M; Kameneva, Tatiana

    2014-01-01

    This research takes a step towards discovering underlying ionic channel changes in the glaucomatous ganglion cells. Glaucoma is characterized by a gradual death of retinal ganglion cells. In this paper, we propose a hypothesis that the ionic channel concentrations change during the progression of glaucoma. We use computer simulation of a multi-compartment morphologically correct model of a mouse retinal ganglion cell to verify our hypothesis. Using published experimental data, we alter the morphology of healthy ganglion cells to replicate glaucomatous cells. Our results suggest that in glaucomatous cell, the sodium channel concentration decreases in the soma by 30% and by 60% in the dendrites, calcium channel concentration decreases by 10% in all compartments, and leak channel concentration increases by 40% in the soma and by 100% in the dendrites.

  4. Polymodal Sensory Integration in Retinal Ganglion Cells.

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    Križaj, David

    2016-01-01

    An animal's ability to perceive the external world is conditioned by its capacity to extract and encode specific features of the visual image. The output of the vertebrate retina is not a simple representation of the 2D visual map generated by photon absorptions in the photoreceptor layer. Rather, spatial, temporal, direction selectivity and color "dimensions" of the original image are distributed in the form of parallel output channels mediated by distinct retinal ganglion cell (RGC) populations. We propose that visual information transmitted to the brain includes additional, light-independent, inputs that reflect the functional states of the retina, anterior eye and the body. These may include the local ion microenvironment, glial metabolism and systemic parameters such as intraocular pressure, temperature and immune activation which act on ion channels that are intrinsic to RGCs. We particularly focus on light-independent mechanical inputs that are associated with physical impact, cell swelling and intraocular pressure as excessive mechanical stimuli lead to the counterintuitive experience of "pressure phosphenes" and/or debilitating blinding disease such as glaucoma and diabetic retinopathy. We point at recently discovered retinal mechanosensitive ion channels as examples through which molecular physiology brings together Greek phenomenology, modern neuroscience and medicine. Thus, RGC output represents a unified picture of the embodied context within which vision takes place.

  5. Melanopsin retinal ganglion cell loss in Alzheimer's disease

    DEFF Research Database (Denmark)

    La Morgia, Chiara; Ross-Cisneros, Fred N; Koronyo, Yosef

    2015-01-01

    OBJECTIVE: Melanopsin retinal ganglion cells (mRGCs) are photoreceptors driving circadian photoentrainment, and circadian dysfunction characterizes Alzheimer's disease (AD). We investigated mRGCs in AD, hypothesizing their contribution to circadian dysfunction. METHODS: We assessed retinal nerve...

  6. Signaling mechanisms of neurite outgrowth induced by the cell adhesion molecules NCAM and N-cadherin

    DEFF Research Database (Denmark)

    Hansen, S M; Berezin, V; Bock, E

    2008-01-01

    Formation of appropriate neural circuits depends on a complex interplay between extracellular guiding cues and intracellular signaling events that result in alterations of cytoskeletal dynamics and a neurite growth response. Surface-expressed cell adhesion molecules (CAMs) interact with the surro......Formation of appropriate neural circuits depends on a complex interplay between extracellular guiding cues and intracellular signaling events that result in alterations of cytoskeletal dynamics and a neurite growth response. Surface-expressed cell adhesion molecules (CAMs) interact...... extracellular guidance cues to intracellular events and thereby regulating neurite outgrowth. In this review, we focus on two CAMs, the neural cell adhesion molecule (NCAM) and N-cadherin, and their ability to mediate signaling associated with a neurite outgrowth response. In particular, we will focus on direct...

  7. THE MODULATORY ROLE OF TAURINE IN RETINAL GANGLION CELLS

    OpenAIRE

    Jiang, Zheng; Bulley, Simon; Guzzone, Joseph; Ripps, Harris; Shen, Wen

    2013-01-01

    Taurine (2-aminoethylsuphonic acid) is present in nearly all animal tissues, and is the most abundant free amino acid in muscle, heart, CNS and retina. Although it is known to be a major cytoprotectant and essential for normal retinal development, its role in retinal neurotransmission and modulation is not well understood. We investigated the response of taurine in retinal ganglion cells, and its effect on synaptic transmission between ganglion cells and their pre-synaptic neurons. We find th...

  8. Troxler Fading, Eye Movements, and Retinal Ganglion Cell Properties

    Directory of Open Access Journals (Sweden)

    Romain Bachy

    2014-12-01

    Full Text Available We present four movies demonstrating the effect of flicker and blur on the magnitude and speed of adaptation for foveal and peripheral vision along the three color axes that isolate retinal ganglion cells projecting to magno, parvo, and konio layers of the LGN. The demonstrations support the eye movement hypothesis for Troxler fading for brightness and color, and demonstrate the effects of flicker and blur on adaptation of each class of retinal ganglion cells.

  9. Neurite outgrowth in cultured mouse pelvic ganglia - Effects of neurotrophins and bladder tissue.

    Science.gov (United States)

    Ekman, Mari; Zhu, Baoyi; Swärd, Karl; Uvelius, Bengt

    2017-07-01

    Neurotrophic factors regulate survival and growth of neurons. The urinary bladder is innervated via both sympathetic and parasympathetic neurons located in the major pelvic ganglion. The aim of the present study was to characterize the effects of the neurotrophins nerve growth factor (NGF), brain derived neurotrophic factor (BDNF) and neurotrophin 3 (NT-3) on the sprouting rate of sympathetic and parasympathetic neurites from the female mouse ganglion. The pelvic ganglion was dissected out and attached to a petri dish and cultured in vitro. All three factors (BDNF, NT-3 and NGF) stimulated neurite outgrowth of both sympathetic and parasympathetic neurites although BDNF and NT-3 had a higher stimulatory effect on parasympathetic ganglion cells. The neurotrophin receptors TrkA, TrkB and TrkC were all expressed in neurons of the ganglia. Co-culture of ganglia with urinary bladder tissue, but not diaphragm tissue, increased the sprouting rate of neurites. Active forms of BDNF and NT-3 were detected in urinary bladder tissue using western blotting whereas tissue from the diaphragm expressed NGF. Neurite outgrowth from the pelvic ganglion was inhibited by a TrkB receptor antagonist. We therefore suggest that the urinary bladder releases trophic factors, including BDNF and NT-3, which regulate neurite outgrowth via activation of neuronal Trk-receptors. These findings could influence future strategies for developing pharmaceuticals to improve re-innervation due to bladder pathologies. Copyright © 2017. Published by Elsevier B.V.

  10. The modulatory role of taurine in retinal ganglion cells.

    Science.gov (United States)

    Jiang, Zheng; Bulley, Simon; Guzzone, Joseph; Ripps, Harris; Shen, Wen

    2013-01-01

    Taurine (2-aminoethylsuphonic acid) is present in nearly all animal tissues, and is the most abundant free amino acid in muscle, heart, CNS, and retina. Although it is known to be a major cytoprotectant and essential for normal retinal development, its role in retinal neurotransmission and modulation is not well understood. We investigated the response of taurine in retinal ganglion cells, and its effect on synaptic transmission between ganglion cells and their presynaptic neurons. We find that taurine-elicited currents in ganglion cells could be fully blocked by both strychnine and SR95531, glycine and GABA(A) receptor antagonists, respectively. This suggests that taurine-activated receptors might share the antagonists with GABA and glycine receptors. The effect of taurine at micromolar concentrations can effectively suppress spontaneous vesicle release from the presynaptic neurons, but had limited effects on light-evoked synaptic signals in ganglion cells. We also describe a metabotropic effect of taurine in the suppression of light-evoked response in ganglion cells. Clearly, taurine acts in multiple ways to modulate synaptic signals in retinal output neurons, ganglion cells.

  11. THE MODULATORY ROLE OF TAURINE IN RETINAL GANGLION CELLS

    Science.gov (United States)

    Jiang, Zheng; Bulley, Simon; Guzzone, Joseph; Ripps, Harris; Shen, Wen

    2017-01-01

    Taurine (2-aminoethylsuphonic acid) is present in nearly all animal tissues, and is the most abundant free amino acid in muscle, heart, CNS and retina. Although it is known to be a major cytoprotectant and essential for normal retinal development, its role in retinal neurotransmission and modulation is not well understood. We investigated the response of taurine in retinal ganglion cells, and its effect on synaptic transmission between ganglion cells and their pre-synaptic neurons. We find that taurine-elicited currents in ganglion cells could be fully blocked by both strychnine and SR95531, glycine and GABAA receptor antagonists, respectively. This suggests that taurine-activated receptors might share the antagonists with GABA and glycine receptors. The effect of taurine at micromolar concentrations can effectively suppress spontaneous vesicle release from the pre-synaptic neurons, but had limited effects on light-evoked synaptic signals in ganglion cells. We also describe a metabotropic effect of taurine in the suppression of light-evoked response in ganglion cells. Clearly, taurine acts in multiple ways to modulate synaptic signals in retinal output neurons, ganglion cells. PMID:23392924

  12. CHLORHEXIDINE INHIBITS L1 CELL ADHESION MOLECULE MEDIATED NEURITE OUTGROWTH IN VITRO

    Science.gov (United States)

    Milstone, Aaron M.; Bamford, Penny; Aucott, Susan W.; Tang, Ningfeng; White, Kimberly R.; Bearer, Cynthia F.

    2013-01-01

    Background Chlorhexidine is a skin disinfectant that reduces skin and mucous membrane bacterial colonization and inhibits organism growth. Despite numerous studies assessing chlorhexidine safety in term infants, residual concerns have limited its use in hospitalized neonates, especially low birth weight preterm infants. The aim of this study was to assess the potential neurotoxicity of chlorhexidine on the developing central nervous system using a well-established in vitro model of neurite outgrowth that includes laminin and L1 cell adhesion molecule (L1) as neurite outgrowth promoting substrates. Methods Cerebellar granule neurons are plated on either poly L-lysine, L1 or laminin. Chlorhexidine, hexachlorophene or their excipients are added to the media. Neurons are grown for 24 h, then fixed and neurite length measured. Results Chlorhexidine significantly reduced the length of neurites grown on L1 but not laminin. Chlorhexidine concentrations as low as 125 ng/ml statistically significantly reduced neurite length on L1. Hexachlorophene did not affect neurite length. Conclusion Chlorhexidine at concentrations detected in the blood following topical applications in preterm infants specifically inhibited L1 mediated neurite outgrowth of cerebellar granule neurons. It is now vital to determine whether the blood brain barrier is permeable to chlorhexidine in preterm infants. PMID:24126818

  13. Transcription Factor Brn-3b Overexpression Enhances Neurite Outgrowth in PC12 Cells Under Condition of Hypoxia.

    Science.gov (United States)

    Phatak, Nitasha R; Stankowska, Dorota L; Krishnamoorthy, Raghu R

    2015-08-01

    Transcription factor Brn-3b plays a key role in retinal ganglion cell differentiation, survival, and axon outgrowth during development. However, the precise role of Brn-3b in the normal adult retina as well as during neurodegeneration is unclear. In the current study, the effect of overexpression of Brn-3b was assessed in vitro, in PC12 cells under conditions of normoxia and hypoxia. Immunoblot analysis showed that overexpression of Brn-3b in PC12 cells as well as 661W cells produced significant increase in the growth cone marker, growth-associated protein-43 (GAP-43), and acetylated-tubulin (ac-TUBA). In addition, an increased immunostaining for GAP-43 and ac-TUBA was observed in PC12 cells overexpressing Brn-3b, which was accompanied by a marked increase in neurite outgrowth, compared to PC12 cells overexpressing the empty vector. In separate experiments, one set of PC12 cells transfected either with a Brn-3b expression vector or an empty vector was subjected to conditions of hypoxia for 2 h, while another set of similarly transfected PC12 cells was maintained in normoxic conditions. It was found that the upregulation of GAP-43 and ac-TUBA in PC12 cells overexpressing Brn-3b under conditions of normoxia was sustained under conditions of hypoxia. Immunocytochemical analysis revealed not only an upregulation of GAP-43 and ac-TUBA, but also increased neurite outgrowth in PC12 cells transfected with Brn-3b as compared to PC12 cells transfected with empty vector in both normoxia and hypoxia. The findings have implications for a potential role of Brn-3b in neurodegenerative diseases in which hypoxia/ischemia contribute to pathophysiology of the disease.

  14. Establishment of a long-term spiral ganglion neuron culture with reduced glial cell number: Effects of AraC on cell composition and neurons.

    Science.gov (United States)

    Schwieger, Jana; Esser, Karl-Heinz; Lenarz, Thomas; Scheper, Verena

    2016-08-01

    Sensorineural deafness is mainly caused by damage to hair cells and degeneration of the spiral ganglion neurons (SGN). Cochlear implants can functionally replace lost hair cells and stimulate the SGN electrically. The benefit from cochlear implantation depends on the number and excitability of these neurons. To identify potential therapies for SGN protection, in vitro tests are carried out on spiral ganglion cells (SGC). A glial cell-reduced and neuron-enhanced culture of neonatal rat SGC under mitotic inhibition (cytarabine (AraC)) for up to seven days is presented. Serum containing and neurotrophin-enriched cultures with and without AraC-addition were analyzed after 4 and 7 days. The total number of cells was significantly reduced, while the proportion of neurons was greatly increased by AraC-treatment. Cell type-specific labeling demonstrated that nearly all fibroblasts and most of the glial cells were removed. Neither the neuronal survival, nor the neurite outgrowth or soma diameter were negatively affected. Additionally neurites remain partly free of surrounding non-neuronal cells. Recent culture conditions allow only for short-term cultivation of neonatal SGC and lack information on the influence of non-neuronal cells on SGN and of direct contact of neurites with test-materials. AraC-addition reduces the number of non-neuronal cells and increases the ratio of SGN in culture, without negative impact on neuronal viability. This treatment allows longer-term cultivation of SGC and provides deeper insight into SGN-glial cell interaction and the attachment of neurites on test-material surfaces. Copyright © 2016 The Authors. Published by Elsevier B.V. All rights reserved.

  15. An alternative pathway for signal flow from rod photoreceptors to ganglion cells in mammalian retina.

    OpenAIRE

    DeVries, S H; Baylor, D A

    1995-01-01

    Rod signals in the mammalian retina are thought to reach ganglion cells over the circuit rod-->rod depolarizing bipolar cell-->AII amacrine cell-->cone bipolar cells-->ganglion cells. A possible alternative pathway involves gap junctions linking the rods and cones, the circuit being rod-->cone-->cone bipolar cells-->ganglion cells. It is not clear whether this second pathway indeed relays rod signals to ganglion cells. We studied signal flow in the isolated rabbit retina with a multielectrode...

  16. Prostaglandin E2 facilitates neurite outgrowth in a motor neuron-like cell line, NSC-34

    Directory of Open Access Journals (Sweden)

    Hiroshi Nango

    2017-10-01

    Full Text Available Prostaglandin E2 (PGE2 exerts various biological effects by binding to E-prostanoid receptors (EP1-4. Although recent studies have shown that PGE2 induces cell differentiation in some neuronal cells such as mouse DRG neurons and sensory neuron-like ND7/23 cells, it is unclear whether PGE2 plays a role in differentiation of motor neurons. In the present study, we investigated the mechanism of PGE2-induced differentiation of motor neurons using NSC-34, a mouse motor neuron-like cell line. Exposure of undifferentiated NSC-34 cells to PGE2 and butaprost, an EP2-selective agonist, resulted in a reduction of MTT reduction activity without increase the number of propidium iodide-positive cells and in an increase in the number of neurite-bearing cells. Sulprostone, an EP1/3 agonist, also significantly lowered MTT reduction activity by 20%; however, no increase in the number of neurite-bearing cells was observed within the concentration range tested. PGE2-induced neurite outgrowth was attenuated significantly in the presence of PF-0441848, an EP2-selective antagonist. Treatment of these cells with dibutyryl-cAMP increased the number of neurite-bearing cells with no effect on cell proliferation. These results suggest that PGE2 promotes neurite outgrowth and suppresses cell proliferation by activating the EP2 subtype, and that the cAMP-signaling pathway is involved in PGE2-induced differentiation of NSC-34 cells. Keywords: Prostaglandin E2, E-prostanoid receptors, Motor neuron, Neurite outgrowth, cAMP

  17. Dopaminergic modulation of tracer coupling in a ganglion-amacrine cell network

    OpenAIRE

    MILLS, STEPHEN L.; XIA, XIAO-BO; HOSHI, HIDEO; FIRTH, SALLY I.; RICE, MARGARET E.; FRISHMAN, LAURA J.; MARSHAK, DAVID W.

    2007-01-01

    Many retinal ganglion cells are coupled via gap junctions with neighboring amacrine cells and ganglion cells. We investigated the extent and dynamics of coupling in one such network, the OFF α ganglion cell of rabbit retina and its associated amacrine cells. We also observed the relative spread of Neurobiotin injected into a ganglion cell in the presence of modulators of gap junctional permeability. We found that gap junctions between amacrine cells were closed via stimulation of a D1 dopamin...

  18. A model for the receptive field of retinal ganglion cells.

    Science.gov (United States)

    Cho, Myoung Won; Choi, M Y

    2014-01-01

    Most retina ganglion cells have center-surround receptive fields, where the center may be either ON or OFF while the surround is the opposite. We clarify the functional roles of the receptive field structure, on the basis of the modern theory of natural data processing. It is suggested that the retina shares the principal mechanism and performance of image processing with a video codec in computers, where the antagonism in spatial or temporal receptive fields originates from the orthogonality condition between linear filters for optimal coding of visual signals. We also reveal what visual information is multiplexed across the discharges of an ensemble of ganglion cells. Our theory makes it possible to predict the cross-correlations between ganglion cell spikes, which are optimized for LGN cells to respond accurately and quickly to their receptive fields. Copyright © 2013 Elsevier Ltd. All rights reserved.

  19. Staurosporine induces ganglion cell differentiation in part by stimulating urokinase-type plasminogen activator expression and activation in the developing chick retina

    Energy Technology Data Exchange (ETDEWEB)

    Kim, Yeoun-Hee [Department of Biology, College of Natural Sciences, Kyungpook National University, Daegu 702-701 (Korea, Republic of); Chang, Yongmin [Department of Molecular Medicine, Kyungpook National University College of Medicine, Kyungpook National University, 200 Dongduk-Ro Jung-Gu, Daegu 700-714 (Korea, Republic of); Jung, Jae-Chang, E-mail: jcjung@knu.ac.kr [Department of Biology, College of Natural Sciences, Kyungpook National University, Daegu 702-701 (Korea, Republic of)

    2012-06-22

    Highlights: Black-Right-Pointing-Pointer Staurosporine mediates stimulation of RGC differentiation in vitro cultured retinal neuroblasts. Black-Right-Pointing-Pointer Staurosporine mediates uPA activation during RGC differentiation in vitro. Black-Right-Pointing-Pointer Inhibition of uPA blocks the staurosporine mediated RGC differentiation both in vitro and in ovo. Black-Right-Pointing-Pointer Thus, uPA may play a role in the staurosporine-mediated stimulation of RGC differentiation. -- Abstract: Here, we investigated whether staurosporine-mediated urokinase-type plasminogen activator (uPA) activation is involved in retinal ganglion cell (RGC) differentiation. Retinal cells were isolated from developing chick retinas at embryonic day 6 (E6). Relatively few control cells grown in serum-free medium started to form processes by 12 h. In contrast, staurosporine-treated cells had processes within 3 h, and processes were evident at 8 h. Immunofluorescence staining showed that Tuj-1-positive cells with shorter neurites could be detected in control cultures at 18 h, whereas numerous Tuj-1 positive ganglion cells with longer neuritic extensions were seen in staurosporine-treated cultures. BrdU-positive proliferating cells were more numerous in control cultures than in staurosporine-treated cultures, and the BrdU staining was not detected in post-mitotic Tuj-1 positive ganglion cells. Western blotting of cell lysates showed that staurosporine induced high levels of the active form of uPA. The staurosporine-induced uPA signal was localized predominantly in the soma, neurites and axons of Tuj-1-positive ganglion cells. Amiloride, an inhibitor of uPA, markedly reduced staurosporine-induced Tuj-1 staining, neurite length, neurite number, and uPA staining versus controls. In developing retinas in ovo, amiloride administration remarkably reduced the staurosporine-induced uPA staining and RGC differentiation. Taken together, our in vitro and in vivo data collectively indicate that

  20. Neurite outgrowth resistance to rho kinase inhibitors in PC12 Adh cell.

    Science.gov (United States)

    Yin, Hua; Hou, Xiaolin; Tao, Tingrui; Lv, Xiaoman; Zhang, Luyong; Duan, Weigang

    2015-05-01

    Rho kinase (ROCK) inhibitor is a promising agent for neural injury disorders, which mechanism is associated with neurite outgrowth. However, neurite outgrowth resistance occurred when PC12 Adh cell was treated with ROCK inhibitors for a longer time. PC12 Adh cells were treated with ROCK inhibitor Y27632 or NGF for different durations. Neurite outgrowth resistance occurred when PC12 Adh cell exposed to Y27632 (33 µM) for 3 or more days, but not happen when exposed to nerve growth factor (NGF, 100 ng/mL). The gene expression in the PC12 Adh cells treated with Y27632 (33 µM) or NGF (100 ng/mL) for 2 or 4 days was assayed by gene microarray, and the reliability of the results were confirmed by real-time RT-PCR. Cluster analysis proved that the gene expression profile of PC12 Adh cell treated with Y27632 for 4 days was different from that treated with Y27632 for 2 days and those treated with NGF for 2 and 4 days, respectively. Pathway analysis hinted that the neurite outgrowth resistance could be associated with up-regulation of inflammatory pathways, especially rno04610 (complement and coagulation cascades), and down-regulation of cell cycle pathways, especially rno04110. © 2015 The Authors. Cell Biology International Published by John Wiley & Sons Ltd on behalf of International Federation of Cell Biology.

  1. ALS/FTLD-linked TDP-43 regulates neurite morphology and cell survival in differentiated neurons

    International Nuclear Information System (INIS)

    Han, Jeong-Ho; Yu, Tae-Hoon; Ryu, Hyun-Hee; Jun, Mi-Hee; Ban, Byung-Kwan; Jang, Deok-Jin; Lee, Jin-A

    2013-01-01

    Tar-DNA binding protein of 43 kDa (TDP-43) has been characterized as a major component of protein aggregates in brains with neurodegenerative diseases such as frontotemporal lobar degeneration (FTLD) and amyotrophic lateral sclerosis (ALS). However, physiological roles of TDP-43 and early cellular pathogenic effects caused by disease associated mutations in differentiated neurons are still largely unknown. Here, we investigated the physiological roles of TDP-43 and the effects of missense mutations associated with diseases in differentiated cortical neurons. The reduction of TDP-43 by siRNA increased abnormal neurites and decreased cell viability. ALS/FTLD-associated missense mutant proteins (A315T, Q331K, and M337V) were partially mislocalized to the cytosol and neurites when compared to wild-type and showed abnormal neurites similar to those observed in cases of loss of TDP-43. Interestingly, cytosolic expression of wild-type TDP-43 with mutated nuclear localization signals also induced abnormal neurtie morphology and reduction of cell viability. However, there was no significant difference in the effects of cytosolic expression in neuronal morphology and cell toxicity between wild-type and missense mutant proteins. Thus, our results suggest that mislocalization of missense mutant TDP-43 may contribute to loss of TDP-43 function and affect neuronal morphology, probably via dominant negative action before severe neurodegeneration in differentiated cortical neurons. Highlights: • The function of nuclear TDP-43 in neurite morphology in mature neurons. • Partial mislocalization of TDP-43 missense mutants into cytosol from nucleus. • Abnormal neurite morphology caused by missense mutants of TDP-43. • The effect of cytosolic expression of TDP-43 in neurite morphology and in cell survival

  2. Development of rat embryonic spinal ganglion cells in damaged nerve.

    Science.gov (United States)

    Petrova, E S; Isaeva, E N; Korzhevskii, D E

    2014-09-01

    The development of dissociated cells from rat embryonic spinal ganglion after transplantation to damaged nerve of adult animals was studied using immunohistochemical differentiation markers of neural and glial cells. The cell suspension obtained after dissociation of rat embryonic spinal ganglia (embryonic day 15) was injected into the proximal segment of crushed sciatic nerve. The nerve was damaged by ligation for 40 sec. Progenitor cells were labeled with 5-bromo-2'-deoxyuridine (BrdU) before transplantation. BrdU-immunopositive cells were detected in the nerve trunks of recipients on days 1, 21, and 28 after transplantation. Dissociated cells of rat embryonic spinal ganglion (embryonic day 15) survived for at least 4 weeks after transplantation to the nerve and differentiate into NeuN-immunopositive neurons with morphological properties of sensory neurons and satellite cells containing S100 protein.

  3. A Comparative Analysis of Ganglion Cell Complex Parameters in ...

    African Journals Online (AJOL)

    Aim: To evaluate the differences between ganglion cell complex (GCC) of primary open angle glaucoma and ocular disorders affecting the macula. Methods and Materials: Forty-seven patients diagnosed with primary open angle glaucoma and 27 patients with macular diseases of different aetiology were enrolled in this ...

  4. Processing of natural temporal stimuli by macaque retinal ganglion cells

    NARCIS (Netherlands)

    Hateren, J.H. van; Rüttiger, L.; Lee, B.B.

    2002-01-01

    This study quantifies the performance of primate retinal ganglion cells in response to natural stimuli. Stimuli were confined to the temporal and chromatic domains and were derived from two contrasting environments, one typically northern European and the other a flower show. The performance of the

  5. Encoding visual information in retinal ganglion cells with prosthetic stimulation

    Science.gov (United States)

    Freeman, Daniel K.; Rizzo, Joseph F., III; Fried, Shelley I.

    2011-06-01

    Retinal prostheses aim to restore functional vision to those blinded by outer retinal diseases using electric stimulation of surviving retinal neurons. The ability to replicate the spatiotemporal pattern of ganglion cell spike trains present under normal viewing conditions is presumably an important factor for restoring high-quality vision. In order to replicate such activity with a retinal prosthesis, it is important to consider both how visual information is encoded in ganglion cell spike trains, and how retinal neurons respond to electric stimulation. The goal of the current review is to bring together these two concepts in order to guide the development of more effective stimulation strategies. We review the experiments to date that have studied how retinal neurons respond to electric stimulation and discuss these findings in the context of known retinal signaling strategies. The results from such in vitro studies reveal the advantages and disadvantages of activating the ganglion cell directly with the electric stimulus (direct activation) as compared to activation of neurons that are presynaptic to the ganglion cell (indirect activation). While direct activation allows high temporal but low spatial resolution, indirect activation yields improved spatial resolution but poor temporal resolution. Finally, we use knowledge gained from in vitro experiments to infer the patterns of elicited activity in ongoing human trials, providing insights into some of the factors limiting the quality of prosthetic vision.

  6. Thorny ganglion cells in marmoset retina: Morphological and neurochemical characterization with antibodies against calretinin.

    Science.gov (United States)

    Chandra, Ashleigh J; Lee, Sammy C S; Grünert, Ulrike

    2017-12-15

    In primates, over 17 morphological types of retinal ganglion cell have been distinguished by their dendritic morphology and stratification, but reliable markers for specific ganglion cell populations are still rare. The calcium binding protein calretinin is known to be expressed in the inner nuclear and the ganglion cell layer of marmoset retina, however, the specific cell type(s) expressing calretinin in the ganglion cell layer are yet to be determined. Here, we identified calretinin positive retinal ganglion cells in the common marmoset Callithrix jacchus. Double labeling with the ganglion cell marker RBPMS demonstrated that the large majority (80%) of the calretinin positive cells in the ganglion cell layer are ganglion cells, and 20% are displaced amacrine cells. The calretinin positive ganglion cells made up on average 12% of the total ganglion cell population outside of the foveal region and their proportion increased with eccentricity. Prelabeling with antibodies against calretinin and subsequent intracellular injection with DiI revealed that the large majority of the injected cells (n = 74) were either narrow thorny or broad thorny ganglion cells, 14 cells were displaced amacrine cells. Narrow thorny cells were further distinguished into outer and inner stratifying cells. In addition, weakly labeled cells with a large soma were identified as parasol ganglion cells. Our results show that three types of thorny ganglion cells in marmoset retina can be identified with antibodies against calretinin. Our findings are also consistent with the idea that the proportion of wide-field ganglion cell types increases in peripheral retina. © 2017 Wiley Periodicals, Inc.

  7. Starburst cells nondirectionally facilitate the responses of direction-selective retinal ganglion cells.

    Science.gov (United States)

    Chiao, Chuan-Chin; Masland, Richard H

    2002-12-15

    The mechanism of direction selectivity in retinal ganglion cells remains controversial. An important issue is how the starburst amacrine cells, which are known to provide a major synaptic input to the direction-selective ganglion cells, participate in the directional discrimination. Here, we present evidence that the cholinergic outputs of the starburst cells affect the responses of the ganglion cells symmetrically; they provide a feedforward excitation that facilitates the response of the ganglion cells to movement in both the preferred and null directions. This seems to place a constraint on models of the directional discrimination in which the starburst cells participate, namely, that their cholinergic synapses be nondirectional in their effects on the ganglion cells.

  8. Veratridine increases the survival of retinal ganglion cells in vitro

    Directory of Open Access Journals (Sweden)

    S.P.F. Pereira

    1997-12-01

    Full Text Available Neuronal cell death is an important phenomenon involving many biochemical pathways. This degenerative event has been studied to understand how the cells activate the mechanisms that lead to self-destruction. Target cells and afferent cells play a relevant role in the regulation of natural cell death. We studied the effect of veratridine (1.5, 3.0, 4.5 and 6.0 µM on the survival of neonatal rat retinal ganglion cells in vitro. Veratridine (3.0 µM, a well-known depolarizing agent that opens the Na+ channel, promoted a two-fold increase in the survival of retinal ganglion cells kept in culture for 48 h. This effect was dose-dependent and was blocked by 1.0 µM tetrodotoxin (a classical voltage-dependent Na+ channel blocker and 30.0 µM flunarizine (a Na+ and Ca2+ channel blocker. These results indicate that electrical activity is also important for the maintenance of retinal ganglion cell survival in vitro

  9. Luteolin Induces microRNA-132 Expression and Modulates Neurite Outgrowth in PC12 Cells

    Science.gov (United States)

    Wu, Ming-Jiuan; Chen, Pei-Yi; Yen, Jui-Hung

    2012-01-01

    Luteolin (3′,4′,5,7-tetrahydroxyflavone), a food-derived flavonoid, has been reported to exert neurotrophic properties that are associated with its capacity to promote neuronal survival and neurite outgrowth. In this study, we report for the first time that luteolin induces the persistent expression of microRNA-132 (miR-132) in PC12 cells. The correlation between miR-132 knockdown and a decrease in luteolin-mediated neurite outgrowth may indicate a mechanistic link by which miR-132 functions as a mediator for neuritogenesis. Furthermore, we find that luteolin led to the phosphorylation and activation of cAMP response element binding protein (CREB), which is associated with the up-regulation of miR-132 and neurite outgrowth. Moreover, luteolin-induced CREB activation, miR-132 expression and neurite outgrowth were inhibited by adenylate cyclase, protein kinase A (PKA) and MAPK/ERK kinase 1/2 (MEK1/2) inhibitors but not by protein kinase C (PKC) or calcium/calmodulin-dependent protein kinase II (CaMK II) inhibitors. Consistently, we find that luteolin treatment increases ERK phosphorylation and PKA activity in PC12 cells. These results show that luteolin induces the up-regulation of miR-132, which serves as an important regulator for neurotrophic actions, mainly acting through the activation of cAMP/PKA- and ERK-dependent CREB signaling pathways in PC12 cells. PMID:22916239

  10. Agmatine protects retinal ganglion cells from hypoxia-induced apoptosis in transformed rat retinal ganglion cell line

    Directory of Open Access Journals (Sweden)

    Kim Chan

    2007-10-01

    Full Text Available Abstract Background Agmatine is an endogenous polyamine formed by the decarboxylation of L-arginine. We investigated the protective effects of agmatine against hypoxia-induced apoptosis of immortalized rat retinal ganglion cells (RGC-5. RGC-5 cells were cultured in a closed hypoxic chamber (5% O2 with or without agmatine. Cell viability was determined by lactate dehydrogenase (LDH assay and apoptosis was examined by annexin V and caspase-3 assays. Expression and phosphorylation of mitogen-activated protein kinases (MAPKs; JNK, ERK p44/42, and p38 and nuclear factor-kappa B (NF-κB were investigated by Western immunoblot analysis. The effects of agmatine were compared to those of brain-derived neurotrophic factor (BDNF, a well-known protective neurotrophin for retinal ganglion cells. Results After 48 hours of hypoxic culture, the LDH assay showed 52.3% cell loss, which was reduced to 25.6% and 30.1% when agmatine and BDNF were administered, respectively. This observed cell loss was due to apoptotic cell death, as established by annexin V and caspase-3 assays. Although total expression of MAPKs and NF-κB was not influenced by hypoxic injury, phosphorylation of these two proteins was increased. Agmatine reduced phosphorylation of JNK and NF-κB, while BDNF suppressed phosphorylation of ERK and p38. Conclusion Our results show that agmatine has neuroprotective effects against hypoxia-induced retinal ganglion cell damage in RGC-5 cells and that its effects may act through the JNK and NF-κB signaling pathways. Our data suggest that agmatine may lead to a novel therapeutic strategy to reduce retinal ganglion cell injury related to hypoxia.

  11. CD146- and CD105-positive phenotypes of retinal ganglion cells. Are these in situ proofs of neuronal regeneration?

    Science.gov (United States)

    Vrapciu, A D; Rusu, M C; Voinea, L M; Corbu, C G

    2014-10-01

    The in vivo identity of stem cells is not yet clear. Numerous studies involve the perivascular niches as providers of stem cells during regenerative processes. CD146, in humans, as well as gicerin, at chicken, play roles in neuronal development and neurites extension. CD146 is a marker of stemness but also a pericytary marker. Stem cells in vascular niches can differentiate in neural cells. By applying CD146 and CD105 antibodies on human retinas from glaucomatous eyes, CD146-positive retinal ganglion cells (RGCs) were found, some being placed in perivascular positions; ongoing processes of neurites extension were related to these neurons. On other hand, RGCs were positively labeled by CD105 antibodies. These results support the hypothesis that in glaucoma eyes the CD146-positive RGCs result from regenerative processes driven by stem cells in the retinal perivascular niches. Further experiments are needed to evaluate whether CR146-positive neurons indicate also a physiological process of maintenance of retina. Copyright © 2014 Elsevier Ltd. All rights reserved.

  12. Retinal Ganglion Cell Loss in Diabetes Associated with Elevated Homocysteine

    Directory of Open Access Journals (Sweden)

    Kenneth S. Shindler

    2009-11-01

    Full Text Available A number of studies have suggested that homocysteine may be a contributing factor to development of retinopathy in diabetic patients based on observed correlations between elevated homocysteine levels and the presence of retinopathy. The significance of such a correlation remains to be determined, and potential mechanisms by which homocysteine might induce retinopathy have not been well characterized. Ganapathy and colleagues1 used mutant mice that have endogenously elevated homocysteine levels due to heterozygous deletion of the cystathionine-β-synthase gene to examine changes in retinal pathology following induction of diabetes. Their finding that elevated homocysteine levels hastens loss of cells in the retinal ganglion cell layer suggests that toxicity to ganglion cells may warrant further investigation as a potential mechanism of homocysteine enhanced susceptibility to diabetic retinopathy.

  13. High speed coding for velocity by archerfish retinal ganglion cells

    Directory of Open Access Journals (Sweden)

    Kretschmer Viola

    2012-06-01

    Full Text Available Abstract Background Archerfish show very short behavioural latencies in response to falling prey. This raises the question, which response parameters of retinal ganglion cells to moving stimuli are best suited for fast coding of stimulus speed and direction. Results We compared stimulus reconstruction quality based on the ganglion cell response parameters latency, first interspike interval, and rate. For stimulus reconstruction of moving stimuli using latency was superior to using the other stimulus parameters. This was true for absolute latency, with respect to stimulus onset, as well as for relative latency, with respect to population response onset. Iteratively increasing the number of cells used for reconstruction decreased the calculated error close to zero. Conclusions Latency is the fastest response parameter available to the brain. Therefore, latency coding is best suited for high speed coding of moving objects. The quantitative data of this study are in good accordance with previously published behavioural response latencies.

  14. Large enhancement in neurite outgrowth on a cell membrane-mimicking conducting polymer

    Science.gov (United States)

    Zhu, Bo; Luo, Shyh-Chyang; Zhao, Haichao; Lin, Hsing-An; Sekine, Jun; Nakao, Aiko; Chen, Chi; Yamashita, Yoshiro; Yu, Hsiao-Hua

    2014-07-01

    Although electrically stimulated neurite outgrowth on bioelectronic devices is a promising means of nerve regeneration, immunogenic scar formation can insulate electrodes from targeted cells and tissues, thereby reducing the lifetime of the device. Ideally, an electrode material capable of electrically interfacing with neurons selectively and efficiently would be integrated without being recognized by the immune system and minimize its response. Here we develop a cell membrane-mimicking conducting polymer possessing several attractive features. This polymer displays high resistance towards nonspecific enzyme/cell binding and recognizes targeted cells specifically to allow intimate electrical communication over long periods of time. Its low electrical impedance relays electrical signals efficiently. This material is capable to integrate biochemical and electrical stimulation to promote neural cellular behaviour. Neurite outgrowth is enhanced greatly on this new conducting polymer; in addition, electrically stimulated secretion of proteins from primary Schwann cells can also occur on it.

  15. Age-Related Change in Vestibular Ganglion Cell Populations in Individuals With Presbycusis and Normal Hearing.

    Science.gov (United States)

    Gluth, Michael B; Nelson, Erik G

    2017-04-01

    We sought to establish that the decline of vestibular ganglion cell counts uniquely correlates with spiral ganglion cell counts, cochlear hair cell counts, and hearing phenotype in individuals with presbycusis. The relationship between aging in the vestibular system and aging in the cochlea is a topic of ongoing investigation. Histopathologic age-related changes the vestibular system may mirror what is seen in the cochlea, but correlations with hearing phenotype and the impact of presbycusis are not well understood. Vestibular ganglion cells, spiral ganglion cells, and cochlear hair cells were counted in specimens from individuals with presbycusis and normal hearing. These were taken from within a large collection of processed human temporal bones. Correlations between histopathology and hearing phenotype were investigated. Vestibular ganglion cell counts were positively correlated with spiral ganglion cell counts and cochlear hair cell counts and were negatively correlated with hearing phenotype. There was no statistical evidence on linear regression to suggest that the relationship between age and cell populations differed significantly according to whether presbycusis was present or not. Superior vestibular ganglion cells were more negatively correlated with age than inferior ganglion cells. No difference in vestibular ganglion cells was noted based on sex. Vestibular ganglion cell counts progressively deteriorate with age, and this loss correlates closely with changes in the cochlea, as well as hearing phenotype. However, these correlations do not appear to be unique in individuals with presbycusis as compared with those with normal hearing.

  16. Intrinsically photosensitive retinal ganglion cell function in relation to age

    DEFF Research Database (Denmark)

    Herbst, Kristina; Sander, Birgit; Lund-Andersen, Henrik

    2012-01-01

    The activity of melanopsin containing intrinsically photosensitive ganglion retinal cells (ipRGC) can be assessed by a means of pupil responses to bright blue (appr.480 nm) light. Due to age related factors in the eye, particularly, structural changes of the lens, less light reaches retina. The aim...... of this study was to examine how age and in vivo measured lens transmission of blue light might affect pupil light responses, in particular, mediated by the ipRGC....

  17. Pituitary adenylate cyclase-activating peptide stimulates neurite growth in PC12 cells.

    Science.gov (United States)

    Hernandez, A; Kimball, B; Romanchuk, G; Mulholland, M W

    1995-01-01

    The ability of PACAP-38 to stimulate morphological development was studied using rat pheochromocytoma PC12 cells. PACAP-38 produced concentration-dependent increases in percentage of cells exhibiting neurite extension. Similar increases were produced by forskolin (28 +/- 2% at 96 h) and 8-bromo cAMP (30 +/- 2%). Vasoactive intestinal peptide and alpha-calcitonin gene-related peptide were without effect. PACAP-38 produced significant increases in PC12 cell cAMP content and inositol phosphate turnover. Intracellular [Ca2+] increased from 169 +/- 14 nM to 560 +/- 58 nM in response to 1 microM PACAP-38. PACAP-stimulated neurite outgrowth was abolished by RpcAMPS, an inhibitor of cAMP-dependent kinases but was unaffected by the protein kinase C antagonist H7.

  18. Differential intensity-dependent effects of magnetic stimulation on the longest neurites and shorter dendrites in neuroscreen-1 cells

    Science.gov (United States)

    Lin, Ching-Yi; Huang, Whitney J.; Li, Kevin; Swanson, Roy; Cheung, Brian; Lin, Vernon W.; Lee, Yu-Shang

    2015-04-01

    Objective. Magnetic stimulation (MS) is a potential treatment for neuropsychiatric disorders. This study investigates whether MS-regulated neuronal activity can translate to specific changes in neuronal arborization and thus regulate synaptic activity and function. Approach. To test our hypotheses, we examined the effects of MS on neurite growth of neuroscreen-1 (NS-1) cells over the pulse frequencies of 1, 5 and 10 Hz at field intensities controlled via machine output (MO). Cells were treated with either 30% or 40% MO. Due to the nature of circular MS coils, the center region of the gridded coverslip (zone 1) received minimal (∼5%) electromagnetic current density while the remaining area (zone 2) received maximal (∼95%) current density. Plated NS-1 cells were exposed to MS twice per day for three days and then evaluated for length and number of neurites and expression of brain-derived neurotrophic factor (BDNF). Main results. We show that MS dramatically affects the growth of the longest neurites (axon-like) but does not significantly affect the growth of shorter neurites (dendrite-like). Also, MS-induced changes in the longest neurite growth were most evident in zone 1, but not in zone 2. MS effects were intensity-dependent and were most evident in bolstering longest neurite outgrowth, best seen in the 10 Hz MS group. Furthermore, we found that MS-increased BDNF expression and secretion was also frequency-dependent. Taken together, our results show that MS exerts distinct effects when different frequencies and intensities are applied to the neuritic compartments (longest neurite versus shorter dendrite(s)) of NS-1 cells. Significance. These findings support the concept that MS increases BDNF expression and signaling, which sculpts longest neurite arborization and connectivity by which neuronal activity is regulated. Understanding the mechanisms underlying MS is crucial for efficiently incorporating its use into potential therapeutic strategies.

  19. Neurite outgrowth on chromaffin cells applying extremely low frequency magnetic fields by permanent magnets.

    Science.gov (United States)

    Hernández-Hernández, Hugo; Cruces-Solis, Hugo; Elías-Viñas, David; Verdugo-Díaz, Leticia

    2009-10-01

    There is an increasing interest about the effects of electromagnetic fields on health and clinical applications. Electromagnetic fields have been shown to promote differentiation and regeneration of many tissues. The purpose of the present study was to evaluate if a magnetic field (MF) varying in time is able to induce neurite outgrowth in cultured chromaffin cells. For this reason, a stimulation system was developed in order to generate a magnetic field, using permanent magnets as a supply. In this investigation we used a pair of permanent ferrite magnets. These were mounted in a mechanical system in which both magnets rotate around a culture Petri dish. The stimulation device was designed at Centro de Investigación y de Estudios Avanzados, Avanzados del IPN, Mexico City. Primary cultures of chromaffin cells were stimulated with a magnetic field of 6.4 mT and 4, 7, 10 or 12Hz (2h daily, during a 7-day period). After treatment, percentage of neurite outgrowth was calculated. Our results show that the magnetic fields produced by rotating permanent magnets induced neurite outgrowth on chromaffin cells at 7 and 10Hz. The present study provides evidence that MFs varying in time (7 and 10Hz) induce neurite outgrowth in chromaffin cells. These studies will contribute to elucidate the effect of noninvasive MF stimulus in order to apply it in future regeneration therapies. Also, the device designed could be used for different kind of cells and may work as a model for future clinical devices. 2009 IMSS. Published by Elsevier Inc.

  20. Quantitative assessment of neurite outgrowth in human embryonic stem-cell derived neurons using automated high-content image analysis

    Science.gov (United States)

    During development neurons undergo a number of morphological changes including neurite outgrowth from the cell body. Exposure to neurotoxicants that interfere with this process may cause in permanent deficits in nervous system function. While many studies have used rodent primary...

  1. Cometin is a novel neurotrophic factor that promotes neurite outgrowth and neuroblast migration in vitro and supports survival of spiral ganglion neurons in vivo

    DEFF Research Database (Denmark)

    Jørgensen, Jesper Roland; Fransson, Anette; Fjord-Larsen, Lone

    2012-01-01

    properties in vitro, combined with the restricted inner ear expression during development, we further investigated Cometin in relation to deafness. In neomycin deafened guinea pigs, two weeks intracochlear infusion of recombinant Cometin supports spiral ganglion neuron survival and function. In contrast...... to the control group receiving artificial perilymph, Cometin treated animals retain normal electrically-evoked brainstem response which is maintained several weeks after treatment cessation. Neuroprotection is also evident from stereological analysis of the spiral ganglion. Altogether, these studies show...

  2. Pulsed electromagnetic fields potentiate neurite outgrowth in the dopaminergic MN9D cell line.

    Science.gov (United States)

    Lekhraj, Rukmani; Cynamon, Deborah E; DeLuca, Stephanie E; Taub, Eric S; Pilla, Arthur A; Casper, Diana

    2014-06-01

    Pulsed electromagnetic fields (PEMF) exert biological effects and are in clinical use to facilitate bone repair and wound healing. Research has demonstrated that PEMF can induce signaling molecules and growth factors, molecules that play important roles in neuronal differentiation. Here, we tested the effects of a low-amplitude, nonthermal, pulsed radiofrequency signal on morphological neuronal differentiation in MN9D, a dopaminergic cell line. Cells were plated in medium with 10% fetal calf serum. After 1 day, medium was replaced with serum-containing medium, serum-free medium, or medium supplemented with dibutyryl cyclic adenosine monophosphate (Bt2 cAMP), a cAMP analog known to induce neurite outgrowth. Cultures were divided into groups and treated with PEMF signals for either 30 min per day or continuously for 15 min every hour for 3 days. Both serum withdrawal and Bt2 cAMP significantly increased neurite length. PEMF treatment similarly increased neurite length under both serum-free and serum-supplemented conditions, although to a lesser degree in the presence of serum, when continuous treatments had greater effects. PEMF signals also increased cell body width, indicating neuronal maturation, and decreased protein content, suggesting that this treatment was antimitotic, an effect reversed by the inhibitor of cAMP formation dideoxyadenosine. Bt2 cAMP and PEMF effects were not additive, suggesting that neurite elongation was achieved through a common pathway. PEMF signals increased cAMP levels from 3 to 5 hr after treatment, supporting this mechanism of action. Although neuritogenesis is considered a developmental process, it may also represent the plasticity required to form and maintain synaptic connections throughout life. Copyright © 2014 Wiley Periodicals, Inc.

  3. Melanopsin-expressing retinal ganglion cells: implications for human diseases

    DEFF Research Database (Denmark)

    La Morgia, Chiara; Ross-Cisneros, Fred N; Hannibal, Jens

    2011-01-01

    interest on these cells, mainly focused on animal models. Only recently, a few studies have started to address the relevance of the mRGC system in humans and related diseases. We recently discovered that mRGCs resist neurodegeneration in two inherited mitochondrial disorders that cause blindness, i......In the last decade, there was the seminal discovery of melanopsin-expressing retinal ganglion cells (mRGCs) as a new class of photoreceptors that subserve the photoentrainment of circadian rhythms and other non-image forming functions of the eye. Since then, there has been a growing research...

  4. Melanopsin expressing human retinal ganglion cells

    DEFF Research Database (Denmark)

    Hannibal, Jens; Christiansen, Anders Tolstrup; Heegaard, Steffen

    2017-01-01

    microscopy and 3D reconstruction of melanopsin immunoreactive (-ir) RGCs, we applied the criteria used in mouse on human melanopsin-ir RGCs. We identified M1, displaced M1, M2, and M4 cells. We found two other subtypes of melanopsin-ir RGCs, which were named "gigantic M1 (GM1)" and "gigantic displaced M1...

  5. Sympathetic and sensory innervation of small intensely fluorescent (SIF) cells in rat superior cervical ganglion.

    Science.gov (United States)

    Takaki, Fumiya; Nakamuta, Nobuaki; Kusakabe, Tatsumi; Yamamoto, Yoshio

    2015-02-01

    The sympathetic ganglion contains small intensely fluorescent (SIF) cells derived from the neural crest. We morphologically characterize SIF cells and focus on their relationship with ganglionic cells, preganglionic nerve fibers and sensory nerve endings. SIF cells stained intensely for tyrosine hydroxylase (TH), with a few cells also being immunoreactive for dopamine β-hydroxylase (DBH). Vesicular acetylcholine transporter (VAChT)-immunoreactive puncta were distributed around some clusters of SIF cells, whereas some SIF cells closely abutted DBH-immunoreactive ganglionic cells. SIF cells contained bassoon-immunoreactive products beneath the cell membrane at the attachments and on opposite sites to the ganglionic cells. Ganglion neurons and SIF cells were immunoreactive to dopamine D2 receptors. Immunohistochemistry for P2X3 revealed ramified nerve endings with P2X3 immunoreactivity around SIF cells. Triple-labeling for P2X3, TH and VAChT allowed the classification of SIF cells into three types based on their innervation: (1) with only VAChT-immunoreactive puncta, (2) with only P2X3-immunoreactive nerve endings, (3) with both P2X3-immunoreactive nerve endings and VAChT-immunoreactive puncta. The results of retrograde tracing with fast blue dye indicated that most of these nerve endings originated from the petrosal ganglion. Thus, SIF cells in the superior cervical ganglion are innervated by preganglionic fibers and glossopharyngeal sensory nerve endings and can be classified into three types. SIF cells might modulate sympathetic activity in the superior cervical ganglion.

  6. Neurite outgrowth induced by a synthetic peptide ligand of neural cell adhesion molecule requires fibroblast growth factor receptor activation

    DEFF Research Database (Denmark)

    Rønn, L C; Doherty, P; Holm, A

    2000-01-01

    identified a neuritogenic ligand, termed the C3 peptide, of the first immunoglobulin (lg) module of NCAM using a combinatorial library of synthetic peptides. Here we investigate whether stimulation of neurite outgrowth by this synthetic ligand of NCAM involves FGFRs. In primary cultures of cerebellar neurons...... from wild-type mice, the C3 peptide stimulated neurite outgrowth. This response was virtually absent in cultures of cerebellar neurons from transgenic mice expressing a dominant-negative form of the FGFR1. Likewise, in PC12E2 cells transiently expressing a dominant-negative form of the mouse FGFR1......, induction of neurites by the C3 peptide was abrogated. These findings suggest that the neuritogenic effect of the C3 peptide requires the presence of functional FGFRs and support the hypothesis that FGFRs are essential in cell adhesion molecule-stimulated neurite outgrowth. The C3 peptide appears...

  7. Bone Marrow Stromal Cells Combined With a Honeycomb Collagen Sponge Facilitate Neurite Elongation In Vitro and Neural Restoration in the Hemisected Rat Spinal Cord.

    Science.gov (United States)

    Onuma-Ukegawa, Madoka; Bhatt, Kush; Hirai, Takashi; Kaburagi, Hidetoshi; Sotome, Shinichi; Wakabayashi, Yoshiaki; Ichinose, Shizuko; Shinomiya, Kenichi; Okawa, Atsushi; Enomoto, Mitsuhiro

    2015-01-01

    In the last decade, researchers and clinicians have reported that transplantation of bone marrow stromal cells (BMSCs) promotes functional recovery after brain or spinal cord injury (SCI). However, an appropriate scaffold designed for the injured spinal cord is needed to enhance the survival of transplanted BMSCs and to promote nerve regeneration. We previously tested a honeycomb collagen sponge (HC), which when applied to the transected spinal cord allowed bridging of the gap with nerve fibers. In this study, we examined whether the HC implant combined with rat BMSCs increases nerve regeneration in vitro and enhances functional recovery in vivo. We first evaluated the neurite outgrowth of rat dorsal root ganglion (DRG) explants cultured on HC with or without BMSCs in vitro. Regeneration of neurites from the DRGs was increased by BMSCs combined with HC scaffolds. In the in vivo study, 3-mm-long HC scaffolds with or without BMSCs were implanted into the hemisected rat thoracic spinal cord. Four weeks after the procedure, rats implanted with HC scaffolds containing BMSCs displayed better motor and sensory recovery than those implanted with HC scaffolds only. Histologically, more CGRP-positive sensory fibers at the implanted site and 5-HT-positive serotonergic fibers contralateral to the implanted site were observed in spinal cords receiving BMSCs. Furthermore, more rubrospinal neurons projected distally to the HC implant containing BMSCs. Our study indicates that the application of BMSCs in a HC scaffold in the injured spinal cord directly promoted sensory nerve and rubrospinal tract regeneration, thus resulting in functional recovery.

  8. Induction of Neurite Outgrowth in PC12 Cells Treated with Temperature-Controlled Repeated Thermal Stimulation

    Science.gov (United States)

    Kudo, Tada-aki; Kanetaka, Hiroyasu; Mochizuki, Kentaro; Tominami, Kanako; Nunome, Shoko; Abe, Genji; Kosukegawa, Hiroyuki; Abe, Toshihiko; Mori, Hitoshi; Mori, Kazumi; Takagi, Toshiyuki; Izumi, Shin-ichi

    2015-01-01

    To promote the functional restoration of the nervous system following injury, it is necessary to provide optimal extracellular signals that can induce neuronal regenerative activities, particularly neurite formation. This study aimed to examine the regulation of neuritogenesis by temperature-controlled repeated thermal stimulation (TRTS) in rat PC12 pheochromocytoma cells, which can be induced by neurotrophic factors to differentiate into neuron-like cells with elongated neurites. A heating plate was used to apply thermal stimulation, and the correlation of culture medium temperature with varying surface temperature of the heating plate was monitored. Plated PC12 cells were exposed to TRTS at two different temperatures via heating plate (preset surface temperature of the heating plate, 39.5°C or 42°C) in growth or differentiating medium for up to 18 h per day. We then measured the extent of growth, neuritogenesis, or acetylcholine esterase (AChE) activity (a neuronal marker). To analyze the mechanisms underlying the effects of TRTS on these cells, we examined changes in intracellular signaling using the following: tropomyosin-related kinase A inhibitor GW441756; p38 mitogen-activated protein kinase (MAPK) inhibitor SB203580; and MAPK/extracellular signal-regulated kinase (ERK) kinase (MEK) inhibitor U0126 with its inactive analog, U0124, as a control. While a TRTS of 39.5°C did not decrease the growth rate of cells in the cell growth assay, it did increase the number of neurite-bearing PC12 cells and AChE activity without the addition of other neuritogenesis inducers. Furthermore, U0126, and SB203580, but not U0124 and GW441756, considerably inhibited TRTS-induced neuritogenesis. These results suggest that TRTS can induce neuritogenesis and that participation of both the ERK1/2 and p38 MAPK signaling pathways is required for TRTS-dependent neuritogenesis in PC12 cells. Thus, TRTS may be an effective technique for regenerative neuromedicine. PMID:25879210

  9. Navigating neurites utilize cellular topography of Schwann cell somas and processes for optimal guidance

    Science.gov (United States)

    Lopez-Fagundo, Cristina; Mitchel, Jennifer A.; Ramchal, Talisha D.; Dingle, Yu-Ting L.; Hoffman-Kim, Diane

    2013-01-01

    The path created by aligned Schwann cells (SCs) after nerve injury underlies peripheral nerve regeneration. We developed geometric bioinspired substrates to extract key information needed for axon guidance by deconstructing the topographical cues presented by SCs. We have previously reported materials that directly replicate SC topography with micro- and nanoscale resolution, but a detailed explanation of the means of directed axon extension on SC topography has not yet been described. Here, using neurite tracing and time-lapse microscopy, we analyzed the SC features that influence axon guidance. Novel poly(dimethylsiloxane) materials, fabricated via photolithography, incorporated bioinspired topographical components with the shapes and sizes of aligned SCs, namely somas and processes, where the length of the processes were varied but the soma geometry and dimensions were kept constant. Rat dorsal root ganglia neurites aligned to all materials presenting bioinspired topography after a 5 days in culture and to bioinspired materials presenting soma and process features after only 17 hours in culture. Key findings of this study were: Neurite response to underlying bioinspired topographical features was time dependent, where at 5 days, neurites aligned most strongly to materials presenting combinations of soma and process features, with higher than average density of either process or soma features; but at 17 hours they aligned more strongly to materials presenting average densities of soma and process features and to materials presenting process features only. These studies elucidate the influence of SC topography on axon guidance in a time-dependent setting and have implications for the optimization of nerve regeneration strategies. PMID:23557939

  10. Incomplete segregation of endorgan-specific vestibular ganglion cells in mice and rats

    Science.gov (United States)

    Maklad, A.; Fritzsch, B.

    1999-01-01

    The endorgan-specific distribution of vestibular ganglion cells was studied in neonatal and postnatal rats and mice using indocarbocyanine dye (DiI) and dextran amines for retrograde and anterograde labeling. Retrograde DiI tracing from the anterior vertical canal labeled neurons scattered throughout the whole superior vestibular ganglion, with denser labeling at the dorsal and central regions. Horizontal canal neurons were scattered along the dorsoventral axis with more clustering toward the dorsal and ventral poles of this axis. Utricular ganglion cells occupied predominantly the central region of the superior vestibular ganglion. This utricular population overlapped with both the anterior vertical and horizontal canals' ganglion cells. Posterior vertical canal neurons were clustered in the posterior part of the inferior vestibular ganglion. The saccular neurons were distributed in the two parts of the vestibular ganglion, the superior and inferior ganglia. Within the inferior ganglion, the saccular neurons were clustered in the anterior part. In the superior ganglion, the saccular neurons were widely scattered throughout the whole ganglion with more numerous neurons at the posterior half. Small and large neurons were labeled from all endorgans. Examination of the fiber trajectory within the superior division of the vestibular nerve showed no clear lamination of the fibers innervating the different endorgans. These results demonstrate an overlapping pattern between the different populations within the superior ganglion, while in the inferior ganglion, the posterior canal and saccular neurons show tighter clustering but incomplete segregation. This distribution implies that the ganglion cells are assigned for their target during development in a stochastic rather than topographical fashion.

  11. Regional differences in myelination of chick vestibulocochlear ganglion cells.

    Science.gov (United States)

    Sun, Ying-Jie; Kobayashi, Hiroto; Yoshida, Saori; Shirasawa, Nobuyuki; Naito, Akira

    2013-11-01

    In vertebrates, vestibular and cochlear ganglion (VG and CG, respectively) cells are bipolar neurons with myelinated axons and perikarya. The time course of the myelination of the VG and CG cells during development of chick embryos was investigated. Chick VG and CG from embryonic day at 7-20 (E7-20) were prepared for a transmission electron microscopy, myelin basic protein immunohistochemistry, and real-time quantitative RT-PCR. In the VG cells, myelination was first observed on the peripheral axons of the ampullar nerves at E10, on the utricular and saccular nerves at E12, and on the lagenar and neglecta nerves at E13. In the VG central axons, myelination was first seen on the ampullar nerves at E11, on the utricular and saccular nerves at E13, and on the lagenar nerves at E13. In the CG cells, the myelination was first observed on the peripheral and central axons at E14. In both VG and CG, myelination was observed on the perikarya at E17. These results suggest that the onset of the axonal myelination on the VG cells occurred earlier than that on the CG cells, whereas the perikaryal myelination occurred at about the same time on the both types of ganglion cells. Moreover, the myelination on the ampullar nerves occurred earlier than that on the utricular and saccular nerves. The myelination on the peripheral axons occurred earlier than that on the central axons of the VG cells, whereas that on the central and peripheral axons of the CG cells occurred at about the same time. The regional differences in myelination in relation to the onset of functional activities in the VG and CG cells are discussed. Copyright © 2013 ISDN. Published by Elsevier Ltd. All rights reserved.

  12. The circadian response of intrinsically photosensitive retinal ganglion cells.

    Directory of Open Access Journals (Sweden)

    Andrew J Zele

    Full Text Available Intrinsically photosensitive retinal ganglion cells (ipRGC signal environmental light level to the central circadian clock and contribute to the pupil light reflex. It is unknown if ipRGC activity is subject to extrinsic (central or intrinsic (retinal network-mediated circadian modulation during light entrainment and phase shifting. Eleven younger persons (18-30 years with no ophthalmological, medical or sleep disorders participated. The activity of the inner (ipRGC and outer retina (cone photoreceptors was assessed hourly using the pupil light reflex during a 24 h period of constant environmental illumination (10 lux. Exogenous circadian cues of activity, sleep, posture, caffeine, ambient temperature, caloric intake and ambient illumination were controlled. Dim-light melatonin onset (DLMO was determined from salivary melatonin assay at hourly intervals, and participant melatonin onset values were set to 14 h to adjust clock time to circadian time. Here we demonstrate in humans that the ipRGC controlled post-illumination pupil response has a circadian rhythm independent of external light cues. This circadian variation precedes melatonin onset and the minimum ipRGC driven pupil response occurs post melatonin onset. Outer retinal photoreceptor contributions to the inner retinal ipRGC driven post-illumination pupil response also show circadian variation whereas direct outer retinal cone inputs to the pupil light reflex do not, indicating that intrinsically photosensitive (melanopsin retinal ganglion cells mediate this circadian variation.

  13. Uptake of Retrograde Tracers by Intact Optic Nerve Axons: A New Way to Label Retinal Ganglion Cells

    OpenAIRE

    Liang, Yu-Xiang; Yang, Jian; Yuan, Ti-Fei; So, Kwok-Fai

    2015-01-01

    Retrograde labelling of retinal ganglion cells with optic nerve transection often leads to degeneration of ganglion cells in prolonged experiments. Here we report that an intact optic nerve could uptake retrograde tracers applied onto the surface of the nerve, leading to high efficiency labelling of ganglion cells in the retina with long-term survival of cells. This method labelled a similar number of ganglion cells (2289 +/- 174 at 2 days) as the retrograde labeling technique from the superi...

  14. Identification of ganglion cell neurites in human subretinal and epiretinal membranes

    NARCIS (Netherlands)

    Lewis, Geoffrey P.; Betts, Kellen E.; Sethi, Charanjit S.; Charteris, David G.; Lesnik-Oberstein, Sarit Y.; Avery, Robert L.; Fisher, Steven K.

    2007-01-01

    To determine whether neural elements are present in subretinal and epiretinal proliferative vitreoretinopathy (PVR) membranes as well as in diabetic, fibrovascular membranes removed from patients during vitrectomy surgery. Human subretinal and epiretinal membranes of varying durations were

  15. Diversity in spatial scope of contrast adaptation among mouse retinal ganglion cells.

    Science.gov (United States)

    Khani, Mohammad Hossein; Gollisch, Tim

    2017-12-01

    Retinal ganglion cells adapt to changes in visual contrast by adjusting their response kinetics and sensitivity. While much work has focused on the time scales of these adaptation processes, less is known about the spatial scale of contrast adaptation. For example, do small, localized contrast changes affect a cell's signal processing across its entire receptive field? Previous investigations have provided conflicting evidence, suggesting that contrast adaptation occurs either locally within subregions of a ganglion cell's receptive field or globally over the receptive field in its entirety. Here, we investigated the spatial extent of contrast adaptation in ganglion cells of the isolated mouse retina through multielectrode-array recordings. We applied visual stimuli so that ganglion cell receptive fields contained regions where the average contrast level changed periodically as well as regions with constant average contrast level. This allowed us to analyze temporal stimulus integration and sensitivity separately for stimulus regions with and without contrast changes. We found that the spatial scope of contrast adaptation depends strongly on cell identity, with some ganglion cells displaying clear local adaptation, whereas others, in particular large transient ganglion cells, adapted globally to contrast changes. Thus, the spatial scope of contrast adaptation in mouse retinal ganglion cells appears to be cell-type specific. This could reflect differences in mechanisms of contrast adaptation and may contribute to the functional diversity of different ganglion cell types. NEW & NOTEWORTHY Understanding whether adaptation of a neuron in a sensory system can occur locally inside the receptive field or whether it always globally affects the entire receptive field is important for understanding how the neuron processes complex sensory stimuli. For mouse retinal ganglion cells, we here show that both local and global contrast adaptation exist and that this diversity in

  16. Rabies virus infection of cultured adult mouse dorsal root ganglion neurons

    Directory of Open Access Journals (Sweden)

    Jaime Castellanos

    1996-10-01

    Full Text Available An in vitro model of adult dorsal root ganglion neurons infection by rabies virus is described. Viral marked neurotropism is observed, and the percentage and the degree of infection of the neurons is higher than in non neuronal cells, even if neurons are the minority of the cells in the culture. The neuritic tree is also heavily infected by the virus.

  17. Enhancement of the nerve growth factor-mediated neurite outgrowth from PC12D cells by Chinese and Paraguayan medicinal plants.

    Science.gov (United States)

    Li, P; Matsunaga, K; Ohizumi, Y

    1999-07-01

    It is very important to search for natural compounds possessing nerve growth factor (NGF)-potentiating activity. Extracts of 7 Chinese and 10 Paraguayan medicinal plants were examined for their effects on the NGF-mediated neurite outgrowth from PC12D cells to evaluate their NGF-potentiating activities. In the methanol extracts, Gymmopteris rufa (LINN.) BERNH, Ruta graveolens LINN. and Picrorhiza scrophulariiflora PENNELL markedly increased the proportion of neurite-bearing cells. In the case of ethyl acetate fractions, Equisetum giganteum LINN. produced the most powerful enhancement of the proportion of the neurite-bearing cells, and the activities were in the following decreasing order: Equisetum giganteum LINN., Gymmopteris rufa (LINN.) BERNH, Ruta graveolens LINN., and Picrorhiza scrophulariiflora PENNELL. In the water fractions, Imperata cylindrica, Ginseng Radix, Gymmopteris rufa (LINN.) BERNH, Gochnatia polymorpha (LESS) CAB and Picrorhiza scrophulariiflora PENNELL caused a weak enhancement of the proportion of PC12D cells with neurites. Of all the extracts and fractions, the methanol extract of Picrorhiza scrophulariiflora PENNELL induced the longest neurites in PC12D cells. In the ethyl acetate and water fractions of Nardostachys chinensis, long neurites were observed although only a small proportion of PC12D cells had neurites. On the other hand, in the ethyl acetate fraction of Equisetum gigantheum LINN., while the length of the neurites was short, the proportion of neurite-bearing cells was largest among all the extracts and fractions.

  18. Regulation of Taurine transporter activity in cultured rat retinal ganglion cells and rat retinal Muller Cells

    International Nuclear Information System (INIS)

    Eissa, Laila A.; Smith, Sylvia B.; El-sherbeny, Amira A.

    2006-01-01

    Diabetic retinopathy is one of the most common complications of diabetes. The amino acid taurine is believed to play an antioxidant protective role in diabetic retinopathy through the scavenging of the reactive species. It is not well established whether taurine uptake is altered in retina cells during diabetic conditions. Thus, the present study was designed to investigate the changes in taurine transport in cultures of rat retinal Muller cells and rat retinal ganglion cells under conditions associated with diabetes. Taurine was abundantly taken up by retinal Muller cells and rat retinal ganglion cells under normal glycemic condition. Taurine was actively transported to rat Muller cells and rat retinal ganglion cells in a Na and Cl dependant manner. Taurine uptake further significantly elevated in both type of cells after the incubation with high glucose concentration. This effect could be attributed to the increase in osmolarity. Because Nitric Oxide (NO) is a molecule implicated in the pathogenesis of diabetes, we also determined the activity of taurine transporter in cultured rat retinal Muller cells and rat retinal ganglion cells in the presence of the NO donors, SIN-1 and SNAP. Taurine uptake was elevated above control value after 24-h incubation with low concentration of NO donors. We finally investigated the ability of neurotoxic glutamate to change taurine transporter activity in both types of cells. Uptake of taurine was significantly increased in rat retinal ganglion cells when only incubated with high concentration of glutamate. Our data provide evidence that taurine transporter is present in cultured rat retinal ganglion and Muller cells and is regulated by hyperosmolarity. The data are relevant to disease such as diabetes and neuronal degeneration where retinal cell volume may dramatically change. (author)

  19. Organic Photovoltaics and Bioelectrodes Providing Electrical Stimulation for PC12 Cell Differentiation and Neurite Outgrowth.

    Science.gov (United States)

    Hsiao, Yu-Sheng; Liao, Yan-Hao; Chen, Huan-Lin; Chen, Peilin; Chen, Fang-Chung

    2016-04-13

    Current bioelectronic medicines for neurological therapies generally involve treatment with a bioelectronic system comprising a power supply unit and a bioelectrode device. Further integration of wireless and self-powered units is of practical importance for implantable bioelectronics. In this study, we developed biocompatible organic photovoltaics (OPVs) for serving as wireless electrical power supply units that can be operated under illumination with near-infrared (NIR) light, and organic bioelectronic interface (OBEI) electrode devices as neural stimulation electrodes. The OPV/OBEI integrated system is capable to provide electrical stimulation (ES) as a means of enhancing neuron-like PC12 cell differentiation and neurite outgrowth. For the OPV design, we prepared devices incorporating two photoactive material systems--β-carotene/N,N'-dioctyl-3,4,9,10-perylenedicarboximide (β-carotene/PTCDI-C8) and poly(3-hexylthiophene)/phenyl-C61-butyric acid methyl ester (P3HT/PCBM)--that exhibited open circuit voltages of 0.11 and 0.49 V, respectively, under NIR light LED (NLED) illumination. Then, we connected OBEI devices with different electrode gaps, incorporating biocompatible poly(hydroxymethylated-3,4-ethylenedioxythiophene), to OPVs to precisely tailor the direct current electric field conditions during the culturing of PC12 cells. This NIR light-driven OPV/OBEI system could be engineered to provide tunable control over the electric field (from 220 to 980 mV mm(-1)) to promote 64% enhancement in the neurite length, direct the neurite orientation on chips, or both. The OPV/OBEI integrated systems under NIR illumination appear to function as effective power delivery platforms that should meet the requirements for wirelessly offering medical ES to a portion of the nervous system; they might also be a key technology for the development of next-generation implantable bioelectronics.

  20. Hypoxia-ischemia and retinal ganglion cell damage

    Directory of Open Access Journals (Sweden)

    Charanjit Kaur

    2008-08-01

    Full Text Available Charanjit Kaur1, Wallace S Foulds2, Eng-Ang Ling11Department of Anatomy, Yong Loo Lin School of Medicine, National University of Singapore, Singapore; 2Singapore Eye Research Institute, SingaporeAbstract: Retinal hypoxia is the potentially blinding mechanism underlying a number of sight-threatening disorders including central retinal artery occlusion, ischemic central retinal vein thrombosis, complications of diabetic eye disease and some types of glaucoma. Hypoxia is implicated in loss of retinal ganglion cells (RGCs occurring in such conditions. RGC death occurs by apoptosis or necrosis. Hypoxia-ischemia induces the expression of hypoxia inducible factor-1α and its target genes such as vascular endothelial growth factor (VEGF and nitric oxide synthase (NOS. Increased production of VEGF results in disruption of the blood retinal barrier leading to retinal edema. Enhanced expression of NOS results in increased production of nitric oxide which may be toxic to the cells resulting in their death. Excess glutamate release in hypoxic-ischemic conditions causes excitotoxic damage to the RGCs through activation of ionotropic and metabotropic glutamate receptors. Activation of glutamate receptors is thought to initiate damage in the retina by a cascade of biochemical effects such as neuronal NOS activation and increase in intracellular Ca2+ which has been described as a major contributing factor to RGC loss. Excess production of proinflammatory cytokines also mediates cell damage. Besides the above, free-radicals generated in hypoxic-ischemic conditions result in RGC loss because of an imbalance between antioxidant- and oxidant-generating systems. Although many advances have been made in understanding the mediators and mechanisms of injury, strategies to improve the damage are lacking. Measures to prevent neuronal injury have to be developed.Keywords: retinal hypoxia, retinal ganglion cells, glutamate receptors, neuronal injury, retina

  1. Activation of ganglion cells and axon bundles using epiretinal electrical stimulation.

    Science.gov (United States)

    Grosberg, Lauren E; Ganesan, Karthik; Goetz, Georges A; Madugula, Sasidhar S; Bhaskhar, Nandita; Fan, Victoria; Li, Peter; Hottowy, Pawel; Dabrowski, Wladyslaw; Sher, Alexander; Litke, Alan M; Mitra, Subhasish; Chichilnisky, E J

    2017-09-01

    Epiretinal prostheses for treating blindness activate axon bundles, causing large, arc-shaped visual percepts that limit the quality of artificial vision. Improving the function of epiretinal prostheses therefore requires understanding and avoiding axon bundle activation. This study introduces a method to detect axon bundle activation on the basis of its electrical signature and uses the method to test whether epiretinal stimulation can directly elicit spikes in individual retinal ganglion cells without activating nearby axon bundles. Combined electrical stimulation and recording from isolated primate retina were performed using a custom multielectrode system (512 electrodes, 10-μm diameter, 60-μm pitch). Axon bundle signals were identified by their bidirectional propagation, speed, and increasing amplitude as a function of stimulation current. The threshold for bundle activation varied across electrodes and retinas, and was in the same range as the threshold for activating retinal ganglion cells near their somas. In the peripheral retina, 45% of electrodes that activated individual ganglion cells (17% of all electrodes) did so without activating bundles. This permitted selective activation of 21% of recorded ganglion cells (7% of expected ganglion cells) over the array. In one recording in the central retina, 75% of electrodes that activated individual ganglion cells (16% of all electrodes) did so without activating bundles. The ability to selectively activate a subset of retinal ganglion cells without axon bundles suggests a possible novel architecture for future epiretinal prostheses. NEW & NOTEWORTHY Large-scale multielectrode recording and stimulation were used to test how selectively retinal ganglion cells can be electrically activated without activating axon bundles. A novel method was developed to identify axon activation on the basis of its unique electrical signature and was used to find that a subset of ganglion cells can be activated at single-cell

  2. Glial cell line-derived neurotrophic factor (GDNF) induces neuritogenesis in the cochlear spiral ganglion via neural cell adhesion molecule (NCAM)

    Science.gov (United States)

    Euteneuer, Sara; Yang, Kuo H.; Chavez, Eduardo; Leichtle, Anke; Loers, Gabriele; Olshansky, Adel; Pak, Kwang; Schachner, Melitta; Ryan, Allen F.

    2013-01-01

    Glial cell line-derived neurotrophic factor (GDNF) increases survival and neurite extension of spiral ganglion neurons (SGNs), the primary neurons of the auditory system, via yet unknown signaling mechanisms. In other cell types, signaling is achieved by the GPI-linked GDNF family receptor α1 (GFRα1) via recruitment of transmembrane receptors: Ret (re-arranged during transformation) and/or NCAM (neural cell adhesion molecule). Here we show that GDNF enhances neuritogenesis in organotypic cultures of spiral ganglia from 5-day-old rats and mice. Addition of GFRα1-Fc increases this effect. GDNF/GFRα1-Fc stimulation activates intracellular PI3K/Akt and MEK/Erk signaling cascades as detected by Western blot analysis of cultures prepared from rats at postnatal days 5 (P5, before the onset of hearing) and 20 (P20, after the onset of hearing). Both cascades mediate GDNF stimulation of neuritogenesis, since application of the Akt inhibitor Wortmannin or the Erk inhibitor U0126 abolished GDNF/GFRα1-Fc stimulated neuritogenesis in P5 rats. Since cultures of P5 NCAM-deficient mice failed to respond by neuritogenesis to GDNF/GFRα1-Fc, we conclude that NCAM serves as a receptor for GDNF signaling responsible for neuritogenesis in early postnatal spiral ganglion. PMID:23262364

  3. Neurite outgrowth of murine cerebellar granule cells can be enhanced by aniracetam with or without alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid (AMPA).

    Science.gov (United States)

    Fushiki, S; Matsumoto, K; Nagata, A

    1995-10-27

    To assess the neurotrophic effects of a nootropic drug, aniracetam, we studied neurite extension of mouse cerebellar granule cells in culture with low or with high K+ under different combinations of drugs and then immunohistochemically stained the cells with an antibody against L1, a neural cell adhesion molecule on cerebellar granule cells. Quantitative analyses using parameters of the total neurite length, maximal neurite length and number of branches disclosed that aniracetam, in the presence of high K+ and alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid (AMPA), significantly enhanced neurite extension of cultured granule neurons. Aniracetam alone also stimulated neurite extension of cerebellar granule cells at a longer period of culture with low K+ showing a bell-shaped dose response curve with maximal effects at 10 microM. Aniracetam may influence remodeling of the neural network after injury.

  4. Quantitative assessment of neurite outgrowth in human embryonic stem cell derived hN2 cells using automated high-content image analysis

    Science.gov (United States)

    Throughout development neurons undergo a number of morphological changes including neurite outgrowth from the cell body. Exposure to neurotoxic chemicals that interfere with this process may result in permanent deficits in nervous system function. Traditionally, rodent primary ne...

  5. Neurite outgrowth induced by a synthetic peptide ligand of neural cell adhesion molecule requires fibroblast growth factor receptor activation

    DEFF Research Database (Denmark)

    Rønn, L C; Doherty, P; Holm, A

    2000-01-01

    The neural cell adhesion molecule NCAM is involved in axonal outgrowth and target recognition in the developing nervous system. In vitro, NCAM-NCAM binding has been shown to induce neurite outgrowth, presumably through an activation of fibroblast growth factor receptors (FGFRs). We have recently...

  6. Comparison of PC12 and Cerebellar Granule Cell Cultures for Evaluating Neurite Outgrowth Using High Content Screening

    Science.gov (United States)

    Development of high-throughput assays for chemical screening and hazard identification is a pressing priority worldwide. One approach uses in vitro, cell-based assays which recapitulate biological events observed in vivo. Neurite outgrowth is one such critical cellular process un...

  7. Spinal motor neurite outgrowth over glial scar inhibitors is enhanced by coculture with bone marrow stromal cells.

    Science.gov (United States)

    Wright, Karina T; Uchida, Kenzo; Bara, Jennifer J; Roberts, Sally; El Masri, Wagih; Johnson, William E B

    2014-08-01

    Transplantation of bone marrow cells into spinal cord lesions promotes functional recovery in animal models, and recent clinical trials suggest possible recovery also in humans. The mechanisms responsible for these improvements are still unclear. To characterize spinal cord motor neurite interactions with human bone marrow stromal cells (MSCs) in an in vitro model of spinal cord injury (SCI). Previously, we have reported that human MSCs promote the growth of extending sensory neurites from dorsal root ganglia (DRG), in the presence of some of the molecules present in the glial scar, which are attributed with inhibiting axonal regeneration after SCI. We have adapted and optimized this system replacing the DRG with a spinal cord culture to produce a central nervous system (CNS) model, which is more relevant to the SCI situation. We have developed and characterized a novel spinal cord culture system. Human MSCs were cocultured with spinal motor neurites in substrate choice assays containing glial scar-associated inhibitors of nerve growth. In separate experiments, MSC-conditioned media were analyzed and added to spinal motor neurites in substrate choice assays. As has been reported previously with DRG, substrate-bound neurocan and Nogo-A repelled spinal neuronal adhesion and neurite outgrowth, but these inhibitory effects were abrogated in MSC/spinal cord cocultures. However, unlike DRG, spinal neuronal bodies and neurites showed no inhibition to substrates of myelin-associated glycoprotein. In addition, the MSC secretome contained numerous neurotrophic factors that stimulated spinal neurite outgrowth, but these were not sufficient stimuli to promote spinal neurite extension over inhibitory concentrations of neurocan or Nogo-A. These findings provide novel insight into how MSC transplantation may promote regeneration and functional recovery in animal models of SCI and in the clinic, especially in the chronic situation in which glial scars (and associated neural

  8. Retinal Ganglion Cell Distribution and Spatial Resolving Power in Deep-Sea Lanternfishes (Myctophidae)

    KAUST Repository

    De Busserolles, Fanny

    2014-01-01

    Topographic analyses of retinal ganglion cell density are very useful in providing information about the visual ecology of a species by identifying areas of acute vision within the visual field (i.e. areas of high cell density). In this study, we investigated the neural cell distribution in the ganglion cell layer of a range of lanternfish species belonging to 10 genera. Analyses were performed on wholemounted retinas using stereology. Topographic maps were constructed of the distribution of all neurons and both ganglion and amacrine cell populations in 5 different species from Nissl-stained retinas using cytological criteria. Amacrine cell distribution was also examined immunohistochemically in 2 of the 5 species using anti-parvalbumin antibody. The distributions of both the total neuron and the amacrine cell populations were aligned in all of the species examined, showing a general increase in cell density toward the retinal periphery. However, when the ganglion cell population was topographically isolated from the amacrine cell population, which comprised up to 80% of the total neurons within the ganglion cell layer, a different distribution was revealed. Topographic maps of the true ganglion cell distribution in 18 species of lanternfishes revealed well-defined specializations in different regions of the retina. Different species possessed distinct areas of high ganglion cell density with respect to both peak density and the location and/or shape of the specialized acute zone (i.e. elongated areae ventro-temporales, areae temporales and large areae centrales). The spatial resolving power was calculated to be relatively low (varying from 1.6 to 4.4 cycles per degree), indicating that myctophids may constitute one of the less visually acute groups of deep-sea teleosts. The diversity in retinal specializations and spatial resolving power within the family is assessed in terms of possible ecological functions and evolutionary history.

  9. Pleurotus giganteus (Berk.) Karunarathna & K.D. Hyde: Nutritional value and in vitro neurite outgrowth activity in rat pheochromocytoma cells.

    Science.gov (United States)

    Phan, Chia-Wei; Wong, Wei-Lun; David, Pamela; Naidu, Murali; Sabaratnam, Vikineswary

    2012-07-19

    Drugs dedicated to alleviate neurodegenerative diseases like Parkinson's and Alzheimer's have always been associated with debilitating side effects. Medicinal mushrooms which harness neuropharmacological compounds offer a potential possibility for protection against such diseases. Pleurotus giganteus (formerly known as Panus giganteus) has been consumed by the indigenous people in Peninsular Malaysia for many years. Domestication of this wild mushroom is gaining popularity but to our knowledge, medicinal properties reported for this culinary mushroom are minimal. The fruiting bodies P. giganteus were analysed for its nutritional values. Cytotoxicity of the mushroom's aqueous and ethanolic extracts towards PC12, a rat pheochromocytoma cell line was assessed by using 3-[4,5-dimethythiazol-2-yl]-2,5-diphenyltetrazolium bromide (MTT) assay. Neurite outgrowth stimulation assay was carried out with nerve growth factor (NGF) as control. To elucidate signaling mechanisms involved by mushroom extract-induced neurite outgrowth, treatment of specific inhibitor for MEK/ERK and PI3K signalling pathway was carried out. The fruiting bodies of P. giganteus were found to have high carbohydrate, dietary fibre, potassium, phenolic compounds and triterpenoids. Both aqueous and ethanolic extracts induced neurite outgrowth of PC12 cells in a dose- and time-dependant manner with no detectable cytotoxic effect. At day 3, 25 μg/ml of aqueous extract and 15 μg/ml of ethanolic extract showed the highest percentage of neurite-bearing cells, i.e. 31.7 ± 1.1% and 33.3 ± 0.9%; respectively. Inhibition treatment results suggested that MEK/ERK and PI3K/Akt are responsible for neurite outgrowth of PC12 cells stimulated by P. giganteus extract. The high potassium content (1345.7 mg/100 g) may be responsible for promoting neurite extension, too. P. giganteus contains bioactive compounds that mimic NGF and are responsible for neurite stimulation. Hence, this mushroom may be

  10. Brimonidine prevents axonal and somatic degeneration of retinal ganglion cell neurons

    Directory of Open Access Journals (Sweden)

    Crish Samuel D

    2011-01-01

    Full Text Available Abstract Background Brimonidine is a common drug for lowering ocular pressure and may directly protect retinal ganglion cells in glaucoma. The disease involves early loss of retinal ganglion cell transport to brain targets followed by axonal and somatic degeneration. We examined whether brimonidine preserves ganglion cell axonal transport and abates degeneration in rats with elevated ocular pressure induced by laser cauterization of the episcleral veins. Results Ocular pressure was elevated unilaterally by 90% for a period of 8 weeks post- cauterization. During this time, brimonidine (1mg/kg/day or vehicle (phosphate-buffered saline was delivered systemically and continuously via subcutaneous pump. Animals received bilateral intravitreal injections of fluorescent cholera toxin subunit β (CTB two days before sacrifice to assess anterograde transport. In retinas from the vehicle group, elevated pressure induced a 44% decrease in the fraction of ganglion cells with intact uptake of CTB and a 14-42% reduction in the number of immuno-labelled ganglion cell bodies, with the worst loss occurring nasally. Elevated pressure also caused a 33% loss of ganglion cell axons in vehicle optic nerves and a 70% decrease in CTB transport to the superior colliculus. Each of these components of ganglion cell degeneration was either prevented or significantly reduced in the brimonidine treatment group. Conclusions Continuous and systemic treatment with brimonidine by subcutaneous injection significantly improved retinal ganglion cell survival with exposure to elevated ocular pressure. This effect was most striking in the nasal region of the retina. Brimonidine treatment also preserved ganglion cell axon morphology, sampling density and total number in the optic nerve with elevated pressure. Consistent with improved outcome in the optic projection, brimonidine also significantly reduced the deficits in axonal transport to the superior colliculus associated with

  11. Melanopsin ganglion cells extend dendrites into the outer retina during early postnatal development.

    Science.gov (United States)

    Renna, Jordan M; Chellappa, Deepa K; Ross, Christopher L; Stabio, Maureen E; Berson, David M

    2015-09-01

    Melanopsin ganglion cells express the photopigment melanopsin and are the first functional photoreceptors to develop in the mammalian retina. They have been shown to play a variety of important roles in visual development and behavior in the early postnatal period (Johnson et al., 2010; Kirkby and Feller, 2013; Rao et al., 2013; Renna et al., 2011). Here, we probed the maturation of the dendritic arbors of melanopsin ganglion cells during this developmental period in mice. We found that some melanopsin ganglion cells (mainly the M1-subtype) transiently extend their dendrites not only into the inner plexiform layer (where they receive synaptic inputs from bipolar and amacrine cells) but also into the outer plexiform layer, where in mature retina, rod and cone photoreceptors are thought to contact only bipolar and horizontal cells. Thus, some immature melanopsin ganglion cells are biplexiform. This feature is much less common although still present in the mature retina. It reaches peak incidence 8-12 days after birth, before the eyes open and bipolar cells are sufficiently mature to link rods and cones to ganglion cells. At this age, some outer dendrites of melanopsin ganglion cells lie in close apposition to the axon terminals of cone photoreceptors and express a postsynaptic marker of glutamatergic transmission, postsynaptic density-95 protein (PSD-95). These findings raise the possibility of direct, monosynaptic connections between cones and melanopsin ganglion cells in the early postnatal retina. We provide a detailed description of the developmental profile of these processes and consider their possible functional and evolutionary significance. © 2015 Wiley Periodicals, Inc.

  12. Spatial distribution of excitatory synapses on the dendrites of ganglion cells in the mouse retina.

    Directory of Open Access Journals (Sweden)

    Yin-Peng Chen

    Full Text Available Excitatory glutamatergic inputs from bipolar cells affect the physiological properties of ganglion cells in the mammalian retina. The spatial distribution of these excitatory synapses on the dendrites of retinal ganglion cells thus may shape their distinct functions. To visualize the spatial pattern of excitatory glutamatergic input into the ganglion cells in the mouse retina, particle-mediated gene transfer of plasmids expressing postsynaptic density 95-green fluorescent fusion protein (PSD95-GFP was used to label the excitatory synapses. Despite wide variation in the size and morphology of the retinal ganglion cells, the expression of PSD95 puncta was found to follow two general rules. Firstly, the PSD95 puncta are regularly spaced, at 1-2 µm intervals, along the dendrites, whereby the presence of an excitatory synapse creates an exclusion zone that rules out the presence of other glutamatergic synaptic inputs. Secondly, the spatial distribution of PSD95 puncta on the dendrites of diverse retinal ganglion cells are similar in that the number of excitatory synapses appears to be less on primary dendrites and to increase to a plateau on higher branch order dendrites. These observations suggest that synaptogenesis is spatially regulated along the dendritic segments and that the number of synaptic contacts is relatively constant beyond the primary dendrites. Interestingly, we also found that the linear puncta density is slightly higher in large cells than in small cells. This may suggest that retinal ganglion cells with a large dendritic field tend to show an increased connectivity of excitatory synapses that makes up for their reduced dendrite density. Mapping the spatial distribution pattern of the excitatory synapses on retinal ganglion cells thus provides explicit structural information that is essential for our understanding of how excitatory glutamatergic inputs shape neuronal responses.

  13. Ionomycin-induced calcium influx induces neurite degeneration in mouse neuroblastoma cells: analysis of a time-lapse live cell imaging system.

    Science.gov (United States)

    Nakamura, Saki; Nakanishi, Ayumi; Takazawa, Minami; Okihiro, Shunsuke; Urano, Shiro; Fukui, Koji

    2016-01-01

    Reactive oxygen species induce neuronal cell death. However, the detailed mechanisms of cell death have not yet been elucidated. Previously, we reported neurite degeneration before the induction of cell death. Here, we attempted to elucidate the mechanisms of neurite degeneration before the induction of cell death using the neuroblastoma N1E-115 cell line and a time-lapse live cell imaging system. Treatment with the calcium ionophore ionomycin induced cell death and neurite degeneration in a concentration- and time-dependent manner. Treatment with a low concentration of ionomycin immediately produced a significant calcium influx into the intracellular region in N1E-115 cells. After 1-h incubation with ionomycin, the fluorescence emission of MitoSOX TM increased significantly compared to the control. Finally, analysis using a new mitochondrial specific fluorescence dye, MitoPeDPP, indicated that treatment with ionomycin significantly increased the mitochondrial lipid hydroperoxide production in N1E-115 cells. The fluorescence emissions of Fluo-4 AM and MitoPeDPP were detected in the cell soma and neurite regions in ionomycin-treated N1E-115 cells. However, the emissions of neurites were much lower than those of the cell soma. TBARS values of ionomycin-treated cells significantly increased compared to the control. These results indicate that ionomycin induces calcium influx into the intracellular region and reactive oxygen species production in N1E-115 cells. Lipid hydroperoxide production was induced in ionomycin-treated N1E-115 cells. Calcium influx into the intracellular region is a possible activator of neurite degeneration.

  14. The prescriptions from Shenghui soup enhanced neurite growth and GAP-43 expression level in PC12 cells.

    Science.gov (United States)

    Zhang, Qi; Zhang, Zi-Jian; Wang, Xing-Hua; Ma, Jie; Song, Yue-Han; Liang, Mi; Lin, Sen-Xiang; Zhao, Jie; Zhang, Ao-Zhe; Li, Feng; Hua, Qian

    2016-09-20

    Shenghui soup is a traditional Chinese herbal medicine used in clinic for the treatment of forgetfulness. In order to understanding the prescription principle, the effects of "tonifying qi and strengthening spleen" group (TQSS) including Poria cocos (Schw.) Wolf. and Panax ginseng C.A.Mey and "eliminating phlegm and strengthening intelligence" group (EPSI) composed of Polygala tenuifolia Willd., Acorus calamus L. and Sinapis alba L from the herb complex on neurite growth in PC12 cells, two disassembled prescriptions derived from Shenghui soup and their molecular mechanisms were investigated. Firstly, CCK-8 kit was used to detect the impact of the two prescriptions on PC12 cell viability; and Flow cytometry was performed to measure the cell apoptosis when PC12 cells were treated with these drugs. Secondly, the effect of the two prescriptions on the differentiation of PC12 cells was observed. Finally, the mRNA and protein expression levels of GAP-43 were analyzed by RT-PCR and western blot, respectively. "Tonifying qi and strengthening spleen" prescription decreased cell viability in a dose-dependent manner, but had no significant effect on cell apoptosis. Meanwhile, it could improve neurite growth and elevate the mRNA and protein expression level of GAP-43. "Eliminating phlegm and strengthening intelligence" prescription also exerted the similar effects on cell viability and apoptosis. Furthermore, it could also enhance cell neurite growth, with a higher expression level of GAP-43 mRNA and protein. "Tonifying qi and strengthening spleen" and "eliminating phlegm and strengthening intelligence" prescriptions from Shenghui soup have a positive effect on neurite growth. Their effects are related to the up-regulating expression of GAP-43.

  15. Retinal ganglion cell projections to the hamster suprachiasmatic nucleus, intergeniculate leaflet, and visual midbrain: bifurcation and melanopsin immunoreactivity

    Science.gov (United States)

    Morin, Lawrence P.; Blanchard, Jane H.; Provencio, Ignacio

    2003-01-01

    The circadian clock in the suprachiasmatic nucleus (SCN) receives direct retinal input via the retinohypothalamic tract (RHT), and the retinal ganglion cells contributing to this projection may be specialized with respect to direct regulation of the circadian clock. However, some ganglion cells forming the RHT bifurcate, sending axon collaterals to the intergeniculate leaflet (IGL) through which light has secondary access to the circadian clock. The present studies provide a more extensive examination of ganglion cell bifurcation and evaluate whether ganglion cells projecting to several subcortical visual nuclei contain melanopsin, a putative ganglion cell photopigment. The results showed that retinal ganglion cells projecting to the SCN send collaterals to the IGL, olivary pretectal nucleus, and superior colliculus, among other places. Melanopsin-immunoreactive (IR) ganglion cells are present in the hamster retina, and some of these cells project to the SCN, IGL, olivary pretectal nucleus, or superior colliculus. Triple-label analysis showed that melanopsin-IR cells bifurcate and project bilaterally to each SCN, but not to the other visual nuclei evaluated. The melanopsin-IR cells have photoreceptive characteristics optimal for circadian rhythm regulation. However, the presence of moderately widespread bifurcation among ganglion cells projecting to the SCN, and projection by melanopsin-IR cells to locations distinct from the SCN and without known rhythm function, suggest that this ganglion cell type is generalized, rather than specialized, with respect to the conveyance of photic information to the brain. Copyright 2003 Wiley-Liss, Inc.

  16. Pigment epithelium-derived factor protects retinal ganglion cells

    Directory of Open Access Journals (Sweden)

    Fleenor Debra L

    2007-01-01

    Full Text Available Abstract Background Retinal ganglion cells (RGCs are responsible for the transmission of visual signals to the brain. Progressive death of RGCs occurs in glaucoma and several other retinal diseases, which can lead to visual impairment and blindness. Pigment epithelium-derived factor (PEDF is a potent antiangiogenic, neurotrophic and neuroprotective protein that can protect neurons from a variety of pathologic insults. We tested the effects of PEDF on the survival of cultured adult rat RGCs in the presence of glaucoma-like insults, including cytotoxicity induced by glutamate or withdrawal of trophic factors. Results Cultured adult rat RGCs exposed to glutamate for 3 days showed signs of cytotoxicity and death. The toxic effect of glutamate was concentration-dependent (EC50 = 31 μM. In the presence of 100 μM glutamate, RGC number decreased to 55 ± 4% of control (mean ± SEM, n = 76; P 50 values of 13.6 ng/mL (glutamate and 3.4 ng/mL (trophic factor withdrawal, respectively. At 100 ng/mL, PEDF completely protected the cells from both insults. Inhibitors of the nuclear factor κB (NFκB and extracellular signal-regulated kinases 1/2 (ERK1/2 significantly reduced the protective effects of PEDF. Conclusion We demonstrated that PEDF potently and efficaciously protected adult rat RGCs from glutamate- and trophic factor withdrawal-mediated cytotoxicity, via the activation of the NFκB and ERK1/2 pathways. The neuroprotective effect of PEDF represents a novel approach for potential treatment of retinopathies, such as glaucoma.

  17. Potentiation of nerve growth factor-induced neurite outgrowth in PC12 cells by ifenprodil: the role of sigma-1 and IP3 receptors.

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    Tamaki Ishima

    Full Text Available In addition to both the α1 adrenergic receptor and N-methyl-D-aspartate (NMDA receptor antagonists, ifenprodil binds to the sigma receptor subtypes 1 and 2. In this study, we examined the effects of ifenprodil on nerve growth factor (NGF-induced neurite outgrowth in PC12 cells. Ifenprodil significantly potentiated NGF-induced neurite outgrowth, in a concentration-dependent manner. In contrast, the α1 adrenergic receptor antagonist, prazosin and the NMDA receptor NR2B antagonist, Ro 25-6981 did not alter NGF-induced neurite outgrowth. Potentiation of NGF-induced neurite outgrowth mediated by ifenprodil was significantly antagonized by co-administration of the selective sigma-1 receptor antagonist, NE-100, but not the sigma-2 receptor antagonist, SM-21. Similarly, ifenprodil enhanced NGF-induced neurite outgrowth was again significantly reduced by the inositol 1,4,5-triphosphate (IP(3 receptor antagonists, xestospongin C and 2-aminoethoxydiphenyl borate (2-APB treatment. Furthermore, BAPTA-AM, a chelator of intracellular Ca(2+, blocked the effects of ifenprodil on NGF-induced neurite outgrowth, indicating the role of intracellular Ca(2+ in the neurite outgrowth. These findings suggest that activation at sigma-1 receptors and subsequent interaction with IP(3 receptors may mediate the pharmacological effects of ifenprodil on neurite outgrowth.

  18. Retinal ganglion cells: Energetics, compartmentation, axonal transport, cytoskeletons and vulnerability.

    Science.gov (United States)

    Yu, Dao-Yi; Cringle, Stephen J; Balaratnasingam, Chandrakumar; Morgan, William H; Yu, Paula K; Su, Er-Ning

    2013-09-01

    Retinal ganglion cells (RGCs) are specialized projection neurons that relay an immense amount of visual information from the retina to the brain. RGC signal inputs are collected by dendrites and output is distributed from the cell body via very thin (0.5-1 μm) and long (∼50 mm) axons. The RGC cell body is larger than other retinal neurons, but is still only a very small fraction (one ten thousandths) of the length and total surface area of the axon. The total distance traversed by RGCs extends from the retina, starting from synapses with bipolar and amacrine cells, to the brain, to synapses with neurons in the lateral geniculate nucleus. This review will focus on the energy demands of RGCs and the relevant tissues that surround them. RGC survival and function unexceptionally depends upon free energy, predominantly adenosine triphosphate (ATP). RGC energy metabolism is vastly different when compared to that of the photoreceptors. Each subcellular component of the RGC is remarkably different in terms of structure, function and extracellular environment. The energy demands and distribution of each component are also distinct as evidenced by the uneven distribution of mitochondria and ATP within the RGC - signifying the presence of intracellular energy gradients. In this review we will describe RGCs as having four subcellular components, (1) Dendrites, (2) Cell body, (3) Non-myelinated axon, including intraocular and optic nerve head portions, and (4) Myelinated axon, including the intra-orbital and intracranial portions. We will also describe how RGCs integrate information from each subcellular component in order achieve intracellular homeostatic stability as well as respond to perturbations in the extracellular environment. The possible cellular mechanisms such as axonal transport and axonal cytoskeleton proteins that are involved in maintaining RGC energy homeostasis during normal and disease conditions will also be discussed in depth. The emphasis of this

  19. Ganglion cell adaptability: does the coupling of horizontal cells play a role?

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    Karin Dedek

    Full Text Available BACKGROUND: The visual system can adjust itself to different visual environments. One of the most well known examples of this is the shift in spatial tuning that occurs in retinal ganglion cells with the change from night to day vision. This shift is thought to be produced by a change in the ganglion cell receptive field surround, mediated by a decrease in the coupling of horizontal cells. METHODOLOGY/PRINCIPAL FINDINGS: To test this hypothesis, we used a transgenic mouse line, a connexin57-deficient line, in which horizontal cell coupling was abolished. Measurements, both at the ganglion cell level and the level of behavioral performance, showed no differences between wild-type retinas and retinas with decoupled horizontal cells from connexin57-deficient mice. CONCLUSION/SIGNIFICANCE: This analysis showed that the coupling and uncoupling of horizontal cells does not play a dominant role in spatial tuning and its adjustability to night and day light conditions. Instead, our data suggest that another mechanism, likely arising in the inner retina, must be responsible.

  20. Taurine Provides Neuroprotection against Retinal Ganglion Cell Degeneration

    Science.gov (United States)

    Froger, Nicolas; Cadetti, Lucia; Lorach, Henri; Martins, Joao; Bemelmans, Alexis-Pierre; Dubus, Elisabeth; Degardin, Julie; Pain, Dorothée; Forster, Valérie; Chicaud, Laurent; Ivkovic, Ivana; Simonutti, Manuel; Fouquet, Stéphane; Jammoul, Firas; Léveillard, Thierry; Benosman, Ryad; Sahel, José-Alain; Picaud, Serge

    2012-01-01

    Retinal ganglion cell (RGC) degeneration occurs in numerous retinal diseases leading to blindness, either as a primary process like in glaucoma, or secondary to photoreceptor loss. However, no commercial drug is yet directly targeting RGCs for their neuroprotection. In the 70s, taurine, a small sulfonic acid provided by nutrition, was found to be essential for the survival of photoreceptors, but this dependence was not related to any retinal disease. More recently, taurine deprivation was incriminated in the retinal toxicity of an antiepileptic drug. We demonstrate here that taurine can improve RGC survival in culture or in different animal models of RGC degeneration. Taurine effect on RGC survival was assessed in vitro on primary pure RCG cultures under serum-deprivation conditions, and on NMDA-treated retinal explants from adult rats. In vivo, taurine was administered through the drinking water in two glaucomatous animal models (DBA/2J mice and rats with vein occlusion) and in a model of Retinitis pigmentosa with secondary RGC degeneration (P23H rats). After a 6-day incubation, 1 mM taurine significantly enhanced RGCs survival (+68%), whereas control RGCs were cultured in a taurine-free medium, containing all natural amino-acids. This effect was found to rely on taurine-uptake by RGCs. Furthermore taurine (1 mM) partly prevented NMDA-induced RGC excitotoxicity. Finally, taurine supplementation increased RGC densities both in DBA/2J mice, in rats with vein occlusion and in P23H rats by contrast to controls drinking taurine-free water. This study indicates that enriched taurine nutrition can directly promote RGC survival through RGC intracellular pathways. It provides evidence that taurine can positively interfere with retinal degenerative diseases. PMID:23115615

  1. Retinal ganglion cells electrophysiology: the effect of cell morphology on impulse waveform.

    Science.gov (United States)

    Maturana, Matias I; Wong, Raymond; Cloherty, Shaun L; Ibbotson, Michael R; Hadjinicolaou, Alex E; Grayden, David B; Burkitt, Anthony N; Meffin, Hamish; O'Brien, Brendan J; Kameneva, Tatiana

    2013-01-01

    There are 16 morphologically defined classes of rats retinal ganglion cells (RGCs). Using computer simulation of a realistic anatomically correct A1 mouse RGC, we investigate the effect of the cell's morphology on its impulse waveform, using the first-, and second-order time derivatives as well as the phase plot features. Using whole cell patch clamp recordings, we recorded the impulse waveform for each of the rat RGCs types. While we found some clear differences in many features of the impulse waveforms for A2 and B2 cells compared to other cell classes, many cell types did not show clear differences.

  2. The morphological characterization of orientation-biased displaced large-field ganglion cells in the central part of goldfish retina.

    Science.gov (United States)

    Hoshi, Hideo; Sato, Fumi

    2018-02-01

    The vertebrate retina has about 30 subtypes of ganglion cells. Each ganglion cell receives synaptic inputs from specific types of bipolar and amacrine cells ramifying at the same depth of the inner plexiform layer (IPL), each of which is thought to process a specific aspect of visual information. Here, we identified one type of displaced ganglion cell in the goldfish retina which had a large and elongated dendritic field. As a population, all of these ganglion cells were oriented in the horizontal axis and perpendicular to the dorsal-ventral axis of the goldfish eye in the central part of retina. This ganglion cell has previously been classified as Type 1.2. However, the circuit elements which synapse with this ganglion cell are not yet characterized. We found that this displaced ganglion cell was directly tracer-coupled only with homologous ganglion cells at sites containing Cx35/36 puncta. We further illustrated that the processes of dopaminergic neurons often terminated next to intersections between processes of ganglion cells, close to where dopamine D1 receptors were localized. Finally, we showed that Mb1 ON bipolar cells had ribbon synapses in the axonal processes passing through the IPL and made ectopic synapses with this displaced ganglion cell that stratified into stratum 1 of the IPL. These results suggest that the displaced ganglion cell may synapse with both Mb1 cells using ectopic ribbon synapses and OFF cone bipolar cells with regular ribbon synapses in the IPL to function in both scotopic and photopic light conditions. © 2017 Wiley Periodicals, Inc.

  3. Hypothermia Protects and Prolongs the Tolerance Time of Retinal Ganglion Cells against Ischemia.

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    Maximilian Schultheiss

    Full Text Available Hypothermia has been shown to be neuroprotective in the therapy of ischemic stroke in the brain. To date no studies exist on the level of the inner retina and it is unclear if hypothermia would prolong the ischemic tolerance time of retinal ganglion cells, which are decisive in many ischemic retinopathies.Bovine eyes were enucleated and stored either at 21°C or 37°C for 100 or 340 minutes, respectively. Afterwards the globes were dissected, the retina was prepared and either the spontaneous ganglion cell responses were measured or the retina was incubated as an organotypic culture for additional 24 hours. After incubation the retina was either processed for histology (H&E and DAPI staining or real-time PCR (Thy-1 expression was performed.Hypothermia prolonged ganglion cell survival up to 340 minutes under ischemic conditions. In contrast to eyes kept at 37°C the eyes stored at 21°C still showed spontaneous ganglion cell spiking (56.8% versus 0%, a 5.8 fold higher Thy-1 mRNA expression (not significant, but a trend and a preserved retinal structure after 340 minutes of ischemia.Hypothermia protects retinal ganglion cells against ischemia and prolongs their ischemic tolerance time.

  4. Expression of EFR3A in the mouse cochlea during degeneration of spiral ganglion following hair cell loss.

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    Chen Nie

    Full Text Available Retrograde degeneration of spiral ganglion cells in the cochlea following hair cell loss is similar to dying back in pathology. The EFR3A gene has recently been discovered to be involved in the pathogenesis of dying back. The relationship of EFR3A and spiral ganglion degeneration, however, was rarely investigated. In this study, we destroyed the hair cells of the mouse cochlea by co-administration of kanamycin and furosemide and then investigated the EFR3A expression during the induced spiral ganglion cell degeneration. Our results revealed that co-administration of kanamycin and furosemide quickly induced hair cell loss in the C57BL/6J mice and then resulted in progressive degeneration of the spiral ganglion beginning at day 5 following drug administration. The number of the spiral ganglion cells began to decrease at day 15. The expression of EFR3A increased remarkably in the spiral ganglion at day 5 and then decreased to near normal level within the next 10 days. Our study suggested that the change of EFR3A expression in the spiral ganglion was coincident with the time of the spiral ganglion degeneration, which implied that high expression of EFR3A may be important to prompt initiation of spiral ganglion degeneration following hair cell loss.

  5. Displaced retinal ganglion cells in albino and pigmented rats

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    Francisco Manuel Nadal-Nicolás

    2014-10-01

    Full Text Available We have studied in parallel the population of displaced retinal ganglion cells (dRGCs and normally placed (orthotopic RGCs, oRGCs in albino and pigmented rats. Using retrograde tracing from the optic nerve, from both superior colliculi (SC or from the ipsilateral SC in conjunction with Brn3 and melanopsin immunodetection, we report for the first time their total number and topography as well as the number and distribution of those dRGCs and oRGCs that project ipsi- or contralaterally and/or that express any of the three Brn3 isoforms or melanopsin. The total number of RGCs (oRGCs+dRGCs is 84,706±1,249 in albino and 90,440±2,236 in pigmented, out of which 2,383 and 2,428 are melanopsin positive (m-RGCs, respectively. Regarding dRGCs: i/ albino rats have a significantly lower number of dRGCs than pigmented animals (0.5% of the total number of RGCs vs. 2.5%, respectively, ii/ dRGCs project massively to the contralateral SC, iii/ the percentage of ipsilaterality is higher for dRGCs than for oRGCs, iv/ a higher proportion of ipsilateral dRGCs is observed in albino than pigmented animals, v/ dRGC topography is very specific, they predominate in the equatorial temporal retina, being densest where the oRGCs are densest, vi/ Brn3a detects all dRGCs except half of the ipsilateral ones and those that express melanopsin, vii/ the proportion of dRGCs that express Brn3b or Brn3c is slightly lower than in the oRGC population, viii/ a higher percentage of dRGCs (13% albino, 9% pigmented than oRGCs (2.6% express melanopsin, ix/ few m-RGCs (displaced and orthotopic project to the ipsilateral SC, x/ the topography of m-dRGCs does not resemble the general distribution of dRGCs, xi/ The soma size in m-oRGCs ranges from 10 to 21 µm and in m-dRGCs from 8 to 15 µm, xii/ oRGCs and dRGCs have the same susceptibility to axonal injury and hypertension. Although the role of mammalian dRGCs remains to be determined, our data suggest that they are not misplaced by an

  6. Stimulation of neuronal neurite outgrowth using functionalized carbon nanotubes

    Energy Technology Data Exchange (ETDEWEB)

    Matsumoto, K; Sato, C; Shimizu, N [Graduate School of Life Sciences, Toyo University, 1-1-1 Izumino, Itakura-machi, Ora-gun, Gunma 374-0193 (Japan); Naka, Y [Bio-Nano Electronics Research Center, Toyo University, 2100 Kujirai, Kawagoe-shi, Saitama 350-8585 (Japan); Whitby, R, E-mail: shimizu@toyonet.toyo.ac.jp [School of Pharmacy and Biomolecular Sciences, University of Brighton, Cockroft Building, Lewes Road, Brighton BN2 4GJ (United Kingdom)

    2010-03-19

    Low concentrations (0.11-1.7 {mu}g ml{sup -1}) of functionalized carbon nanotubes (CNTs), which are multi-walled CNTs modified by amino groups, when added with nerve growth factor (NGF), promoted outgrowth of neuronal neurites in dorsal root ganglion (DRG) neurons and rat pheochromocytoma cell line PC12h cells in culture media. The quantity of active extracellular signal-regulated kinase (ERK) was higher after the addition of both 0.85 {mu}g ml{sup -1} CNTs and NGF than that with NGF alone. CNTs increased the number of cells with neurite outgrowth in DRG neurons and PC12h cells after the inhibition of the ERK signaling pathway using a mitogen-activated protein kinase (MAPK)/ERK kinase (MEK) inhibitor. Active ERK proteins were detected in MEK inhibitor-treated neurons after the addition of CNTs to the culture medium. These results demonstrate that CNTs may stimulate neurite outgrowth by activation of the ERK signaling pathway. Thus, CNTs are biocompatible and are promising candidates for biological applications and devices.

  7. Stimulation of neuronal neurite outgrowth using functionalized carbon nanotubes

    International Nuclear Information System (INIS)

    Matsumoto, K; Sato, C; Shimizu, N; Naka, Y; Whitby, R

    2010-01-01

    Low concentrations (0.11-1.7 μg ml -1 ) of functionalized carbon nanotubes (CNTs), which are multi-walled CNTs modified by amino groups, when added with nerve growth factor (NGF), promoted outgrowth of neuronal neurites in dorsal root ganglion (DRG) neurons and rat pheochromocytoma cell line PC12h cells in culture media. The quantity of active extracellular signal-regulated kinase (ERK) was higher after the addition of both 0.85 μg ml -1 CNTs and NGF than that with NGF alone. CNTs increased the number of cells with neurite outgrowth in DRG neurons and PC12h cells after the inhibition of the ERK signaling pathway using a mitogen-activated protein kinase (MAPK)/ERK kinase (MEK) inhibitor. Active ERK proteins were detected in MEK inhibitor-treated neurons after the addition of CNTs to the culture medium. These results demonstrate that CNTs may stimulate neurite outgrowth by activation of the ERK signaling pathway. Thus, CNTs are biocompatible and are promising candidates for biological applications and devices.

  8. Dominant inheritance of retinal ganglion cell resistance to optic nerve crush in mice

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    Schlamp Cassandra L

    2007-03-01

    Full Text Available Abstract Background Several neurodegenerative diseases are influenced by complex genetics that affect an individual's susceptibility, disease severity, and rate of progression. One such disease is glaucoma, a chronic neurodegenerative condition of the eye that targets and stimulates apoptosis of CNS neurons called retinal ganglion cells. Since ganglion cell death is intrinsic, it is reasonable that the genes that control this process may contribute to the complex genetics that affect ganglion cell susceptibility to disease. To determine if genetic background influences susceptibility to optic nerve damage, leading to ganglion cell death, we performed optic nerve crush on 15 different inbred lines of mice and measured ganglion cell loss. Resistant and susceptible strains were used in a reciprocal breeding strategy to examine the inheritance pattern of the resistance phenotype. Because earlier studies had implicated Bax as a susceptibility allele for ganglion cell death in the chronic neurodegenerative disease glaucoma, we conducted allelic segregation analysis and mRNA quantification to assess this gene as a candidate for the cell death phenotype. Results Inbred lines showed varying levels of susceptibility to optic nerve crush. DBA/2J mice were most resistant and BALB/cByJ mice were most susceptible. F1 mice from these lines inherited the DBA/2J phenotype, while N2 backcross mice exhibited the BALB/cByJ phenotype. F2 mice exhibited an intermediate phenotype. A Wright Formula calculation suggested as few as 2 dominant loci were linked to the resistance phenotype, which was corroborated by a Punnett Square analysis of the distribution of the mean phenotype in each cross. The levels of latent Bax mRNA were the same in both lines, and Bax alleles did not segregate with phenotype in N2 and F2 mice. Conclusion Inbred mice show different levels of resistance to optic nerve crush. The resistance phenotype is heritable in a dominant fashion involving

  9. TRPC1 is required for survival and proliferation of cochlear spiral ganglion stem/progenitor cells.

    Science.gov (United States)

    Chen, Hsin-Chien; Wang, Chih-Hung; Shih, Cheng-Ping; Chueh, Sheau-Huei; Liu, Shu-Fan; Chen, Hang-Kang; Lin, Yi-Chun

    2015-12-01

    The present studies were designed to test the hypothesis that canonical transient receptor potential channel 1 (TRPC1) is required for the proliferation of cochlear spiral ganglion stem/progenitor cells (SPCs). TRPC1 were detected and evaluated in postnatal day 1 CBA/CaJ mice pups derived-cochlear spiral ganglion SPCs by reverse transcription-polymerase chain reaction, Western blot, immunocytochemistry, and calcium imaging. The cell viability and proliferation of the spiral ganglion SPCs following si-RNA mediated knockdown of TRPC1 or addition of TRPC channel blocker SKF9635 were compared to controls. In spiral ganglion SPCs, TRPC1 was found to be the most abundantly expressed TRPC subunit and shown to contribute to store-operated calcium entry. Silencing of TRPC1 or addition of TRPC channel blockers significantly decreased the rate of cell proliferation. The results suggest that TRPC1 might serve as an essential molecule in regulating the proliferation of spiral ganglion SPCs. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

  10. Strychnine blocks transient but not sustained inhibition in mudpuppy retinal ganglion cells.

    Science.gov (United States)

    Belgum, J H; Dvorak, D R; McReynolds, J S

    1984-01-01

    Transient and sustained inhibitory synaptic inputs to on-centre, off-centre, and on-off ganglion cells in the mudpuppy retina were studied using intracellular recording in the superfused eye-cup preparation. When chemical transmission was blocked with 4 mM-Co2+, application of either glycine or gamma-aminobutyric acid (GABA) caused a hyperpolarization and conductance increase in all ganglion cells. For both amino acids, the responses were dose dependent in the range 0.05-10 mM, with a half-maximal response at about 0.7 mM. Glycine and GABA sensitivities were very similar in all three types of ganglion cells. The response to applied glycine was selectively antagonized by 10(-5) M-strychnine and the response to applied GABA was selectively antagonized by 10(-5) M-picrotoxin. In all ganglion cells, 10(-5) M-strychnine eliminated the transient inhibitory events which occur at the onset and termination of a light stimulus. The block of transient inhibition was associated with a relative depolarization of membrane potential and decrease in conductance at these times. Strychnine had no effect on membrane potential or conductance in darkness or during sustained inhibitory responses to light. Picrotoxin (10(-5) M) did not block transient inhibitory events in any ganglion cells, but did affect other components of their responses. The results suggest that in all three classes of ganglion cells transient inhibition, but not sustained inhibition, may be mediated by glycine or a closely related substance. PMID:6481635

  11. Spiral ganglion cell morphology in guinea pigs after deafening and neurotrophic treatment

    NARCIS (Netherlands)

    van Loon, M.C.; Ramekers, D.; Agterberg, M.J.H.; de Groot, J.C.M.J.; Grolman, W.; Klis, S.F.L.; Versnel, H.

    2013-01-01

    It is well known that spiral ganglion cells (SGCs) degenerate in hair-cell-depleted cochleas and that treatment with exogenous neurotrophins can prevent this degeneration. Several studies reported that, in addition, SGC size decreases after deafening and increases after neurotrophic treatment. The

  12. Temporal response of protein-based artificial ganglion cell receptive field (Conference Presentation)

    Science.gov (United States)

    Okada-Shudo, Yoshiko

    2016-10-01

    We propose ganglion cell receptive-field-type filters with the use of the photoreceptor protein bacteriorhodopsin. Visual image processing is possible with the use of only one sensing element. We also demonstrate that our difference of Gaussians (DOG) filter, which mimics on-center off-suround ganglion cell receptive fields, has the function of a Laplacian filter and can act as an edge detecor. The X-type receptive field responses obtained by the filter, for a variety of stimuli, are compared with available electrophysiological recodings.

  13. Retinal Ganglion Cell Diversity and Subtype Specification from Human Pluripotent Stem Cells

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    Kirstin B. Langer

    2018-04-01

    Full Text Available Summary: Retinal ganglion cells (RGCs are the projection neurons of the retina and transmit visual information to postsynaptic targets in the brain. While this function is shared among nearly all RGCs, this class of cell is remarkably diverse, comprised of multiple subtypes. Previous efforts have identified numerous RGC subtypes in animal models, but less attention has been paid to human RGCs. Thus, efforts of this study examined the diversity of RGCs differentiated from human pluripotent stem cells (hPSCs and characterized defined subtypes through the expression of subtype-specific markers. Further investigation of these subtypes was achieved using single-cell transcriptomics, confirming the combinatorial expression of molecular markers associated with these subtypes, and also provided insight into more subtype-specific markers. Thus, the results of this study describe the derivation of RGC subtypes from hPSCs and will support the future exploration of phenotypic and functional diversity within human RGCs. : In this article, Langer and colleagues present extensive characterization of RGC subtypes derived from human pluripotent stem cells, with multiple subtypes identified by subtype-specific molecular markers. Their results present a more detailed analysis of RGC diversity in human cells and yield the use of different markers to identify RGC subtypes. Keywords: iPSC, retina, retinal ganglion cell, RGC subtype, stem cell, ipRGC, alpha RGC, direction selective RGC, RNA-seq

  14. Light-evoked somatosensory perception of transgenic rats that express channelrhodopsin-2 in dorsal root ganglion cells.

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    Zhi-Gang Ji

    Full Text Available In vertebrate somatosensory systems, each mode of touch-pressure, temperature or pain is sensed by sensory endings of different dorsal root ganglion (DRG neurons, which conducted to the specific cortical loci as nerve impulses. Therefore, direct electrical stimulation of the peripheral nerve endings causes an erroneous sensation to be conducted by the nerve. We have recently generated several transgenic lines of rat in which channelrhodopsin-2 (ChR2 transgene is driven by the Thy-1.2 promoter. In one of them, W-TChR2V4, some neurons were endowed with photosensitivity by the introduction of the ChR2 gene, coding an algal photoreceptor molecule. The DRG neurons expressing ChR2 were immunohistochemically identified using specific antibodies to the markers of mechanoreceptive or nociceptive neurons. Their peripheral nerve endings in the plantar skin as well as the central endings in the spinal cord were also examined. We identified that ChR2 is expressed in a certain population of large neurons in the DRG of W-TChR2V4. On the basis of their morphology and molecular markers, these neurons were classified as mechanoreceptive but not nociceptive. ChR2 was also distributed in their peripheral sensory nerve endings, some of which were closely associated with CK20-positive cells to form Merkel cell-neurite complexes or with S-100-positive cells to form structures like Meissner's corpuscles. These nerve endings are thus suggested to be involved in the sensing of touch. Each W-TChR2V4 rat showed a sensory-evoked behavior in response to blue LED flashes on the plantar skin. It is thus suggested that each rat acquired an unusual sensory modality of sensing blue light through the skin as touch-pressure. This light-evoked somatosensory perception should facilitate study of how the complex tactile sense emerges in the brain.

  15. Visual Field Defects and Retinal Ganglion Cell Losses in Human Glaucoma Patients

    Science.gov (United States)

    Harwerth, Ronald S.; Quigley, Harry A.

    2007-01-01

    Objective The depth of visual field defects are correlated with retinal ganglion cell densities in experimental glaucoma. This study was to determine whether a similar structure-function relationship holds for human glaucoma. Methods The study was based on retinal ganglion cell densities and visual thresholds of patients with documented glaucoma (Kerrigan-Baumrind, et al.) The data were analyzed by a model that predicted ganglion cell densities from standard clinical perimetry, which were then compared to histologic cell counts. Results The model, without free parameters, produced accurate and relatively precise quantification of ganglion cell densities associated with visual field defects. For 437 sets of data, the unity correlation for predicted vs. measured cell densities had a coefficient of determination of 0.39. The mean absolute deviation of the predicted vs. measured values was 2.59 dB, the mean and SD of the distribution of residual errors of prediction was -0.26 ± 3.22 dB. Conclusions Visual field defects by standard clinical perimetry are proportional to neural losses caused by glaucoma. Clinical Relevance The evidence for quantitative structure-function relationships provides a scientific basis of interpreting glaucomatous neuropathy from visual thresholds and supports the application of standard perimetry to establish the stage of the disease. PMID:16769839

  16. Thrombospondin-1 modified bone marrow mesenchymal stem cells (BMSCs) promote neurite outgrowth and functional recovery in rats with spinal cord injury.

    Science.gov (United States)

    Pu, Yujie; Meng, Ke; Gu, Chuanlong; Wang, Linlin; Zhang, Xiaoming

    2017-11-10

    Stem cell therapies are currently gaining momentum in the treatment of spinal cord injury (SCI). However, unsatisfied intrinsic neurite growth capacity constitutes significant obstacles for injured spinal cord repair and ultimately results in neurological dysfunction. The present study assessed the efficacy of thrombospondin-1 (TSP-1), a neurite outgrowth-promoting molecule, modified bone marrow mesenchymal stem cells (BMSCs) on promoting neurite outgrowth in vitro and in vivo of Oxygen-Glucose Deprivation (OGD) treated motor neurons and SCI rat models. The present results demonstrated that the treatment of BMSCs+TSP-1 could promote the neurite length, neuronal survival, and functional recovery after SCI. Additionally, TSP-1 could activate transforming growth factor-β1 (TGF-β1) then induced the smad2 phosphorylation, and expedited the expression of GAP-43 to promote neurite outgrowth. The present study for the first time demonstrated that BMSCs+TSP-1 could promote neurite outgrowth and functional recovery after SCI partly through the TGF-β1/p-Samd2 pathway. The study provided a novel encouraging evidence for the potential treatment of BMSCs modification with TSP-1 in patients with SCI.

  17. NMDA Receptors Contribute to Retrograde Synaptic Transmission from Ganglion Cell Photoreceptors to Dopaminergic Amacrine Cells

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    Lei-Lei Liu

    2017-09-01

    Full Text Available Recently, a line of evidence has demonstrated that the vertebrate retina possesses a novel retrograde signaling pathway. In this pathway, phototransduction is initiated by the photopigment melanopsin, which is expressed in a small population of retinal ganglion cells. These ganglion cell photoreceptors then signal to dopaminergic amacrine cells (DACs through glutamatergic synapses, influencing visual light adaptation. We have previously demonstrated that in Mg2+-containing solution, α-amino-3-hydroxyl-5-methyl-4-isoxazole-propionate (AMPA receptors mediate this glutamatergic transmission. Here, we demonstrate that removing extracellular Mg2+ enhances melanopsin-based DAC light responses at membrane potentials more negative than −40 mV. Melanopsin-based responses in Mg2+-free solution were profoundly suppressed by the selective N-methyl-D-aspartate (NMDA receptor antagonist D-AP5. In addition, application of NMDA to the retina produced excitatory inward currents in DACs. These data strongly suggest that DACs express functional NMDA receptors. We further found that in the presence of Mg2+, D-AP5 reduced the peak amplitude of melanopsin-based DAC responses by ~70% when the cells were held at their resting membrane potential (−50 mV, indicating that NMDA receptors are likely to contribute to retrograde signal transmission to DACs under physiological conditions. Moreover, our data show that melanopsin-based NMDA-receptor-mediated responses in DACs are suppressed by antagonists specific to either the NR2A or NR2B receptor subtype. Immunohistochemical results show that NR2A and NR2B subunits are expressed on DAC somata and processes. These results suggest that DACs express functional NMDA receptors containing both NR2A and NR2B subunits. Collectively, our data reveal that, along with AMPA receptors, NR2A- and NR2B-containing NMDA receptors mediate retrograde signal transmission from ganglion cell photoreceptors to DACs.

  18. [Vimentin and S100 protein in the developing rat dorsal root ganglion cells].

    Science.gov (United States)

    Kolos, Ye A; Korzhevskiy, D E

    2013-01-01

    The aim of this research was to study the expression of vimentin, one of the intermediate filament proteins, and S100 protein in the cells of developing dorsal root ganglion in prenatal ontogenesis in Wistar rat embryos (n=22) on 12-19 days of development and in newborn rats (n=6). Immunohistochemical staining methods were used to study the formation of rat dorsal root ganglion glial cells. Vimentin-immunopositive neural and glial progenitor cells were present in dorsal root ganglion primordia in rat embryos at 12-17 prenatal days. At the later stages (day 19 of prenatal life and day 1 of postnatal life) vimentin was found to be synthesized only by the differentiating glial cells. It was shown that at the same time, S100 protein was first detected in the glial cells of rat dorsal root ganglion. On postnatal day 1, the tendency was noted for the accumulation of S100 protein in the larger neurons, that was characteristic to mature dorsal root ganglia.

  19. Retinal ganglion cell neuroprotection by an angiotensin II blocker in an ex vivo retinal explant model.

    Science.gov (United States)

    White, Andrew J R; Heller, Janosch P; Leung, Johahn; Tassoni, Alessia; Martin, Keith R

    2015-12-01

    An ex vivo organotypic retinal explant model was developed to examine retinal survival mechanisms relevant to glaucoma mediated by the renin angiotensin system in the rodent eye. Eyes from adult Sprague Dawley rats were enucleated immediately post-mortem and used to make four retinal explants per eye. Explants were treated either with irbesartan (10 µM), vehicle or angiotensin II (2 μM) for four days. Retinal ganglion cell density was estimated by βIII tubulin immunohistochemistry. Live imaging of superoxide formation with dihydroethidium (DHE) was performed. Protein expression was determined by Western blotting, and mRNA expression was determined by RT-PCR. Irbesartan (10 µM) almost doubled ganglion cell survival after four days. Angiotensin II (2 µM) reduced cell survival by 40%. Sholl analysis suggested that irbesartan improved ganglion cell dendritic arborisation compared to control and angiotensin II reduced it. Angiotensin-treated explants showed an intense DHE fluorescence not seen in irbesartan-treated explants. Analysis of protein and mRNA expression determined that the angiotensin II receptor At1R was implicated in modulation of the NADPH-dependent pathway of superoxide generation. Angiotensin II blockers protect retinal ganglion cells in this model and may be worth further investigation as a neuroprotective treatment in models of eye disease. © The Author(s) 2015.

  20. Adult human nasal mesenchymal-like stem cells restore cochlear spiral ganglion neurons after experimental lesion.

    Science.gov (United States)

    Bas, Esperanza; Van De Water, Thomas R; Lumbreras, Vicente; Rajguru, Suhrud; Goss, Garrett; Hare, Joshua M; Goldstein, Bradley J

    2014-03-01

    A loss of sensory hair cells or spiral ganglion neurons from the inner ear causes deafness, affecting millions of people. Currently, there is no effective therapy to repair the inner ear sensory structures in humans. Cochlear implantation can restore input, but only if auditory neurons remain intact. Efforts to develop stem cell-based treatments for deafness have demonstrated progress, most notably utilizing embryonic-derived cells. In an effort to bypass limitations of embryonic or induced pluripotent stem cells that may impede the translation to clinical applications, we sought to utilize an alternative cell source. Here, we show that adult human mesenchymal-like stem cells (MSCs) obtained from nasal tissue can repair spiral ganglion loss in experimentally lesioned cochlear cultures from neonatal rats. Stem cells engraft into gentamicin-lesioned organotypic cultures and orchestrate the restoration of the spiral ganglion neuronal population, involving both direct neuronal differentiation and secondary effects on endogenous cells. As a physiologic assay, nasal MSC-derived cells engrafted into lesioned spiral ganglia demonstrate responses to infrared laser stimulus that are consistent with those typical of excitable cells. The addition of a pharmacologic activator of the canonical Wnt/β-catenin pathway concurrent with stem cell treatment promoted robust neuronal differentiation. The availability of an effective adult autologous cell source for inner ear tissue repair should contribute to efforts to translate cell-based strategies to the clinic.

  1. REDUCED GANGLION CELL VOLUME ON OPTICAL COHERENCE TOMOGRAPHY IN PATIENTS WITH GEOGRAPHIC ATROPHY.

    Science.gov (United States)

    Ramkumar, Hema L; Nguyen, Brian; Bartsch, Dirk-Uwe; Saunders, Luke J; Muftuoglu, Ilkay Kilic; You, Qisheng; Freeman, William R

    2017-11-07

    Geographic atrophy (GA) is the sequelae of macular degeneration. Automated inner retinal analysis using optical coherence tomography is flawed because segmentation software is calibrated for normal eyes. The purpose of this study is to determine whether ganglion cell layer (GCL) volume is reduced in GA using manual analysis. Nineteen eyes with subfoveal GA and 22 controls were selected for morphometric analyses. Heidelberg scanning laser ophthalmoscope optical coherence tomography images of the optic nerve and macula were obtained, and the Viewing Module was used to manually calibrate retinal layer segmentation. Retinal layer volumes in the central 3-mm and surrounding 6-mm diameter were measured. Linear mixed models were used for statistics. The GCL volume in the central 3 mm of the macula is less (P = 0.003), and the retinal nerve fiber layer volume is more (P = 0.02) in patients with GA when compared with controls. Ganglion cell layer volume positively correlated with outer nuclear layer volume (P = 0.020). The patients with geographic atrophy have a small significant loss of the GCL. Ganglion cell death may precede axonal loss, and increased macular retinal nerve fiber layer volumes are not indicative of GCL volume. Residual ganglion cell stimulation by interneurons may enable vision in patients with GA.

  2. Loss of Melanopsin-Expressing Retinal Ganglion Cells in Patients With Diabetic Retinopathy

    DEFF Research Database (Denmark)

    Obara, Elisabeth Anne; Hannibal, Jens; Heegaard, Steffen

    2017-01-01

    Purpose: Photo-entrainment of the circadian clock is mediated by melanopsin-expressing retinal ganglion cells (mRGCs) located in the retina. Patients suffering from diabetic retinopathy (DR) show impairment of light regulated circadian activity such as sleep disorders, altered blood pressure...

  3. Melanopsin retinal ganglion cells are resistant to neurodegeneration in mitochondrial optic neuropathies

    DEFF Research Database (Denmark)

    La Morgia, C; Ross-Cisneros, F.N.; Sadun, A.A.

    2010-01-01

    Mitochondrial optic neuropathies, that is, Leber hereditary optic neuropathy and dominant optic atrophy, selectively affect retinal ganglion cells, causing visual loss with relatively preserved pupillary light reflex. The mammalian eye contains a light detection system based on a subset of retina...

  4. Modulation of H2O2-Induced Neurite Outgrowth Impairment and Apoptosis in PC12 Cells by a 1,2,4-Triazine Derivative

    Directory of Open Access Journals (Sweden)

    Solaleh Khoramian Tusi

    2012-12-01

    Full Text Available Increased oxidative stress is widely accepted to be a factor in the development and progression of Alzheimer’s disease. Triazine derivatives possess a wide range of pharmacological activities including anti-oxidative and anti-inflammatory actions. In this study, we aimed to investigate the possible protective effect of 3-thioethyl-5,6-dimethoxyphenyl-1,2,4-triazine (TEDMT on H2O2-induced neurite outgrowth impairment and apoptosis in neuron-like PC12 cells. We pretreated PC12 cells with 5, 7, and 10 µM of TEDMT followed by adding H2O2 as an oxidative stress agent. We found that TEDMT contributed to up-regulation of Bcl-2, down regulation of Bax protein and reduction of cleaved Caspase-3 and PARP proteins. Moreover, TEDMT could inhibit the phosphorylation of different mitogen activated protein kinases (extracellular signal-regulated kinase, c-Jun N-terminal kinase and P38 mitogen-activated protein kinase. TEDMT induced heat shock protein 70 while decreased heat shock protein 90 level. Besides we measured six different parameters of neurite outgrowth and complexity. We showed that H2O2 increased cell body area, average neurite width and the proportion of bipolar cells, while decreased average neurite length, the numbers of primary neurites and the ratio of the total neurite branching nodes to the total number of primary neurites. Interestingly, we found that TEDMT not only protects PC12 cell against H2O2-induced apoptosis, but also defends against the destructive effect of oxidative stress on the criteria of neural differentiation. Protective effect of this compound could represent a promising approach for treatment of neurodegenerative diseases.

  5. Dendritic maturation in cat retinal ganglion cells: a Lucifer yellow study.

    Science.gov (United States)

    Dann, J F; Buhl, E H; Peichl, L

    1987-09-11

    The dendritic morphology of developing cat alpha- and beta-retinal ganglion cells was investigated by intracellular injection of Lucifer yellow. In both cell classes the basic pattern of adult morphology was present at birth. However, the presence of transient small spiny protrusions along the dendrites was characteristic of early postnatal cells. Many alpha-cells were further distinguished by a small degree of dendritic bi-stratification which disappeared within the first 5 postnatal days. Therefore during the period before the eyes opened (P7-P10) there was a considerable degree of modification and maturation in dendritic morphology in both classes of retinal ganglion cells. alpha- and beta-cells exhibited differing temporal patterns of dendritic growth, which argues against a 'passive-stretching' hypothesis that explains dendritic field enlargement solely as an effect of retinal areal growth.

  6. [Study on the induced differentiation of induced pluripotent stem cells into cochlear hair cell-like cells and spiral ganglion neuron-like cells in vitro].

    Science.gov (United States)

    Guan, Lina; Chen, Yanhong; Zhu, Hengtao; Chen, Jing; Jiang, Hongqun

    2014-08-01

    In this study, we investigated the potential of mouse induced pluripotent stem cells (iPSC) for use as a source of transplants for the restoration of auditory hair cells and spiral ganglion neurons. We co-cultured the mouse iPSC with the cells of the cochlear organ of Corti or the modiolus in vitro. The cochlear organ of Corti (which contains cochlear hair cells) and the modiolus (which contains auditory spiral ganglion neurons) were obtained from postnatal day 3 (P3) CD-1 ICR mice. After 18 days of coculture with the cells of newborn mouse cochleae. The expressions of hair cell markers (Myosin VIIa, Math1, Calretinin, Espin) and Spiral ganglion neuron markers [Nestin, Neurofilament-M, β-III Tubulin, Vesicular glutamate transporter 1(VGluT1)] were detected by immunocytochemical analysis. Immunocytochemical analysis results indicated that the differentiated iPSC expressed auditory hair cell markers (MyosinVIIa,Math1, Calretinin, Espin ) and spiral ganglion markers (Nestin, Neurofilament-M,β-III Tubulin,VGluT1). Mouse iPSC in virto cultured could successfully be induced to differentiate into hair cell-like cells and spiral ganglion-like cells with hair cell and spiral ganglion molecular markers.

  7. Role of the GM1 ganglioside oligosaccharide portion in the TrkA-dependent neurite sprouting in neuroblastoma cells.

    Science.gov (United States)

    Chiricozzi, Elena; Pomè, Diego Yuri; Maggioni, Margherita; Di Biase, Erika; Parravicini, Chiara; Palazzolo, Luca; Loberto, Nicoletta; Eberini, Ivano; Sonnino, Sandro

    2017-12-01

    GM1 ganglioside (II 3 NeuAc-Gg 4 Cer) is known to promote neurite formation in neuroblastoma cells by activating TrkA-MAPK pathway. The molecular mechanism by which GM1 is involved in the neurodifferentiation process is still unknown, however, in vitro and in vivo evidences have suggested that the oligosaccharide portion of this ganglioside could be involved. Here, we report that, similarly to the entire GM1 molecule, its oligosaccharide II 3 NeuAc-Gg 4, rather than its ceramide (Cer) portion is responsible for the neurodifferentiation process by augmenting neurite elongation and increasing the neurofilament protein expression in murine neuroblastoma cells, Neuro2a. Conversely, asialo-GM1, GM2 and GM3 oligosaccharides are not effective in neurite elongation on Neuro2a cells, whereas the effect exerted by the Fuc-GM1 oligosaccharide (IV 2 αFucII 3 Neu5Ac-Gg 4 ) is similar to that exerted by GM1 oligosaccharide. The neurotrophic properties of GM1 oligosaccharide are exerted by activating the TrkA receptor and the following phosphorylation cascade. By photolabeling experiments performed with a nitrophenylazide containing GM1 oligosaccharide, labeled with tritium, we showed a direct interaction between the GM1 oligosaccharide and the extracellular domain of TrkA receptor. Moreover, molecular docking analyses confirmed that GM1 oligosaccharide binds the TrkA-nerve growth factor complex leading to a binding free energy of approx. -11.5 kcal/mol, acting as a bridge able to increase and stabilize the TrkA-nerve growth factor molecular interactions. © 2017 International Society for Neurochemistry.

  8. Iridoids and sesquiterpenoids of Valeriana stenoptera and their effects on NGF-induced neurite outgrowth in PC12 cells.

    Science.gov (United States)

    Dong, Fa-Wu; Wu, Zhi-Kun; Yang, Liu; Zi, Chen-Ting; Yang, Dan; Ma, Rui-Jing; Liu, Zhen-Hua; Luo, Huai-Rong; Zhou, Jun; Hu, Jiang-Miao

    2015-10-01

    Twenty-one compounds (nine iridoids and twelve sesquiterpenoids), including ten previously unknown (five iridoids and five sesquiterpenoids) were isolated from whole dried material of Valeriana stenoptera. Structures were established on the basis of extensive spectroscopic analysis and the relative stereochemistry of 13-hydroxypatchoulol A was further confirmed by X-ray crystallographic data. All isolates were evaluated for their effects on nerve growth factor (NGF)-mediated neurite outgrowth in pheochromocytoma (PC12) cells and seven compounds showed potent promoting effects. Copyright © 2015 Elsevier Ltd. All rights reserved.

  9. Eliminating Glutamatergic Input onto Horizontal Cells Changes the Dynamic Range and Receptive Field Organization of Mouse Retinal Ganglion Cells.

    Science.gov (United States)

    Ströh, Sebastian; Puller, Christian; Swirski, Sebastian; Hölzel, Maj-Britt; van der Linde, Lea I S; Segelken, Jasmin; Schultz, Konrad; Block, Christoph; Monyer, Hannah; Willecke, Klaus; Weiler, Reto; Greschner, Martin; Janssen-Bienhold, Ulrike; Dedek, Karin

    2018-02-21

    In the mammalian retina, horizontal cells receive glutamatergic inputs from many rod and cone photoreceptors and return feedback signals to them, thereby changing photoreceptor glutamate release in a light-dependent manner. Horizontal cells also provide feedforward signals to bipolar cells. It is unclear, however, how horizontal cell signals also affect the temporal, spatial, and contrast tuning in retinal output neurons, the ganglion cells. To study this, we generated a genetically modified mouse line in which we eliminated the light dependency of feedback by deleting glutamate receptors from mouse horizontal cells. This genetic modification allowed us to investigate the impact of horizontal cells on ganglion cell signaling independent of the actual mode of feedback in the outer retina and without pharmacological manipulation of signal transmission. In control and genetically modified mice (both sexes), we recorded the light responses of transient OFF-α retinal ganglion cells in the intact retina. Excitatory postsynaptic currents (EPSCs) were reduced and the cells were tuned to lower temporal frequencies and higher contrasts, presumably because photoreceptor output was attenuated. Moreover, receptive fields of recorded cells showed a significantly altered surround structure. Our data thus suggest that horizontal cells are responsible for adjusting the dynamic range of retinal ganglion cells and, together with amacrine cells, contribute to the center/surround organization of ganglion cell receptive fields in the mouse. SIGNIFICANCE STATEMENT Horizontal cells represent a major neuronal class in the mammalian retina and provide lateral feedback and feedforward signals to photoreceptors and bipolar cells, respectively. The mode of signal transmission remains controversial and, moreover, the contribution of horizontal cells to visual processing is still elusive. To address the question of how horizontal cells affect retinal output signals, we recorded the light

  10. Retinal ganglion cell topography and spatial resolving power in the white rhinoceros (Ceratotherium simum).

    Science.gov (United States)

    Coimbra, João Paulo; Manger, Paul R

    2017-08-01

    This study sought to determine whether the retinal organization of the white rhinoceros (Ceratotherium simum), a large African herbivore with lips specialized for grazing in open savannahs, relates to its foraging ecology and habitat. Using stereology and retinal wholemounts, we estimated a total of 353,000 retinal ganglion cells. Their density distribution reveals an unusual topographic organization of a temporal (2,000 cells/mm 2 ) and a nasal (1,800 cells/mm 2 ) area embedded within a well-defined horizontal visual streak (800 cells/mm 2 ), which is remarkably similar to the retinal organization in the black rhinoceros. Alpha ganglion cells comprise 3.5% (12,300) of the total population of ganglion cells and show a similar distribution pattern with maximum densities also occurring in the temporal (44 cells/mm 2 ) and nasal (40 cells/mm 2 ) areas. We found higher proportions of alpha cells in the dorsal and ventral retinas. Given their role in the detection of brisk transient stimuli, these higher proportions may facilitate the detection of approaching objects from the front and behind while grazing with the head at 45 °. Using ganglion cell peak density and eye size (29 mm, axial length), we estimated upper limits of spatial resolving power of 7 cycles/deg (temporal area), 6.6 cycles/deg (nasal area), and 4.4 cycles/deg (horizontal streak). The resolution of the temporal area potentially assists with grazing, while the resolution of the streak may be used for panoramic surveillance of the horizon. The nasal area may assist with detection of approaching objects from behind, potentially representing an adaptation compensating for limited neck and head mobility. J. Comp. Neurol., 525:2484-2498, 2017. © 2017 Wiley Periodicals, Inc. © 2017 Wiley Periodicals, Inc.

  11. Muscarinic acetylcholine receptor subtype expression in avian vestibular hair cells, nerve terminals and ganglion cells.

    Science.gov (United States)

    Li, G Q; Kevetter, G A; Leonard, R B; Prusak, D J; Wood, T G; Correia, M J

    2007-04-25

    Muscarinic acetylcholine receptors (mAChRs) are widely expressed in the CNS and peripheral nervous system and play an important role in modulating the cell activity and function. We have shown that the cholinergic agonist carbachol reduces the pigeon's inwardly rectifying potassium channel (pKir2.1) ionic currents in native vestibular hair cells. We have cloned and sequenced pigeon mAChR subtypes M2-M5 and we have studied the expression of all five mAChR subtypes (M1-M5) in the pigeon vestibular end organs (semicircular canal ampullary cristae and utricular maculae), vestibular nerve fibers and the vestibular (Scarpa's) ganglion using tissue immunohistochemistry (IH), dissociated single cell immunocytochemistry (IC) and Western blotting (WB). We found that vestibular hair cells, nerve fibers and ganglion cells each expressed all five (M1-M5) mAChR subtypes. Two of the three odd-numbered mAChRs (M1, M5) were present on the hair cell cilia, supporting cells and nerve terminals. And all three odd numbered mAChRs (M1, M3 and M5) were expressed on cuticular plates, myelin sheaths and Schwann cells. Even-numbered mAChRs were seen on the nerve terminals. M2 was also shown on the cuticular plates and supporting cells. Vestibular efferent fibers and terminals were not identified in our studies. Results from WB of the dissociated vestibular epithelia, nerve fibers and vestibular ganglia were consistent with the results from IH and IC. Our findings suggest that there is considerable co-expression of the subtypes on the neural elements of the labyrinth. Further electrophysiological and pharmacological studies should delineate the mechanisms of action of muscarinic acetylcholine receptors on structures in the labyrinth.

  12. Bone marrow mesenchymal stem cells protect against retinal ganglion cell loss in aged rats with glaucoma

    Directory of Open Access Journals (Sweden)

    Hu Y

    2013-10-01

    Full Text Available Ying Hu,1,2 Hai Bo Tan,1 Xin Mei Wang,3 Hua Rong,1 Hong Ping Cui,1 Hao Cui2 Departments of Ophthalmology, 1Shanghai East Hospital of Tongji University, Shanghai, 2First Affiliated Hospital, 3Fourth Affiliated Hospital, Harbin Medical University, Harbin, People's Republic of China Abstract: Glaucoma is a common eye disease in the aged population and has severe consequences. The present study examined the therapeutic effects of bone marrow mesenchymal stem cell (BMSC transplantation in preventing loss of visual function in aged rats with glaucoma caused by laser-induced ocular hypertension. We found that BMSCs promoted survival of retinal ganglion cells in the transplanted eye as compared with the control eye. Further, in swimming tests guided by visual cues, the rats with a BMSC transplant performed significantly better. We believe that BMSC transplantation therapy is effective in treating aged rats with glaucoma. Keywords: glaucoma, stem cell, transplantation, cell therapy, aging

  13. Biophysical Variation within the M1 Type of Ganglion Cell Photoreceptor

    Directory of Open Access Journals (Sweden)

    Alan J. Emanuel

    2017-10-01

    Full Text Available Intrinsically photosensitive retinal ganglion cells of the M1 type encode environmental irradiance for functions that include circadian and pupillary regulation. Their distinct role, morphology, and molecular markers indicate that they are stereotyped circuit elements, but their physiological uniformity has not been investigated in a systematic fashion. We have profiled the biophysical parameters of mouse M1s and found that extreme variation is their hallmark. Most parameters span 1–3 log units, and the full range is evident in M1s that innervate brain regions serving divergent functions. Biophysical profiles differ among cells possessing similar morphology and between neighboring M1s recorded simultaneously. Variation in each parameter is largely independent of that in others, allowing for flexible individualization. Accordingly, a common stimulus drives heterogeneous spike outputs across cells. By contrast, a population of directionally selective retinal ganglion cells appeared physiologically uniform under similar conditions. Thus, M1s lack biophysical constancy and send diverse signals downstream.

  14. A Learning Model for L/M Specificity in Ganglion Cells

    Science.gov (United States)

    Ahumada, Albert J.

    2016-01-01

    An unsupervised learning model for developing LM specific wiring at the ganglion cell level would support the research indicating LM specific wiring at the ganglion cell level (Reid and Shapley, 2002). Removing the contributions to the surround from cells of the same cone type improves the signal-to-noise ratio of the chromatic signals. The unsupervised learning model used is Hebbian associative learning, which strengthens the surround input connections according to the correlation of the output with the input. Since the surround units of the same cone type as the center are redundant with the center, their weights end up disappearing. This process can be thought of as a general mechanism for eliminating unnecessary cells in the nervous system.

  15. Uniform signal redundancy of parasol and midget ganglion cells in primate retina

    OpenAIRE

    Gauthier, Jeffrey L.; Field, Greg D.; Sher, Alexander; Shlens, Jonathon; Greschner, Martin; Litke, Alan M.; Chichilnisky, E.J.

    2009-01-01

    The collective representation of visual space in high resolution visual pathways was explored by simultaneously measuring the receptive fields of hundreds of ON and OFF midget and parasol ganglion cells in isolated primate retina. As expected, the receptive fields of all four cell types formed regular mosaics uniformly tiling the visual scene. Surprisingly, comparison of all four mosaics revealed that the overlap of neighboring receptive fields was nearly identical, for both the excitatory ce...

  16. Scene from above: retinal ganglion cell topography and spatial resolving power in the giraffe (Giraffa camelopardalis).

    Science.gov (United States)

    Coimbra, João Paulo; Hart, Nathan S; Collin, Shaun P; Manger, Paul R

    2013-06-15

    The giraffe (Giraffa camelopardalis) is a browser that uses its extensible tongue to selectively collect leaves during foraging. As the tallest extant terrestrial mammal, its elevated head height provides panoramic surveillance of the environment. These aspects of the giraffe's ecology and phenotype suggest that vision is of prime importance. Using Nissl-stained retinal wholemounts and stereological methods, we quantitatively assessed the retinal specializations in the ganglion cell layer of the giraffe. The mean total number of retinal ganglion cells was 1,393,779 and their topographic distribution revealed the presence of a horizontal visual streak and a temporal area. With a mean peak of 14,271 cells/mm(2), upper limits of spatial resolving power in the temporal area ranged from 25 to 27 cycles/degree. We also observed a dorsotemporal extension (anakatabatic area) that tapers toward the nasal retina giving rise to a complete dorsal arch. Using neurofilament-200 immunohistochemistry, we also detected a dorsal arch formed by alpha ganglion cells with density peaks in the temporal (14-15 cells/mm(2)) and dorsonasal (10 cells/mm(2)) regions. As with other artiodactyls, the giraffe shares the presence of a horizontal streak and a temporal area which, respectively, improve resolution along the horizon and in the frontal visual field. The dorsal arch is related to the giraffe's head height and affords enhanced resolution in the inferior visual field. The alpha ganglion cell distribution pattern is unique to the giraffe and enhances acquisition of motion information for the control of tongue movement during foraging and the detection of predators. Copyright © 2012 Wiley Periodicals, Inc.

  17. Glia-Neuron Interactions in the Retina Can Be Studied in Cocultures of Muller Cells and Retinal Ganglion Cells

    DEFF Research Database (Denmark)

    Skytt, D. M.; Toft-Kehler, A. K.; Braendstrup, C. T.

    2016-01-01

    Glia-neuron partnership is important for inner retinal homeostasis and any disturbances may result in retinal ganglion cell (RGC) death. Müller cells support RGCs with essential functions such as removing excess glutamate and providing energy sources. The aim was to explore the impact of Müller c...

  18. Diversity of retinal ganglion cells identified by transient GFP transfection in organotypic tissue culture of adult marmoset monkey retina.

    Directory of Open Access Journals (Sweden)

    Satoru Moritoh

    Full Text Available The mammalian retina has more diversity of neurons than scientists had once believed in order to establish complicated vision processing. In the monkey retina, morphological diversity of retinal ganglion cells (RGCs besides dominant midget and parasol cells has been suggested. However, characteristic subtypes of RGCs in other species such as bistratified direction-selective ganglion cells (DSGC have not yet been identified. Increasing interest has been shown in the common marmoset (Callithrix jacchus monkey as a "super-model" of neuroscientific research. Here, we established organotypic tissue culture of the adult marmoset monkey retina with particle-mediated gene transfer of GFP to survey the morphological diversity of RGCs. We successfully incubated adult marmoset monkey retinas for 2 to 4 days ex vivo for transient expression of GFP. We morphologically examined 121 RGCs out of more than 3240 GFP-transfected cells in 5 retinas. Among them, we identified monostratified or broadly stratified ganglion cells (midget, parasol, sparse, recursive, thorny, and broad thorny ganglion cells, and bistratified ganglion cells (recursive, large, and small bistratified ganglion cells [blue-ON/yellow-OFF-like]. By this survey, we also found a candidate for bistratified DSGC whose dendrites were well cofasciculated with ChAT-positive starburst dendrites, costratified with ON and OFF ChAT bands, and had honeycomb-shaped dendritic arbors morphologically similar to those in rabbits. Our genetic engineering method provides a new approach to future investigation for morphological and functional diversity of RGCs in the monkey retina.

  19. ShcA regulates neurite outgrowth stimulated by neural cell adhesion molecule but not by fibroblast growth factor 2: evidence for a distinct fibroblast growth factor receptor response to neural cell adhesion molecule activation

    DEFF Research Database (Denmark)

    Hinsby, Anders M; Lundfald, Line; Ditlevsen, Dorte K

    2004-01-01

    Homophilic binding in trans of the neural cell adhesion molecule (NCAM) mediates adhesion between cells and leads, via activation of intracellular signaling cascades, to neurite outgrowth in primary neurons as well as in the neuronal cell line PC12. NCAM mediates neurite extension in PC12 cells....... Here, we investigated the involvement of adaptor proteins in NCAM and fibroblast growth factor 2 (FGF2)-mediated neurite outgrowth in the PC12-E2 cell line. We found that both FGFR substrate-2 and Grb2 play important roles in NCAM as well as in FGF2-stimulated events. In contrast, the docking protein...... ShcA was pivotal to neurite outgrowth induced by NCAM, but not by FGF2, in PC12 cells. Moreover, in rat cerebellar granule neurons, phosphorylation of ShcA was stimulated by an NCAM mimicking peptide, but not by FGF2. This activation was blocked by inhibitors of both FGFR and Fyn, indicating that NCAM...

  20. Symmetric synaptic patterns between starburst amacrine cells and direction selective ganglion cells in the rabbit retina.

    Science.gov (United States)

    Chen, Yung-Cheng; Chiao, Chuan-Chin

    2008-05-01

    Inputs from starburst amacrine cells (SACs) to ON-OFF direction selective ganglion cells (DSGCs) in the rabbit retina are themselves directional. However, the synaptic asymmetry between SACs and DSGCs required for generating direction selectivity has been controversial. We investigated dendritic contacts and distribution of inhibitory synapses between SACs and their overlapped DSGCs. Double injection of SAC/DSGC pairs and quantitative analysis revealed no obvious asymmetry of dendritic contacts between SACs and DSGCs. Furthermore, examination of the inhibitory input pattern on the dendrites of DSGCs using antibodies against GABA(A) receptors also suggested an isotropic arrangement with the overlapping SACs in both the preferred and the null directions. Therefore, the presynaptic mechanism of direction selectivity upon DSGCs may not result from a simple asymmetric arrangement with overlapping SACs. Multiple layer interactions and sophisticated synaptic connections between SACs and DSGCs are necessary. (c) 2008 Wiley-Liss, Inc.

  1. Seasonally Changing Cryptochrome 1b Expression in the Retinal Ganglion Cells of a Migrating Passerine Bird.

    Directory of Open Access Journals (Sweden)

    Christine Nießner

    Full Text Available Cryptochromes, blue-light absorbing proteins involved in the circadian clock, have been proposed to be the receptor molecules of the avian magnetic compass. In birds, several cryptochromes occur: Cryptochrome 2, Cryptochrome 4 and two splice products of Cryptochrome 1, Cry1a and Cry1b. With an antibody not distinguishing between the two splice products, Cryptochrome 1 had been detected in the retinal ganglion cells of garden warblers during migration. A recent study located Cry1a in the outer segments of UV/V-cones in the retina of domestic chickens and European robins, another migratory species. Here we report the presence of cryptochrome 1b (eCry1b in retinal ganglion cells and displaced ganglion cells of European Robins, Erithacus rubecula. Immuno-histochemistry at the light microscopic and electron microscopic level showed eCry1b in the cell plasma, free in the cytosol as well as bound to membranes. This is supported by immuno-blotting. However, this applies only to robins in the migratory state. After the end of the migratory phase, the amount of eCry1b was markedly reduced and hardly detectable. In robins, the amount of eCry1b in the retinal ganglion cells varies with season: it appears to be strongly expressed only during the migratory period when the birds show nocturnal migratory restlessness. Since the avian magnetic compass does not seem to be restricted to the migratory phase, this seasonal variation makes a role of eCry1b in magnetoreception rather unlikely. Rather, it could be involved in physiological processes controlling migratory restlessness and thus enabling birds to perform their nocturnal flights.

  2. Axonal transmission in the retina introduces a small dispersion of relative timing in the ganglion cell population response.

    Directory of Open Access Journals (Sweden)

    Günther Zeck

    Full Text Available BACKGROUND: Visual stimuli elicit action potentials in tens of different retinal ganglion cells. Each ganglion cell type responds with a different latency to a given stimulus, thus transforming the high-dimensional input into a temporal neural code. The timing of the first spikes between different retinal projection neurons cells may further change along axonal transmission. The purpose of this study is to investigate if intraretinal conduction velocity leads to a synchronization or dispersion of the population signal leaving the eye. METHODOLOGY/PRINCIPAL FINDINGS: We 'imaged' the initiation and transmission of light-evoked action potentials along individual axons in the rabbit retina at micron-scale resolution using a high-density multi-transistor array. We measured unimodal conduction velocity distributions (1.3±0.3 m/sec, mean ± SD for axonal populations at all retinal eccentricities with the exception of the central part that contains myelinated axons. The velocity variance within each piece of retina is caused by ganglion cell types that show narrower and slightly different average velocity tuning. Ganglion cells of the same type respond with similar latency to spatially homogenous stimuli and conduct with similar velocity. For ganglion cells of different type intraretinal conduction velocity and response latency to flashed stimuli are negatively correlated, indicating that differences in first spike timing increase (up to 10 msec. Similarly, the analysis of pair-wise correlated activity in response to white-noise stimuli reveals that conduction velocity and response latency are negatively correlated. CONCLUSION/SIGNIFICANCE: Intraretinal conduction does not change the relative spike timing between ganglion cells of the same type but increases spike timing differences among ganglion cells of different type. The fastest retinal ganglion cells therefore act as indicators of new stimuli for postsynaptic neurons. The intraretinal dispersion

  3. Changes in ganglion cell physiology during retinal degeneration influence excitability by prosthetic electrodes

    Science.gov (United States)

    Cho, Alice; Ratliff, Charles; Sampath, Alapakkam; Weiland, James

    2016-04-01

    Objective. Here we investigate ganglion cell physiology in healthy and degenerating retina to test its influence on threshold to electrical stimulation. Approach. Age-related Macular Degeneration and Retinitis Pigmentosa cause blindness via outer retinal degeneration. Inner retinal pathways that transmit visual information to the central brain remain intact, so direct electrical stimulation from prosthetic devices offers the possibility for visual restoration. Since inner retinal physiology changes during degeneration, we characterize physiological properties and responses to electrical stimulation in retinal ganglion cells (RGCs) of both wild type mice and the rd10 mouse model of retinal degeneration. Main results. Our aggregate results support previous observations that elevated thresholds characterize diseased retinas. However, a physiology-driven classification scheme reveals distinct sub-populations of ganglion cells with thresholds either normal or strongly elevated compared to wild-type. When these populations are combined, only a weakly elevated threshold with large variance is observed. The cells with normal threshold are more depolarized at rest and exhibit periodic oscillations. Significance. During degeneration, physiological changes in RGCs affect the threshold stimulation currents required to evoke action potentials.

  4. Non-cytotoxic Concentration of Cisplatin Decreases Neuroplasticity-Related Proteins and Neurite Outgrowth Without Affecting the Expression of NGF in PC12 Cells.

    Science.gov (United States)

    Ferreira, Rafaela Scalco; Dos Santos, Neife Aparecida Guinaim; Martins, Nádia Maria; Fernandes, Laís Silva; Dos Santos, Antonio Cardozo

    2016-11-01

    Cisplatin is the most effective and neurotoxic platinum chemotherapeutic agent. It induces a peripheral neuropathy characterized by distal axonal degeneration that might progress to degeneration of cell bodies and apoptosis. Most symptoms occur nearby distal axonal branches and axonal degeneration might induce peripheral neuropathy regardless neuronal apoptosis. The toxic mechanism of cisplatin has been mainly associated with DNA damage, but cisplatin might also affect neurite outgrowth. Nevertheless, the neurotoxic mechanism of cisplatin remains unclear. We investigated the early effects of cisplatin on axonal plasticity by using non-cytotoxic concentrations of cisplatin and PC12 cells as a model of neurite outgrowth and differentiation. PC12 cells express NGF-receptors (trkA) and respond to NGF by forming neurites, branches and synaptic vesicles. For comparison, we used a neuronal model (SH-SY5Y cells) that does not express trkA nor responds to NGF. Cisplatin did not change NGF expression in PC12 cells and decreased neurite outgrowth in both models, suggesting a NGF/trkA independent mechanism. It also reduced axonal growth (GAP-43) and synaptic (synapsin I and synaptophysin) proteins in PC12 cells, without inducing mitochondrial damage or apoptosis. Therefore, cisplatin might affect axonal plasticity before DNA damage, NGF/trkA down-regulation, mitochondrial damage or neuronal apoptosis. This is the first study to show that neuroplasticity-related proteins might be early targets of the neurotoxic action of cisplatin and their role on cisplatin-induced peripheral neuropathy should be investigated in vivo.

  5. The Adaptor Protein SH2B3 (Lnk) Negatively Regulates Neurite Outgrowth of PC12 Cells and Cortical Neurons

    Science.gov (United States)

    Wang, Tien-Cheng; Chiu, Hsun; Chang, Yu-Jung; Hsu, Tai-Yu; Chiu, Ing-Ming; Chen, Linyi

    2011-01-01

    SH2B adaptor protein family members (SH2B1-3) regulate various physiological responses through affecting signaling, gene expression, and cell adhesion. SH2B1 and SH2B2 were reported to enhance nerve growth factor (NGF)-induced neuronal differentiation in PC12 cells, a well-established neuronal model system. In contrast, SH2B3 was reported to inhibit cell proliferation during the development of immune system. No study so far addresses the role of SH2B3 in the nervous system. In this study, we provide evidence suggesting that SH2B3 is expressed in the cortex of embryonic rat brain. Overexpression of SH2B3 not only inhibits NGF-induced differentiation of PC12 cells but also reduces neurite outgrowth of primary cortical neurons. SH2B3 does so by repressing NGF-induced activation of PLCγ, MEK-ERK1/2 and PI3K-AKT pathways and the expression of Egr-1. SH2B3 is capable of binding to phosphorylated NGF receptor, TrkA, as well as SH2B1β. Our data further demonstrate that overexpression of SH2B3 reduces the interaction between SH2B1β and TrkA. Consistent with this finding, overexpressing the SH2 domain of SH2B3 is sufficient to inhibit NGF-induced neurite outgrowth. Together, our data demonstrate that SH2B3, unlike the other two family members, inhibits neuronal differentiation of PC12 cells and primary cortical neurons. Its inhibitory mechanism is likely through the competition of TrkA binding with the positive-acting SH2B1 and SH2B2. PMID:22028877

  6. Strychnine, but not PMBA, inhibits neuronal nicotinic acetylcholine receptors expressed by rabbit retinal ganglion cells.

    Science.gov (United States)

    Renna, J M; Strang, C E; Amthor, F R; Keyser, K T

    2007-01-01

    Strychnine is considered a selective competitive antagonist of glycine gated Cl- channels (Saitoh et al., 1994) and studies have used strychnine at low micromolar concentrations to study the role of glycine in rabbit retina (Linn, 1998; Protti et al., 2005). However, other studies have shown that strychnine, in the concentrations commonly used, is also a potent competitive antagonist of alpha7 nicotinic acetylcholine receptors (nAChRs; Matsubayashi et al., 1998). We tested the effects of low micromolar concentrations of strychnine and 3-[2'-phosphonomethyl[1,1'-biphenyl]-3-yl] alanine (PMBA), a specific glycine receptor blocker (Saitoh et al., 1994; Hosie et al., 1999) on the activation of both alpha7 nAChRs on retinal ganglion cells and on ganglion cell responses to a light flash. Extracellular recordings were obtained from ganglion cells in an isolated retina/choroid preparation and 500 microM choline was used as an alpha7 agonist (Alkondon et al., 1997). We recorded from brisk sustained and brisk transient OFF cells, many of which have been previously shown to have alpha7 receptors (Strang et al., 2005). Further, we tested the effect of strychnine, PMBA and alpha-bungarotoxin on the binding of tetramethylrhodamine alpha-bungarotoxin in the inner plexiform layer. Our data indicates that strychnine, at doses as low as 1.0 microM, can inhibit the alpha7 nAChR-mediated response to choline, but PMBA at concentrations as high as 0.4 microM does not. Binding studies show strychnine and alpha-bungarotoxin inhibit binding of labeled alpha-bungarotoxin in the IPL. Thus, the effects of strychnine application may be to inhibit glycine receptors expressed by ganglion cell or to inhibit amacrine cell alpha7 nAChRs, both of which would result in an increase in the ganglion cell responses. Further research will be required to disentangle the effects of strychnine previously believed to be caused by a single mechanism of glycine receptor inhibition.

  7. Minocycline Promotes Neurite Outgrowth of PC12 Cells Exposed to Oxygen-Glucose Deprivation and Reoxygenation Through Regulation of MLCP/MLC Signaling Pathways.

    Science.gov (United States)

    Tao, Tao; Feng, Jin-Zhou; Xu, Guang-Hui; Fu, Jie; Li, Xiao-Gang; Qin, Xin-Yue

    2017-04-01

    Minocycline, a semi-synthetic second-generation derivative of tetracycline, has been reported to exert neuroprotective effects both in animal models and in clinic trials of neurological diseases. In the present study, we first investigated the protective effects of minocycline on oxygen-glucose deprivation and reoxygenation-induced impairment of neurite outgrowth and its potential mechanism in the neuronal cell line, PC12 cells. We found that minocycline significantly increased cell viability, promoted neurite outgrowth and enhanced the expression of growth-associated protein-43 (GAP-43) in PC12 cells exposed to oxygen-glucose deprivation/reoxygenation injury. In addition, immunoblots revealed that minocycline reversed the overexpression of phosphorylated myosin light chain (MLC) and the suppression of activated extracellular signal-regulated kinase 1/2 (ERK1/2) caused by oxygen-glucose deprivation/reoxygenation injury. Moreover, the minocycline-induced neurite outgrowth was significantly blocked by Calyculin A (1 nM), an inhibitor of myosin light chain phosphatase (MLCP), but not by an ERK1/2 inhibitor (U0126; 10 μM). These findings suggested that minocycline activated the MLCP/MLC signaling pathway in PC12 cells after oxygen-glucose deprivation/reoxygenation injury, which resulted in the promotion of neurite outgrowth.

  8. The output of neuronotrophic and neurite-promoting agents from rat brain astroglial cells: a microculture method for screening potential regulatory molecules.

    Science.gov (United States)

    Rudge, J S; Manthorpe, M; Varon, S

    1985-04-01

    Throughout embryonic development, as well as in response to injury of the central nervous system, astroglial cells may present neurons with a critical supply of neuronotrophic and neurite-promoting factors which control, respectively, neuronal survival and axonal growth. The identification of such astroglial cell-derived factors, as well as of specific extrinsic agents regulating their production, will require the use of in vitro techniques. We define here a new microculture system in which added agents can be screened for their ability to enhance or inhibit the output of trophic and neurite-promoting factors from purified neonatal rat brain astroglial cells. With such a procedure, thousands of replicate secondary astroglial cultures can be set-up and maintained in chemically defined medium, on a defined substratum and in a viable, low proliferative stable state. These cultured astroglial cells release into their medium at least three distinct and separable types of agents addressing nerve cells in vitro: (i) high molecular weight trophic factors (Mr greater than 10,000) which support the survival of embryonic peripheral neurons; (ii) low molecular weight trophic agents (Mr less than 10,000) supporting embryonic central neurons; and (iii) polyornithine-binding neurite-promoting factors which enhance neuritic regeneration for both peripheral and central neurons. The temporal release patterns of these three agents from astroglial cultures are quite distinct suggesting that their output is independently regulated.

  9. Recreational drugs, 3,4-Methylenedioxymethamphetamine(MDMA), 3,4-methylenedioxyamphetamine (MDA) and diphenylprolinol, inhibit neurite outgrowth in PC12 cells.

    Science.gov (United States)

    Kaizaki, Asuka; Tanaka, Sachiko; Tsujikawa, Kenji; Numazawa, Satoshi; Yoshida, Takemi

    2010-06-01

    3,4-Methylenedioxymethamphetamine (MDMA) is widely abused as a psychoactive recreational drug. It is well known that MDMA induces neurotoxic damage of serotonergic nerve endings. Although drug abuse is increasing among youths, it is unclear whether recreational drugs affect the development of nerve growth. Thus, the present study examined the effect of recreational drugs, such as MDMA, 3,4-methylenedioxyamphetamine (MDA) and diphenylprolinol, a novel recreational drug with a similar chemical structure as that of psychoactive agent pipradrol, on nerve growth factor (NGF)-induced neurite outgrowth. These recreational drugs induced a dose-dependent cell death in PC12 cells. The IC(50) values of MDMA, MDA, R-diphenylprolinol and S-diphenylprolinol were 4.11 mM, 2.75 mM, 1.00 mM and 0.77 mM, respectively, at 24 hr. To examine the effects of these recreational drugs on NGF-induced neurite outgrowth, PC12 cells were treated with NGF together with MDMA, MDA, S-diphenylprolinol or R-diphenylprolinol at low toxic concentrations. The recreational drugs significantly suppressed neurite outgrowth of PC12 cells induced by NGF. The results suggest that these psychoactive recreational drugs may inhibit neurite growth and thus be implicated in their elicited neurotoxicity.

  10. Msx2 alters the timing of retinal ganglion cells fate commitment and differentiation

    Energy Technology Data Exchange (ETDEWEB)

    Jiang, Shao-Yun, E-mail: jiangshaoyun@yahoo.com [School of Dentistry, Tianjin Medical University, 12 Qi Xiang Tai Street, Tianjin 300070 (China); Wang, Jian-Tao, E-mail: wangjiantao65@hotmail.com [Eye Center, Tianjin Medical University, 64 Tongan Road, Tianjin 300070 (China); Dohney Eye Institute, Keck School of Medicine, University of Southern California, 1355 San Pablo Street, DOH 314, Los Angeles, CA 90033 (United States)

    2010-05-14

    Timing of cell fate commitment determines distinct retinal cell types, which is believed to be controlled by a tightly coordinated regulatory program of proliferation, cell cycle exit and differentiation. Although homeobox protein Msx2 could induce apoptosis of optic vesicle, it is unclear whether Msx2 regulates differentiation and cell fate commitment of retinal progenitor cells (RPCs) to retinal ganglion cells (RGCs). In this study, we show that overexpression of Msx2 transiently suppressed the expression of Cyclin D1 and blocked cell proliferation. Meanwhile, overexpression of Msx2 delayed the expression of RGC-specific differentiation markers (Math5 and Brn3b), which showed that Msx2 could affect the timing of RGCs fate commitment and differentiation by delaying the timing of cell cycle exit of retinal progenitors. These results indicate Msx2 possesses dual regulatory functions in controlling cell cycle progression of retinal RPCs and timing of RGCs differentiation.

  11. Comparative study of photoreceptor and retinal ganglion cell topography and spatial resolving power in Dipsadidae snakes.

    Science.gov (United States)

    Hauzman, Einat; Bonci, Daniela M O; Grotzner, Sonia R; Mela, Maritana; Liber, André M P; Martins, Sonia L; Ventura, Dora F

    2014-01-01

    The diurnal Dipsadidae snakes Philodryas olfersii and P. patagoniensis are closely related in their phylogeny but inhabit different ecological niches. P. olfersii is arboreal, whereas P. patagoniensis is preferentially terrestrial. The goal of the present study was to compare the density and topography of neurons, photoreceptors, and cells in the ganglion cell layer in the retinas of these two species using immunohistochemistry and Nissl staining procedures and estimate the spatial resolving power of their eyes based on the ganglion cell peak density. Four morphologically distinct types of cones were observed by scanning electron microscopy, 3 of which were labeled with anti-opsin antibodies: large single cones and double cones labeled by the antibody JH492 and small single cones labeled by the antibody JH455. The average densities of photoreceptors and neurons in the ganglion cell layer were similar in both species (∼10,000 and 7,000 cells·mm(-2), respectively). The estimated spatial resolving power was also similar, ranging from 2.4 to 2.7 cycles·degree(-1). However, the distribution of neurons had different specializations. In the arboreal P. olfersii, the isodensity maps had a horizontal visual streak, with a peak density in the central region and a lower density in the dorsal retina. This organization might be relevant for locomotion and hunting behavior in the arboreal layer. In the terrestrial P. patagoniensis, a concentric pattern of decreasing cell density emanated from an area centralis located in the naso-ventral retina. Lower densities were observed in the dorsal region. The ventrally high density improves the resolution in the superior visual field and may be an important adaptation for terrestrial snakes to perceive the approach of predators from above. © 2014 S. Karger AG, Basel.

  12. Spatially and Temporally Regulated NRF2 Gene Therapy Using Mcp-1 Promoter in Retinal Ganglion Cell Injury

    Directory of Open Access Journals (Sweden)

    Kosuke Fujita

    2017-06-01

    Full Text Available Retinal ganglion cell degeneration triggered by axonal injury is believed to underlie many ocular diseases, including glaucoma and optic neuritis. In these diseases, retinal ganglion cells are affected unevenly, both spatially and temporally, such that healthy and unhealthy cells coexist in different patterns at different time points. Herein, we describe a temporally and spatially regulated adeno-associated virus gene therapy aiming to reduce undesired off-target effects on healthy retinal neurons. The Mcp-1 promoter previously shown to be activated in stressed retinal ganglion cells following murine optic nerve injury was combined with the neuroprotective intracellular transcription factor Nrf2. In this model, Mcp-1 promoter-driven NRF2 expression targeting only stressed retinal ganglion cells showed efficacy equivalent to non-selective cytomegalovirus promoter-driven therapy for preventing cell death. However, cytomegalovirus promoter-mediated NRF2 transcription induced cellular stress responses and death of Brn3A-positive uninjured retinal ganglion cells. Such undesired effects were reduced substantially by adopting the Mcp-1 promoter. Combining a stress-responsive promoter and intracellular therapeutic gene is a versatile approach for specifically targeting cells at risk of degeneration. This strategy may be applicable to numerous chronic ocular and non-ocular conditions.

  13. Extracellular vesicles from a muscle cell line (C2C12) enhance cell survival and neurite outgrowth of a motor neuron cell line (NSC-34).

    Science.gov (United States)

    Madison, Roger D; McGee, Christopher; Rawson, Renee; Robinson, Grant A

    2014-01-01

    There is renewed interest in extracellular vesicles over the past decade or 2 after initially being thought of as simple cellular garbage cans to rid cells of unwanted components. Although there has been intense research into the role of extracellular vesicles in the fields of tumour and stem cell biology, the possible role of extracellular vesicles in nerve regeneration is just in its infancy. When a peripheral nerve is damaged, the communication between spinal cord motor neurons and their target muscles is disrupted and the result can be the loss of coordinated muscle movement. Despite state-of-the-art surgical procedures only approximately 10% of adults will recover full function after peripheral nerve repair. To improve upon such results will require a better understanding of the basic mechanisms that influence axon outgrowth and the interplay between the parent motor neuron and the distal end organ of muscle. It has previously been shown that extracellular vesicles are immunologically tolerated, display targeting ligands on their surface, and can be delivered in vivo to selected cell populations. All of these characteristics suggest that extracellular vesicles could play a significant role in nerve regeneration. We have carried out studies using 2 very well characterized cell lines, the C2C12 muscle cell line and the motor neuron cell line NSC-34 to ask the question: Do extracellular vesicles from muscle influence cell survival and/or neurite outgrowth of motor neurons? Our results show striking effects of extracellular vesicles derived from the muscle cell line on the motor neuron cell line in terms of neurite outgrowth and survival.

  14. Central projections of intrinsically photosensitive retinal ganglion cells in the macaque monkey

    DEFF Research Database (Denmark)

    Hannibal, J; Kankipati, L; Strang, C E

    2014-01-01

    Circadian rhythms generated by the suprachiasmatic nucleus (SCN) are entrained to the environmental light/dark cycle via intrinsically photosensitive retinal ganglion cells (ipRGCs) expressing the photopigment melanopsin and the neuropeptide pituitary adenylate cyclase-activating polypeptide (PACAP......-expressing cells characterized as inner and outer stratifying melanopsin RGCs. Two macaque monkeys were anesthetized and received a unilateral intravitreal injection of CtB. Bilateral retinal projections containing colocalized CtB and PACAP immunostaining were identified in the SCN, the lateral geniculate complex...

  15. Senso-Neuroendocrine Cells Within The Cerebral Ganglion Of The Earthworm Aporrectodea Caliginosa

    OpenAIRE

    Al Yousuf, Shoa'a [شعاع السيد اليوسف

    1992-01-01

    Ciliated neurones of probable endocrine function are present inside and outside the cerebral ganglion of the earthworm Aporrectodea caliginosa. Cell fibers contain clusters ofsynaptoid vesicles in regions adjacent to the brain capsule or muscles. The cell bodies also send axons to terminate within the neuropile, where synapses and exocytosis of their secretory granules occur. Cytochemical studies indicate the presence of catecholamines within these granules. باستخدام المجهر الالكتروني النف...

  16. Rgcs1, a dominant QTL that affects retinal ganglion cell death after optic nerve crush in mice

    Directory of Open Access Journals (Sweden)

    Schlamp Cassandra L

    2008-07-01

    Full Text Available Abstract Background Intrinsic apoptosis of neuronal somas is one aspect of neurodegenerative diseases that can be influenced by genetic background. Genes that affect this process may act as susceptibility alleles that contribute to the complex genetic nature of these diseases. Retinal ganglion cell death is a defining feature of the chronic and genetically complex neurodegenerative disease glaucoma. Previous studies using an optic nerve crush procedure in inbred mice, showed that ganglion cell resistance to crush was affected by the Mendelian-dominant inheritance of 1–2 predicted loci. To assess this further, we bred and phenotyped a large population of F2 mice derived from a resistant inbred strain (DBA/2J and a susceptible strain (BALB/cByJ. Results Genome wide mapping of the F2 mice using microsatellite markers, detected a single highly significant quantitative trait locus in a 25 cM (58 Mb interval on chromosome 5 (Chr5.loc34-59 cM. No interacting loci were detected at the resolution of this screen. We have designated this locus as Retinal ganglion cell susceptible 1, Rgcs1. In silico analysis of this region revealed the presence of 578 genes or expressed sequence tags, 4 of which are highly expressed in the ganglion cell layer of the mammalian retina, and 2 of which are suspected susceptibility alleles in chronic neurodegenerative diseases. In addition, 25 genes contain 36 known single nucleotide polymorphisms that create nonsynonymous amino acid changes between the two parental strains. Collectively, this analysis has identified 7 potential candidate genes that may affect ganglion cell death. Conclusion The process of ganglion cell death is likely one of the many facets of glaucoma susceptibility. A novel dominant locus has been identified that affects sensitivity of ganglion cells to optic nerve crush. The allele responsible for this sensitivity may also be a susceptibility allele for glaucoma.

  17. Neuroprotective and axon growth-promoting effects following inflammatory stimulation on mature retinal ganglion cells in mice depend on ciliary neurotrophic factor and leukemia inhibitory factor.

    Science.gov (United States)

    Leibinger, Marco; Müller, Adrienne; Andreadaki, Anastasia; Hauk, Thomas G; Kirsch, Matthias; Fischer, Dietmar

    2009-11-11

    After optic nerve injury retinal ganglion cells (RGCs) normally fail to regenerate axons in the optic nerve and undergo apoptosis. However, lens injury (LI) or intravitreal application of zymosan switch RGCs into an active regenerative state, enabling these neurons to survive axotomy and to regenerate axons into the injured optic nerve. Several factors have been proposed to mediate the beneficial effects of LI. Here, we investigated the contribution of glial-derived ciliary neurotrophic factor (CNTF) to LI-mediated regeneration and neuroprotection using wild-type and CNTF-deficient mice. In wild-type mice, CNTF expression was strongly upregulated in retinal astrocytes, the JAK/STAT3 pathway was activated in RGCs, and RGCs were transformed into an active regenerative state after LI. Interestingly, retinal LIF expression was correlated with CNTF expression after LI. In CNTF-deficient mice, the neuroprotective and axon growth-promoting effects of LI were significantly reduced compared with wild-type animals, despite an observed compensatory upregulation of LIF expression in CNTF-deficient mice. The positive effects of LI and also zymosan were completely abolished in CNTF/LIF double knock-out mice, whereas LI-induced glial and macrophage activation was not compromised. In culture CNTF and LIF markedly stimulated neurite outgrowth of mature RGCs. These data confirm a key role for CNTF in directly mediating the neuroprotective and axon regenerative effects of inflammatory stimulation in the eye and identify LIF as an additional contributing factor.

  18. Cerebellar and basal ganglion involvement in Langerhans cell histiocytosis

    Energy Technology Data Exchange (ETDEWEB)

    Saatci, I.; Baskan, O.; Haliloglu, M.; Aydingoz, U. [Department of Radiology, Hacettepe University Hospital, Sihhiye 06100, Ankara (Turkey)

    1999-06-01

    Langerhans cell histiocytosis (LCH) is a disease of unknown cause characterised by proliferation of histiocytic granulomas in tissues; the primary cerebral manifestation is diabetes insipidus caused by hypothalamic infiltration. We present a patient in whom, except for the absence of high signal on T 1 weighting in the posterior pituitary, consistent with central diabetes insipidus, MRI showed no evidence of hypothalamic involvement by histiocytosis, despite the long duration of the disease. However, there was bilateral, symmetrical involvement of the cerebellum and globus pallidus in addition to a calvarial lesion. High signal in the cerebellar white matter on T 2-weighted images may represent demyelination, gliosis and cell loss, as previously reported on pathologic examination. (orig.) With 5 figs., 22 refs.

  19. Cerebellar and basal ganglion involvement in Langerhans cell histiocytosis.

    Science.gov (United States)

    Saatci, I; Baskan, O; Haliloglu, M; Aydingoz, U

    1999-06-01

    Langerhans cell histiocytosis (LCH) is a disease of unknown cause characterised by proliferation of histiocytic granulomas in tissues; the primary cerebral manifestation is diabetes insipidus caused by hypothalamic infiltration. We present a patient in whom, except for the absence of high signal on T1 weighting in the posterior pituitary, consistent with central diabetes insipidus, MRI showed no evidence of hypothalamic involvement by histiocytosis, despite the long duration of the disease. However, there was bilateral, symmetrical involvement of the cerebellum and globus pallidus in addition to a calvarial lesion. High signal in the cerebellar white matter on T2-weighted images may represent demyelination, gliosis and cell loss, as previously reported on pathologic examination.

  20. Dendritic relationship between starburst amacrine cells and direction-selective ganglion cells in the rabbit retina.

    Science.gov (United States)

    Dong, Wei; Sun, Wenzhi; Zhang, Yingye; Chen, Xiaorong; He, Shigang

    2004-04-01

    We investigated the dendritic relationship between starburst amacrine cells (SAs) and morphologically and physiologically characterized ON and ON-OFF direction-selective ganglion cells (DSGCs) in the rabbit retina. ON and ON-OFF DSGCs were found to exhibit tight dendritic cofasciculation with the SA plexus, visualized by immunolabelling of the vesicular acetylcholine transporter (VAChT). The degree of cofasciculation of both types of DSGC dendrites and SA plexus was found to be significant, unlike the relationship between non-DS cells and the SA plexus, which was close to chance distribution. No difference in the degree of cofasciculation in different regions of the DS dendritic field was observed. Individual SAs intracellularly injected both on the 'preferred' and 'null' side of the DSGCs showed the same degree of cofasciculation with the DSGCs. Therefore, the computation of motion direction is unlikely to result from apparent asymmetry in geometric proximity between SAs and DSGCs. Highly selective synaptic connections between SAs and DSGCs are necessary.

  1. Ganglion Cysts

    Science.gov (United States)

    ... cyst Figure 3 - Cross-section of wrist showing the root of a ganglion cyst PDF Ganglion Cysts Related Conditions Trigger Finger Hand Tumors and Wrist Tumors de Quervain's Tenosynovitis Carpometacarpal Boss Other Links CME Mission Statement and ...

  2. Characterization of BASP1-mediated neurite outgrowth

    DEFF Research Database (Denmark)

    Korshunova, Irina; Caroni, Pico; Kolkova, Kateryna

    2008-01-01

    in PC12E2 cells and primary hippocampal neurons. BASP1 overexpression stimulated neurite outgrowth in both cell types. The effects of BASP1 and trans-homophilic NCAM interactions were additive, and BASP1-induced neurite outgrowth was not inhibited by ectopic expression of cytoplasmic NCAM domains......The brain acid-soluble protein BASP1 (CAP-23, NAP-22) belongs to the family of growth-associated proteins, which also includes GAP-43, a protein recently shown to regulate neural cell adhesion molecule (NCAM)-mediated neurite outgrowth. Here, the effects of BASP1 overexpression were investigated...

  3. Dendritic thickness: a morphometric parameter to classify mouse retinal ganglion cells

    Directory of Open Access Journals (Sweden)

    L.D. Loopuijt

    2007-10-01

    Full Text Available To study the dendritic morphology of retinal ganglion cells in wild-type mice we intracellularly injected these cells with Lucifer yellow in an in vitro preparation of the retina. Subsequently, quantified values of dendritic thickness, number of branching points and level of stratification of 73 Lucifer yellow-filled ganglion cells were analyzed by statistical methods, resulting in a classification into 9 groups. The variables dendritic thickness, number of branching points per cell and level of stratification were independent of each other. Number of branching points and level of stratification were independent of eccentricity, whereas dendritic thickness was positively dependent (r = 0.37 on it. The frequency distribution of dendritic thickness tended to be multimodal, indicating the presence of at least two cell populations composed of neurons with dendritic diameters either smaller or larger than 1.8 µm ("thin" or "thick" dendrites, respectively. Three cells (4.5% were bistratified, having thick dendrites, and the others (95.5% were monostratified. Using k-means cluster analysis, monostratified cells with either thin or thick dendrites were further subdivided according to level of stratification and number of branching points: cells with thin dendrites were divided into 2 groups with outer stratification (0-40% and 2 groups with inner (50-100% stratification, whereas cells with thick dendrites were divided into one group with outer and 3 groups with inner stratification. We postulate, that one group of cells with thin dendrites resembles cat ß-cells, whereas one group of cells with thick dendrites includes cells that resemble cat a-cells.

  4. Modeling activity and target-dependent developmental cell death of mouse retinal ganglion cells ex vivo.

    Directory of Open Access Journals (Sweden)

    Sylvie Voyatzis

    Full Text Available Programmed cell death is widespread during the development of the central nervous system and serves multiple purposes including the establishment of neural connections. In the mouse retina a substantial reduction of retinal ganglion cells (RGCs occurs during the first postnatal week, coinciding with the formation of retinotopic maps in the superior colliculus (SC. We previously established a retino-collicular culture preparation which recapitulates the progressive topographic ordering of RGC projections during early post-natal life. Here, we questioned whether this model could also be suitable to examine the mechanisms underlying developmental cell death of RGCs. Brn3a was used as a marker of the RGCs. A developmental decline in the number of Brn3a-immunolabelled neurons was found in the retinal explant with a timing that paralleled that observed in vivo. In contrast, the density of photoreceptors or of starburst amacrine cells increased, mimicking the evolution of these cell populations in vivo. Blockade of neural activity with tetrodotoxin increased the number of surviving Brn3a-labelled neurons in the retinal explant, as did the increase in target availability when one retinal explant was confronted with 2 or 4 collicular slices. Thus, this ex vivo model reproduces the developmental reduction of RGCs and recapitulates its regulation by neural activity and target availability. It therefore offers a simple way to analyze developmental cell death in this classic system. Using this model, we show that ephrin-A signaling does not participate to the regulation of the Brn3a population size in the retina, indicating that eprhin-A-mediated elimination of exuberant projections does not involve developmental cell death.

  5. Horseradish peroxidase dye tracing and embryonic statoacoustic ganglion cell transplantation in the rat auditory nerve trunk.

    Science.gov (United States)

    Palmgren, Björn; Jin, Zhe; Jiao, Yu; Kostyszyn, Beata; Olivius, Petri

    2011-03-04

    At present severe damage to hair cells and sensory neurons in the inner ear results in non-treatable auditory disorders. Cell implantation is a potential treatment for various neurological disorders and has already been used in clinical practice. In the inner ear, delivery of therapeutic substances including neurotrophic factors and stem cells provide strategies that in the future may ameliorate or restore hearing impairment. In order to describe a surgical auditory nerve trunk approach, in the present paper we injected the neuronal tracer horseradish peroxidase (HRP) into the central part of the nerve by an intra cranial approach. We further evaluated the applicability of the present approach by implanting statoacoustic ganglion (SAG) cells into the same location of the auditory nerve in normal hearing rats or animals deafened by application of β-bungarotoxin to the round window niche. The HRP results illustrate labeling in the cochlear nucleus in the brain stem as well as peripherally in the spiral ganglion neurons in the cochlea. The transplanted SAGs were observed within the auditory nerve trunk but no more peripheral than the CNS-PNS transitional zone. Interestingly, the auditory nerve injection did not impair auditory function, as evidenced by the auditory brainstem response. The present findings illustrate that an auditory nerve trunk approach may well access the entire auditory nerve and does not compromise auditory function. We suggest that such an approach might compose a suitable route for cell transplantation into this sensory cranial nerve. Copyright © 2011 Elsevier B.V. All rights reserved.

  6. Calpain Inhibition Attenuates Apoptosis of Retinal Ganglion Cells in Acute Optic Neuritis

    Science.gov (United States)

    Smith, Amena W.; Das, Arabinda; Guyton, M. Kelly; Ray, Swapan K.; Rohrer, Baerbel

    2011-01-01

    Purpose. Optic neuritis (ON), inflammation of the optic nerve, is strongly associated with the pathogenesis of multiple sclerosis (MS) and is initiated by the attack of autoreactive T cells against self-myelin antigens, resulting in demyelination, degeneration of retinal ganglion cells (RGCs), and cumulative visual impairment. Methods. Experimental autoimmune encephalomyelitis (EAE) was induced in Lewis rats on day 0, and animals received daily intraperitoneal injections of calpain inhibitor (calpeptin) or vehicle from day 1 until killed. Retinal cell death was analyzed by DNA fragmentation, and surviving ganglion cells were quantified after double labeling of retinal tissue with TUNEL and Brn3a. The expression of apoptotic and inflammatory proteins was determined by Western blotting. Results. It was demonstrated that calpain inhibition downregulates expression of proapoptotic proteins and the proinflammatory molecule nuclear factor-kappa B (NF-κB) in the retina of Lewis rats with acute EAE. Immunofluorescent labeling revealed that apoptotic cells in the RGC layer of vehicle-treated EAE animals were Brn3a positive, and a moderate dose of calpeptin dramatically reduced the frequency of apoptotic RGCs. Conclusions. These results suggest that calpain inhibition might be a useful supplement to immunomodulatory therapies such as corticosteroids in ON, due to its neuroprotective effect on RGCs. PMID:21613375

  7. [Progression of nerve fiber layer defects in retrobulbar optic neuritis by the macular ganglion cell complex].

    Science.gov (United States)

    Hong, D; Bosc, C; Chiambaretta, F

    2017-11-01

    Recent studies with SD OCT had shown early axonal damage to the macular ganglion cell complex (which consists of the three innermost layers of the retina: Inner Plexiform Layer [IPL], Ganglion Cell Layer [GCL], Retinal Nerve Fibre layer [RNFL]) in optic nerve pathology. Retrobulbar optic neuritis (RBON), occurring frequently in demyelinating diseases, leads to atrophy of the optic nerve fibers at the level of the ganglion cell axons, previously described in the literature. The goal of this study is to evaluate the progression of optic nerve fiber defects and macular ganglion cell complex defects with the SPECTRALIS OCT via a reproducible method by calculating a mean thickness in each quadrant after an episode of retrobulbar optic neuritis. This is a prospective monocentric observational study including 8 patients at the Clermont-Ferrand university medical center. All patients underwent ocular examination with macular and disc OCT analysis and a Goldmann visual field at the time of inclusion (onset or recurrence of RBON), at 3 months and at 6 months. Patients were 40-years-old on average at the time of inclusion. After 6 months of follow-up, there was progression of the atrophy of the macular ganglion cell complex in the affected eye on (11.5% or 11μm) predominantly inferonasally (13.9% or 16μm) and superonasally (12.9% or 14μm) while the other eye remained stable. The decrease in thickness occurred mainly in the most internal 3 layers of the retina. On average, the loss in thickness of the peripapillary RNFL was predominantly inferotemporal (24.9% or 39μm) and superotemporal (21.8% or 28μm). In 3 months of progression, the loss of optic nerve fibers is already seen on macular and disc OCT after an episode of RBON, especially in inferior quadrants in spite of the improvement in the Goldmann visual field and visual acuity. Segmentation by quadrant was used here to compare the progression of the defect by region compared to the fovea in a global and reproducible

  8. Multicompartment retinal ganglion cells response to high frequency bi-phasic pulse train stimulation: Simulation results.

    Science.gov (United States)

    Maturana, Matias I; Grayden, David B; Burkitt, Anthony N; Meffin, Hamish; Kameneva, Tatiana

    2013-01-01

    Retinal ganglion cells (RGCs) are the sole output neurons of the retina that carry information about a visual scene to the brain. By stimulating RGCs with electrical stimulation, it is possible to elicit a sensation of light for people with macular degeneration or retinitis pigmentosa. To investigate the responses of RGCs to high frequency bi-phasic pulse train stimulation, we use previously constrained models of multi-compartment OFF RGCs. The morphologies of mouse RGCs are taken from the Chalupa set of the NeuroMorpho database. The cell models are divided into compartments representing the dendrites, soma and axon that vary between the cells. A total of 132 cells are simulated in the NEURON environment. Results show that the cell morphology plays an important role in the response characteristics of the cell to high frequency bi-phasic pulse train stimulation.

  9. Neonatal functional intestinal obstruction and the presence of severely immature ganglion cells on rectal biopsy: 6 year experience.

    Science.gov (United States)

    Burki, Tariq; Kiho, Liina; Scheimberg, Irene; Phelps, Simon; Misra, Devesh; Ward, Harry; Colmenero, Isabel

    2011-05-01

    We report our experience of managing eight babies who presented with neonatal intestinal obstruction and whose rectal biopsies showed severely immature ganglion cells. Neonatal unit records were reviewed to detect patients with suspected Hirschsprung's disease or functional intestinal obstruction. Those with intestinal atresia, anorectal malformation, malrotation, cystic fibrosis and prematurity were excluded. We identified 73 patients born at term. Twenty-seven did not need a rectal biopsy. Twenty-one had biopsy proven Hirschsprung's disease, while 17 had a normal rectal biopsy. Eight patients, all of whom presented with severe abdominal distension, showed immature ganglion cells. Seven had failed to pass meconium after birth. X-rays in all patients showed distended loops of bowel. Two neonates underwent an emergency laparotomy and a stoma. A repeat biopsy at 3 months showed maturation of ganglion cells and the stoma was reversed. Rectal biopsy was repeated in two other patients 2-9 months after the first biopsy and showed mature ganglion cells. At follow-up, one patient still suffers from severe constipation. Seven are asymptomatic now, including the two patients who needed a stoma. Immature ganglion cells on rectal biopsy may be an indicator of transient functional immaturity of the intestine.

  10. Disassembly of microtubules and inhibition of neurite outgrowth, neuroblastoma cell proliferation, and MAP kinase tyrosine dephosphorylation by dibenzyl trisulphide.

    Science.gov (United States)

    Rösner, H; Williams, L A; Jung, A; Kraus, W

    2001-08-22

    Dibenzyl trisulphide (DTS), a main lipophilic compound in Petiveria alliacea L. (Phytolaccaceae), was identified as one of the active immunomodulatory compounds in extracts of the plant. To learn more about its biological activities and molecular mechanisms, we conducted one-dimensional NMR interaction studies with bovine serum albumin (BSA) and tested DTS and related compounds in two well-established neuronal cell-and-tissue culture systems. We found that DTS preferentially binds to an aromatic region of BSA which is rich in tyrosyl residues. In SH-SY5Y neuroblastoma cells, DTS attenuates the dephosphorylation of tyrosyl residues of MAP kinase (erk1/erk2). In the same neuroblastoma cell line and in Wistar 38 human lung fibroblasts, DTS causes a reversible disassembly of microtubules, but it did not affect actin dynamics. Probably due to the disruption of the microtubule dynamics, DTS also inhibits neuroblastoma cell proliferation and neurite outgrowth from spinal cord explants. Related dibenzyl compounds with none, one, or two sulphur atoms were found to be significantly less effective. These data confirmed that the natural compound DTS has a diverse spectrum of biological properties, including cytostatic and neurotoxic actions in addition to immunomodulatory activities.

  11. Cellular Origin of Spontaneous Ganglion Cell Spike Activity in Animal Models of Retinitis Pigmentosa

    Directory of Open Access Journals (Sweden)

    David J. Margolis

    2011-01-01

    Full Text Available Here we review evidence that loss of photoreceptors due to degenerative retinal disease causes an increase in the rate of spontaneous ganglion spike discharge. Information about persistent spike activity is important since it is expected to add noise to the communication between the eye and the brain and thus impact the design and effective use of retinal prosthetics for restoring visual function in patients blinded by disease. Patch-clamp recordings from identified types of ON and OFF retinal ganglion cells in the adult (36–210 d old rd1 mouse show that the ongoing oscillatory spike activity in both cell types is driven by strong rhythmic synaptic input from presynaptic neurons that is blocked by CNQX. The recurrent synaptic activity may arise in a negative feedback loop between a bipolar cell and an amacrine cell that exhibits resonant behavior and oscillations in membrane potential when the normal balance between excitation and inhibition is disrupted by the absence of photoreceptor input.

  12. Investigation of retinal ganglion cells and axons of normal rats using fluorogold retrograde labeling

    International Nuclear Information System (INIS)

    Yin Xiaolei; Ye Jian; Chen Chunlin

    2006-01-01

    To investigate the retinal ganglion cells (RGCs) by means of fluorogold retrograde labeling, RGCs were labeled by injecting the fluorogold bilaterally into the superficial superior colliculus and lateral genicutate nucleus in six adult SD rats. One and two weeks (3 rats in each group) after injecting the fluorogold, RGCs FG-labeled were observed and the number of them were counted. The results showed that after a week mean density of fluorogold-labeled RGCs was 2210 ± 128/mm 2 , and it was 2164 ± 117/mm 2 after two weeks. Our conclusion is fluorogold retrograde labeling could be very useful in the research of RGCs. (authors)

  13. A Pixel-Encoder Retinal Ganglion Cell with Spatially Offset Excitatory and Inhibitory Receptive Fields

    OpenAIRE

    Keith P. Johnson; Lei Zhao; Daniel Kerschensteiner

    2018-01-01

    The spike trains of retinal ganglion cells (RGCs) are the only source of visual information to the brain. Here, we genetically identify an RGC type in mice that functions as a pixel encoder and increases firing to light increments (PixON-RGC). PixON-RGCs have medium-sized dendritic arbors and non-canonical center-surround receptive fields. From their receptive field center, PixON-RGCs receive only excitatory input, which encodes contrast and spatial information linearly. From their receptive ...

  14. Neural architecture of the "transient" ON directionally selective (class IIb1) ganglion cells in rabbit retina, partly co-stratified with starburst amacrine cells.

    Science.gov (United States)

    Famiglietti, Edward V

    2016-01-01

    Recent physiological studies coupled with intracellular staining have subdivided ON directionally selective (DS) ganglion cells of rabbit retina into two types. One exhibits more "transient" and more "brisk" responses (ON DS-t), and the other has more "sustained' and more "sluggish" responses (ON DS-s), although both represent the same three preferred directions and show preference for low stimulus velocity, as reported in previous studies of ON DS ganglion cells in rabbit retina. ON DS-s cells have the morphology of ganglion cells previously shown to project to the medial terminal nucleus (MTN) of the accessory optic system, and the MTN-projecting, class IVus1 cells have been well-characterized previously in terms of their dendritic morphology, branching pattern, and stratification. ON DS-t ganglion cells have a distinctly different morphology and exhibit heterotypic coupling to amacrine cells, including axon-bearing amacrine cells, with accompanying synchronous firing, while ON DS-s cells are not coupled. The present study shows that ON DS-t cells are morphologically identical to the previously well-characterized, "orphan" class IIb1 ganglion cell, previously regarded as a member of the "brisk-concentric" category of ganglion cells. Its branching pattern, quantitatively analyzed, is similar to that of the morphological counterparts of X and Y cells, and very different from that of the ON DS-s ganglion cell. Close analysis of the dendritic stratification of class IIb1 ganglion cells together with fiducial cells indicates that they differ from that of the ON DS-s cells. In agreement with one of the three previous studies, class IIb1/ON DS-t cells, unlike class IVus1/ON DS-s ganglion cells, in the main do not co-stratify with starburst amacrine cells. As the present study shows, however, portions of their dendrites do deviate from the main substratum, coming within range of starburst boutons. Parsimony favors DS input from starburst amacrine cells both to ON DS

  15. Target genes involved in corticosterone-induced PC12 cell viability and neurite disorders: A potential molecular mechanism of major depressive disorder.

    Science.gov (United States)

    Li, Mingzhen; Zhou, Jianjun; Qian, Jialin; Cheng, Xiaoyan; Wu, Huijuan; Li, Li; Qian, Chunyan; Su, Joyce; Wu, Donald; Burns, Larry; Golden, Teresa; Wu, Ning

    2016-01-30

    Past studies confirmed that hypothalamic-pituitary-adrenal (HPA)-axis hormones involved in major depressive disorder (MDD) development. This study used corticosterone treated PC12 cells to explore the potential role of MAPK signal transduction pathway in response to corticosterone stimulation. The results showed that both live cell numbers and cellular neurite outgrowth were remarkably reduced in response to corticosterone treatments. qPCR results demonstrated that the expression levels of four MAPK pathway genes were significantly increased after corticosterone stimulation. In conclusion, glucocorticoids stimulation can affect neuronal cell viability and neurite outgrowth due to the over expression of a group of MAPK pathway genes. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

  16. A Novel Retinal Ganglion Cell Promoter for Utility in AAV Vectors

    Directory of Open Access Journals (Sweden)

    Killian S. Hanlon

    2017-09-01

    Full Text Available Significant advances in gene therapy have enabled exploration of therapies for inherited retinal disorders, many of which are in preclinical development or clinical evaluation. Gene therapy for retinal conditions has led the way in this growing field. The loss of retinal ganglion cells (RGCs is a hallmark of a number of retinal disorders. As the field matures innovations that aid in refining therapies and optimizing efficacy are in demand. Gene therapies under development for RGC-related disorders, when delivered with recombinant adeno associated vectors (AAV, have typically been expressed from ubiquitous promoter sequences. Here we describe how a novel promoter from the murine Nefh gene was selected to drive transgene expression in RGCs. The Nefh promoter, in an AAV2/2 vector, was shown to drive preferential EGFP expression in murine RGCs in vivo following intravitreal injection. In contrast, EGFP expression from a CMV promoter was observed not only in RGCs, but throughout the inner nuclear layer and in amacrine cells located within the ganglion cell layer (GCL. Of note, the Nefh promoter sequence is sufficiently compact to be readily accommodated in AAV vectors, where transgene size represents a significant constraint. Moreover, this promoter should in principle provide a more targeted and potentially safer alternative for RGC-directed gene therapies.

  17. Autophagy in retinal ganglion cells in a rhesus monkey chronic hypertensive glaucoma model.

    Directory of Open Access Journals (Sweden)

    Shuifeng Deng

    Full Text Available Primary open angle glaucoma (POAG is a neurodegenerative disease characterized by physiological intraocular hypertension that causes damage to the retinal ganglion cells (RGCs. In the past, RGC damage in POAG was suggested to have been attributed to RGC apoptosis. However, in the present study, we applied a model closer to human POAG through the use of a chronic hypertensive glaucoma model in rhesus monkeys to investigate whether another mode of progressive cell death, autophagy, was activated in the glaucomatous retinas. First, in the glaucomatous retinas, the levels of LC3B-II, LC3B-II/LC3B-I and Beclin 1 increased as demonstrated by Western blot analyses, whereas early or initial autophagic vacuoles (AVi and late or degraded autophagic vacuoles (AVd accumulated in the ganglion cell layer (GCL and in the inner plexiform layer (IPL as determined by transmission electron microscopy (TEM analysis. Second, lysosome activity and autophagosome-lysosomal fusion increased in the RGCs of the glaucomatous retinas, as demonstrated by Western blotting against lysosome associated membrane protein-1 (LAMP1 and double labeling against LC3B and LAMP1. Third, apoptosis was activated in the glaucomatous eyes with increased levels of caspase-3 and cleaved caspase-3 and an increased number of TUNEL-positive RGCs. Our results suggested that autophagy was activated in RGCs in the chronic hypertensive glaucoma model of rhesus monkeys and that autophagy may have potential as a new target for intervention in glaucoma treatment.

  18. Glial cell line-derived neurotrophic factor (GDNF) enhances sympathetic neurite growth in rat hearts at early developmental stages

    NARCIS (Netherlands)

    Miwa, Keiko; Lee, Jong-Kook; Takagishi, Yoshiko; Opthof, Tobias; Fu, Xianming; Kodama, Itsuo

    2010-01-01

    Molecular signaling of sympathetic innervation of myocardium is an unresolved issue. The purpose of this study was to investigate the effect of neurotrophic factors on sympathetic neurite growth towards cardiomyocytes. Cardiomyocytes (CMs) and sympathetic neurons (SNs) were isolated from neonatal

  19. Primary culture of glial cells from mouse sympathetic cervical ganglion: a valuable tool for studying glial cell biology.

    Science.gov (United States)

    de Almeida-Leite, Camila Megale; Arantes, Rosa Maria Esteves

    2010-12-15

    Central nervous system glial cells as astrocytes and microglia have been investigated in vitro and many intracellular pathways have been clarified upon various stimuli. Peripheral glial cells, however, are not as deeply investigated in vitro despite its importance role in inflammatory and neurodegenerative diseases. Based on our previous experience of culturing neuronal cells, our objective was to standardize and morphologically characterize a primary culture of mouse superior cervical ganglion glial cells in order to obtain a useful tool to study peripheral glial cell biology. Superior cervical ganglia from neonatal C57BL6 mice were enzymatically and mechanically dissociated and cells were plated on diluted Matrigel coated wells in a final concentration of 10,000cells/well. Five to 8 days post plating, glial cell cultures were fixed for morphological and immunocytochemical characterization. Glial cells showed a flat and irregular shape, two or three long cytoplasm processes, and round, oval or long shaped nuclei, with regular outline. Cell proliferation and mitosis were detected both qualitative and quantitatively. Glial cells were able to maintain their phenotype in our culture model including immunoreactivity against glial cell marker GFAP. This is the first description of immunocytochemical characterization of mouse sympathetic cervical ganglion glial cells in primary culture. This work discusses the uses and limitations of our model as a tool to study many aspects of peripheral glial cell biology. Copyright © 2010 Elsevier B.V. All rights reserved.

  20. The RNA binding protein RBPMS is a selective marker of ganglion cells in the mammalian retina

    Science.gov (United States)

    Rodriguez, Allen R.; de Sevilla Müller, Luis Pérez; Brecha, Nicholas C.

    2014-01-01

    There are few neurochemical markers that reliably identify retinal ganglion cells (RGCs), which are a heterogeneous population of cells that integrate and transmit the visual signal from the retina to the central visual nuclei. We have developed and characterized a new set of affinity purified guinea pig and rabbit antibodies against RNA-binding protein with multiple splicing (RBPMS). On Western blots these antibodies recognize a single band at ~24 kDa, corresponding to RBPMS, and they strongly label RGC and displaced RGC (dRGC) somata in mouse, rat, guinea pig, rabbit and monkey retina. RBPMS immunoreactive cells and RGCs identified by other techniques have a similar range of somal diameters and areas. The density of RBPMS cells in mouse and rat retina is comparable to earlier semi-quantitative estimates of RGCs. RBPMS is mainly expressed in medium and large DAPI-, DRAQ5-, NeuroTrace- and NeuN-stained cells in the ganglion cell layer (GCL), and RBPMS is not expressed in syntaxin (HPC-1) immunoreactive cells in the inner nuclear layer (INL) and GCL, consistent with their identity as RGCs, and not displaced amacrine cells. In mouse and rat retina, most RBPMS cells are lost following optic nerve crush or transection at three weeks, and all Brn3a, SMI-32 and melanopsin immunoreactive RGCs also express RBPMS immunoreactivity. RBPMS immunoreactivity is localized to CFP-fluorescent RGCs in the B6.Cg-Tg(Thy1-CFP)23Jrs/J mouse line. These findings show that antibodies against RBPMS are robust reagents that exclusively identify RGCs and dRGCs in multiple mammalian species, and they will be especially useful for quantification of RGCs. PMID:24318667

  1. Expression of polysialylated neural cell adhesion molecules on adult stem cells after neuronal differentiation of inner ear spiral ganglion neurons

    Energy Technology Data Exchange (ETDEWEB)

    Park, Kyoung Ho [Department of Otolaryngology Head and Neck Surgery, College of Medicine, Catholic University, Seoul (Korea, Republic of); Yeo, Sang Won, E-mail: swyeo@catholic.ac.kr [Department of Otolaryngology Head and Neck Surgery, College of Medicine, Catholic University, Seoul (Korea, Republic of); Troy, Frederic A., E-mail: fatroy@ucdavis.edu [Department of Biochemistry and Molecular Medicine, University of California, School of Medicine, Davis, CA 95616 (United States); Xiamen University, School of Medicine, Xiamen City (China)

    2014-10-17

    Highlights: • PolySia expressed on neurons primarily during early stages of neuronal development. • PolySia–NCAM is expressed on neural stem cells from adult guinea pig spiral ganglion. • PolySia is a biomarker that modulates neuronal differentiation in inner ear stem cells. - Abstract: During brain development, polysialylated (polySia) neural cell adhesion molecules (polySia–NCAMs) modulate cell–cell adhesive interactions involved in synaptogenesis, neural plasticity, myelination, and neural stem cell (NSC) proliferation and differentiation. Our findings show that polySia–NCAM is expressed on NSC isolated from adult guinea pig spiral ganglion (GPSG), and in neurons and Schwann cells after differentiation of the NSC with epidermal, glia, fibroblast growth factors (GFs) and neurotrophins. These differentiated cells were immunoreactive with mAb’s to polySia, NCAM, β-III tubulin, nestin, S-100 and stained with BrdU. NSC could regenerate and be differentiated into neurons and Schwann cells. We conclude: (1) polySia is expressed on NSC isolated from adult GPSG and on neurons and Schwann cells differentiated from these NSC; (2) polySia is expressed on neurons primarily during the early stage of neuronal development and is expressed on Schwann cells at points of cell–cell contact; (3) polySia is a functional biomarker that modulates neuronal differentiation in inner ear stem cells. These new findings suggest that replacement of defective cells in the inner ear of hearing impaired patients using adult spiral ganglion neurons may offer potential hope to improve the quality of life for patients with auditory dysfunction and impaired hearing disorders.

  2. Glutamate release from satellite glial cells of the murine trigeminal ganglion.

    Science.gov (United States)

    Wagner, Lysann; Warwick, Rebekah A; Pannicke, Thomas; Reichenbach, Andreas; Grosche, Antje; Hanani, Menachem

    2014-08-22

    It has been proposed that glutamate serves as a mediator between neurons and satellite glial cells (SGCs) in sensory ganglia and that SGCs release glutamate. Using a novel method, we studied glutamate release from SGCs from murine trigeminal ganglia. Sensory neurons with adhering SGCs were enzymatically isolated from wild type and transgenic mice in which vesicular exocytosis was suppressed in glial cells. Extracellular glutamate was detected by microfluorimetry. After loading the cells with a photolabile Ca(2+) chelator, the intracellular Ca(2+) concentration was raised in SGCs by a UV pulse, which resulted in glutamate release. The amount of released glutamate was decreased in cells with suppressed exocytosis and after pharmacological block of hemichannels. The data demonstrate that SGCs of the trigeminal ganglion release glutamate in a Ca(2+)-dependent manner. Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.

  3. Piperine-like alkamides from Piper nigrum induce BDNF promoter and promote neurite outgrowth in Neuro-2a cells.

    Science.gov (United States)

    Yun, Young Sook; Noda, Sachie; Takahashi, Shigeru; Takahashi, Yuji; Inoue, Hideshi

    2018-01-01

    Black pepper (Piper nigrum) contains a variety of alkamides. Among them, piperine has been reported to have antidepressant-like effects in chronically stressed mice, but little is known about the biological activity of other alkamides. In this study, we investigated the effects of alkamides from white pepper (P. nigrum) on neuronal cells. Twelve alkamides were isolated from white pepper MeOH extracts, and their chemical structures were identified by NMR and MS analyses. The compounds were subjected to assays using the luciferase-reporter gene under the control of the BDNF promoter or cAMP response element in mouse neuroblastoma Neuro-2a cells. In both assays, marked reporter-inducing activity was observed for piperine (1), piperettine (2) and piperylin (7), all of which have in common an (E)-5-(buta-1,3-dien-1-yl)benzo[d] [1, 3] dioxole moiety. Piperettine (2) and piperylin (7) tended to increase endogenous BDNF protein levels. Furthermore, piperylin (7) promoted retinoic acid-induced neurite outgrowth. These results suggest that piperylin (7), or analogues thereof, may have a beneficial effect on disorders associated with dysregulation of BDNF expression, such as depression.

  4. Vesicouterine ligament contains abundant autonomic nerve ganglion cells: the distribution in histology concerning nerve-sparing radical hysterectomy.

    Science.gov (United States)

    Katahira, A; Niikura, H; Ito, K; Takano, T; Nagase, S; Murakami, G; Yaegashi, N

    2008-01-01

    The aim of this study is to describe the histologic architecture of the tissues corresponding to the surgically developed connective tissue bundle commonly referred to as the posterior leaf of the vesico-uterine ligament (VUL), and to examine distribution of ganglion cells. Serial macroscopic slices, each 15-20 mm in thickness, were made from eight specimens (obtained from six female elderly cadavers). In these macroslices, the location of the deep uterine vein was used to identify the deep leaf of the VUL. The specimens were trimmed and semi-serial histologic sections in thickness were prepared at 1 mm intervals. Vesical veins and the associated nerve elements were enclosed by fascia and formed a common pedicle. The base of the pedicle contained the deep uterine vein trunk. The fascia encircling the pedicle varied in thickness and connective intensity between specimens. This vesical neurovascular bundle contained abundant ganglion cells. On average, 48.0% of the ganglion cells along the vesical tributaries of the deep uterine vein were located on the medial or vaginal side of the veins, 19.2% were located between veins, 13.0% on the lateral side of the veins, and 19.8% on the dorsal side. The interindividual variability was greatest on the dorsal side of vesical veins and ranged 11-202 cells. We conclude that in order to achieve maximal preservation of the ganglion cells during the surgical dissection of the posterior leaf of the VUL, care must be taken when the medial or vesical aspect of the ligament is separated. The standard nerve-sparing radical hysterectomy should be modified to reflect differences in the distribution of ganglion cells and in connective intensity between ganglions and veins.

  5. Retinal vessel diameters decrease with macular ganglion cell layer thickness in autosomal dominant optic atrophy and in healthy subjects

    DEFF Research Database (Denmark)

    Rönnbäck, Cecilia; Grønskov, Karen; Larsen, Michael

    2014-01-01

    diameters (central retinal artery equivalent, CRAE, and central retinal vein equivalent, CRVE). Statistical analysis was corrected for age, gender, spherical equivalent refraction, axial length and mean arterial blood pressure (MABP) in a mixed model analysis. RESULTS: Retinal arteries and veins were...... ganglion cell-inner plexiform layer (GC-IPL) thickness (p = 0.0017 and p = 0.0057, respectively). CONCLUSION: Narrow retinal arteries and veins were associated not only with the severity of ADOA but with ganglion cell volume in patients with ADOA and in healthy subjects. This suggests that narrow vessels...

  6. A role for complexes of survival of motor neurons (SMN) protein with gemins and profilin in neurite-like cytoplasmic extensions of cultured nerve cells

    International Nuclear Information System (INIS)

    Sharma, Aarti; Lambrechts, Anja; Le thi Hao; Le, Thanh T.; Sewry, Caroline A.; Ampe, Christophe; Burghes, Arthur H.M.; Morris, Glenn E.

    2005-01-01

    Spinal muscular atrophy (SMA) is caused by reduced levels of SMN (survival of motor neurons protein) and consequent loss of motor neurons. SMN is involved in snRNP transport and nuclear RNA splicing, but axonal transport of SMN has also been shown to occur in motor neurons. SMN also binds to the small actin-binding protein, profilin. We now show that SMN and profilin II co-localise in the cytoplasm of differentiating rat PC12 cells and in neurite-like extensions, especially at their growth cones. Many components of known SMN complexes were also found in these extensions, including gemin2 (SIP-1), gemin6, gemin7 and unrip (unr-interacting protein). Coilin p80 and Sm core protein immunoreactivity, however, were seen only in the nucleus. SMN is known to associate with β-actin mRNA and specific hnRNPs in axons and in neurite extensions of cultured nerve cells, and SMN also stimulates neurite outgrowth in cultures. Our results are therefore consistent with SMN complexes, rather than SMN alone, being involved in the transport of actin mRNPs along the axon as in the transport of snRNPs into the nucleus by similar SMN complexes. Antisense knockdown of profilin I and II isoforms inhibited neurite outgrowth of PC12 cells and caused accumulation of SMN and its associated proteins in cytoplasmic aggregates. BIAcore studies demonstrated a high affinity interaction of SMN with profilin IIa, the isoform present in developing neurons. Pathogenic missense mutations in SMN, or deletion of exons 5 and 7, prevented this interaction. The interaction is functional in that SMN can modulate actin polymerisation in vitro by reducing the inhibitory effect of profilin IIa. This suggests that reduced SMN in SMA might cause axonal pathfinding defects by disturbing the normal regulation of microfilament growth by profilins

  7. Increased population of immature enteric glial cells in the resected proximal ganglionic bowel of Hirschsprung's disease patients.

    Science.gov (United States)

    Tani, Gakuto; Tomuschat, Christian; O'Donnell, Anne Marie; Coyle, David; Puri, Prem

    2017-10-01

    Enteric glial cells are essential for normal gastrointestinal function. Abnormalities in glial structure, development, or function lead to disturbances in gastrointestinal physiology. Fatty acid-binding protein 7 (FABP7) is a marker of immature enteric glial cells, whereas S100 is expressed only by mature glial cells. Patients with Hirschsprung's disease (HSCR) often suffer from dysmotility and enterocolitis despite proper surgery. We designed this study to determine the distribution and expression of glial cells in patients with HSCR compared to normal controls. We investigated FABP7, S100, and PGP 9.5 expressions in both the ganglionic and aganglionic bowel of patients with HSCR (n = 6) versus normal control colon (n = 6). Protein distribution was assessed by using immunofluorescence and confocal microscopy. Gene and protein expressions were quantified using quantitative real-time polymerase chain reaction (qPCR), Western blot analysis, and densitometry. qPCR and Western blot analysis demonstrated a significantly increased FABP7 expression in ganglionic specimens compared to control specimen (P cells lie under the colonic epithelium and in close apposition to enteric neurons in the ganglionic bowel. The significantly increased number of immature enteric glial cells (EGCs) in the ganglionic bowel of HSCR patients may have adverse effect on the function of enteric neurons and intestinal barrier and thus predispose these patients to intestinal motility problems and enterocolitis. Copyright © 2017 Elsevier Inc. All rights reserved.

  8. The human trigeminal ganglion: c-kit positive neurons and interstitial cells.

    Science.gov (United States)

    Rusu, M C; Pop, F; Hostiuc, S; Dermengiu, D; Lală, A I; Ion, D A; Mănoiu, V S; Mirancea, N

    2011-10-20

    The presence of c-kit positive neurons in sensory ganglia has been verified in various species but not in humans. Our aim has been to identify whether human primary trigeminal neurons label with c-kit/CD117 and thus, whether data gathered in animal studies can be extrapolated to humans. We also intended to establish whether, and which non-neuronal cells also label with c-kit in the trigeminal ganglion. Human adult trigeminal ganglia from eight cadavers were processed for immunohistochemistry on paraffin embedded samples using monoclonal antibodies for CD117/c-kit, and three additional trigeminal ganglia were used for transmission electron microscopy (TEM). To evaluate which neuronal type (A or B) was labeled with c-kit, we evaluated the same neurons on adjacent sections labeled with antibodies for neurofilaments (NF). c-kit has labeled trigeminal neurons (TNs), mast cells and interstitial cells (ICs) within the trigeminal ganglion. c-kit+TNs were NF-and thus were strongly presumed to be nociceptive, as such neurons are known to be NF-poor. c-kit+ICs with long and moniliform processes intermingled with the satellite glial cells (SGCs) of the neuronal envelopes. TEM evaluations confirmed this mixed composition of the neuronal envelopes and demonstrated that the perineuronal ICs are in fact interstitial Cajal-like cells (ICLCs) and/or telocytes. c-kit+TNs were objectified in humans and strongly presumed to be nociceptive. TNs envelopes mostly consist of SGCs, but are also combined with ICLCs/telocytes. Copyright © 2011 Elsevier GmbH. All rights reserved.

  9. Transglial transmission at the dorsal root ganglion sandwich synapse: glial cell to postsynaptic neuron communication.

    Science.gov (United States)

    Rozanski, Gabriela M; Li, Qi; Stanley, Elise F

    2013-04-01

    The dorsal root ganglion (DRG) contains a subset of closely-apposed neuronal somata (NS) separated solely by a thin satellite glial cell (SGC) membrane septum to form an NS-glial cell-NS trimer. We recently reported that stimulation of one NS with an impulse train triggers a delayed, noisy and long-lasting response in its NS pair via a transglial signaling pathway that we term a 'sandwich synapse' (SS). Transmission could be unidirectional or bidirectional and facilitated in response to a second stimulus train. We have shown that in chick or rat SS the NS-to-SGC leg of the two-synapse pathway is purinergic via P2Y2 receptors but the second SGC-to-NS synapse mechanism remained unknown. A noisy evoked current in the target neuron, a reversal potential close to 0 mV, and insensitivity to calcium scavengers or G protein block favored an ionotropic postsynaptic receptor. Selective block by D-2-amino-5-phosphonopentanoate (AP5) implicated glutamatergic transmission via N-methyl-d-aspartate receptors. This agent also blocked NS responses evoked by puff of UTP, a P2Y2 agonist, directly onto the SGC cell, confirming its action at the second synapse of the SS transmission pathway. The N-methyl-d-aspartate receptor NR2B subunit was implicated by block of transmission with ifenprodil and by its immunocytochemical localization to the NS membrane, abutting the glial septum P2Y2 receptor. Isolated DRG cell clusters exhibited daisy-chain and branching NS-glial cell-NS contacts, suggestive of a network organization within the ganglion. The identification of the glial-to-neuron transmitter and receptor combination provides further support for transglial transmission and completes the DRG SS molecular transmission pathway. © 2013 Federation of European Neuroscience Societies and Blackwell Publishing Ltd.

  10. Atoh7 promotes the differentiation of Müller cells-derived retinal stem cells into retinal ganglion cells in a rat model of glaucoma.

    Science.gov (United States)

    Song, Wei-tao; Zhang, Xue-yong; Xia, Xiao-bo

    2015-05-01

    Glaucoma is one of the leading eye diseases resulting in blindness due to the death of retinal ganglion cells. This study aimed to develop novel protocol to promote the differentiation of retinal Müller cells into ganglion cells in vivo in a rat model of glaucoma. The stem cells dedifferentiated from rat retinal Müller cells were randomized to receive transfection with empty lentivirus PGC-FU-GFP or lentivirus PGC-FU-Atoh7-GFP, or no transfection. The stem cells were induced further to differentiate. Ocular hypertension was induced using laser photocoagulation. The eyes were injected with Atoh7 expression vector lentivirus PGC-FU-Atoh7-GFP. Eyeball frozen sections, immunohistochemistry, RT-PCR, Western bolt, and apoptosis assay were performed. We found that the proportion of ganglion cells differentiated from Atoh7-tranfected stem cells was significantly higher than that of the other two groups. The mean intraocular pressure of glaucomatous eyes was elevated significantly compared with those of contralateral eyes. Some retinal Müller cells in the inner nuclear layer entered the mitotic cell cycle in rat chronic ocular hypertension glaucoma model. Atoh7 contributes to the differentiation of retinal Müller cells into retinal ganglion cells in rat model of glaucoma. In conclusion, Atoh7 promotes the differentiation of Müller cells-derived retinal stem cells into retinal ganglion cells in a rat model of glaucoma, thus opening up a new avenue for gene therapy and optic nerve regeneration in glaucoma. © 2015 by the Society for Experimental Biology and Medicine.

  11. The trophic effect of ouabain on retinal ganglion cells is mediated by IL-1β and TNF-α

    International Nuclear Information System (INIS)

    Salles von-Held-Ventura, Juliana; Mázala-de-Oliveira, Thalita; Cândida da Rocha Oliveira, Amanda; Granja, Marcelo Gomes; Gonçalves-de-Albuquerque, Cassiano Felippe; Castro-Faria-Neto, Hugo Caire; Giestal-de-Araujo, Elizabeth

    2016-01-01

    Ouabain is a steroid hormone that binds to the enzyme Na + , K + – ATPase and stimulates different intracellular pathways controlling growth, proliferation and cell survival. IL-1β and TNF-α are pleiotropic molecules, conventionally regarded as pro-inflammatory cytokines with well-known effects in the immune system. In addition, IL-1β and TNF-α also play important roles in the nervous system including neuroprotective effects. Previous data from our group showed that ouabain treatment is able to induce an increase in retinal ganglion cell survival kept in mixed retinal cell cultures. The aim of this work was to investigate if IL-1β and TNF-α could be mediating the trophic effect of ouabain on retinal ganglion cells. Our results show that the trophic effect of ouabain on retinal ganglion cell was inhibited by either anti-IL-1β or anti-TNF-α antibodies. In agreement, IL-1β or TNF-α increased the retinal ganglion cells survival in a dose-dependent manner. Accordingly, ouabain treatment induces a temporal release of TNF-α and IL-1β from retinal cell cultures. Interestingly, TNF-α and IL-1β regulate each other intracellular levels. Our results suggest that ouabain treatment triggers the activation of TNF-α and IL-1β signaling pathways leading to an increase in retinal ganglion cell survival. - Highlights: • Pro-inflammatory cytokines regulates the ouabain effect on RGC survival. • Ouabain treatment modulates the intracellular levels of TNF-α and IL-1β. • Ouabain induces the release of TNF-α and IL-1β in retinal cell cultures.

  12. The trophic effect of ouabain on retinal ganglion cells is mediated by IL-1β and TNF-α

    Energy Technology Data Exchange (ETDEWEB)

    Salles von-Held-Ventura, Juliana; Mázala-de-Oliveira, Thalita; Cândida da Rocha Oliveira, Amanda; Granja, Marcelo Gomes [Departamento de Neurobiologia, Programa de Neurociências, Outeiro de São João Batista s/n CEP: 24020-150, Universidade Federal Fluminense, Niterói, RJ (Brazil); Gonçalves-de-Albuquerque, Cassiano Felippe; Castro-Faria-Neto, Hugo Caire [Fundação Oswaldo Cruz, Instituto Oswaldo Cruz, Departamento de Fisiologia e Farmacodinâmica, Av., no 4365, Manguinhos, 21045-900, Rio de Janeiro, RJ (Brazil); Giestal-de-Araujo, Elizabeth, E-mail: egiestal@vm.uff.br [Departamento de Neurobiologia, Programa de Neurociências, Outeiro de São João Batista s/n CEP: 24020-150, Universidade Federal Fluminense, Niterói, RJ (Brazil)

    2016-09-09

    Ouabain is a steroid hormone that binds to the enzyme Na{sup +}, K{sup +} – ATPase and stimulates different intracellular pathways controlling growth, proliferation and cell survival. IL-1β and TNF-α are pleiotropic molecules, conventionally regarded as pro-inflammatory cytokines with well-known effects in the immune system. In addition, IL-1β and TNF-α also play important roles in the nervous system including neuroprotective effects. Previous data from our group showed that ouabain treatment is able to induce an increase in retinal ganglion cell survival kept in mixed retinal cell cultures. The aim of this work was to investigate if IL-1β and TNF-α could be mediating the trophic effect of ouabain on retinal ganglion cells. Our results show that the trophic effect of ouabain on retinal ganglion cell was inhibited by either anti-IL-1β or anti-TNF-α antibodies. In agreement, IL-1β or TNF-α increased the retinal ganglion cells survival in a dose-dependent manner. Accordingly, ouabain treatment induces a temporal release of TNF-α and IL-1β from retinal cell cultures. Interestingly, TNF-α and IL-1β regulate each other intracellular levels. Our results suggest that ouabain treatment triggers the activation of TNF-α and IL-1β signaling pathways leading to an increase in retinal ganglion cell survival. - Highlights: • Pro-inflammatory cytokines regulates the ouabain effect on RGC survival. • Ouabain treatment modulates the intracellular levels of TNF-α and IL-1β. • Ouabain induces the release of TNF-α and IL-1β in retinal cell cultures.

  13. Research progress on protecting effect of taurine on retinal ganglion cells

    Directory of Open Access Journals (Sweden)

    Zong-Li Hu

    2014-03-01

    Full Text Available Taurine is a common amino acid in almost all kinds of tissues of animals. Several researches have pointed out that taurine is essential for protecting retinal ganglion cells(RGCs. The mechanisms are summarized as follows: 1Reducing the NMDA-induced RGC excitotoxicity 2Regulating the function of Müller cells 3Protecting function of mitochondria 4Stimulating regeneration of the optic nerve 5Promoting accommodative regulation 6Improving the microcirculation of retina. Glaucoma is one kind of ophthalmopathy leading to apoptosis of RGCs. Recently, taurine has been proved to be effective in protecting the RGC in several aspects, both in vitro and vivo experiments. This indicates a potential treatment of optic nerve in glaucoma or other ophthalmic diseases.

  14. Neural cell adhesion molecule-stimulated neurite outgrowth depends on activation of protein kinase C and the Ras-mitogen-activated protein kinase pathway

    DEFF Research Database (Denmark)

    Kolkova, K; Novitskaya, V; Pedersen, N

    2000-01-01

    , inhibitors of the nonreceptor tyrosine kinase p59(fyn), PLC, PKC and MEK and an activator of PKC, phorbol-12-myristate-13-acetate (PMA). MEK2 transfection rescued cells treated with all inhibitors. The same was found for PMA treatment, except when cells concomitantly were treated with the MEK inhibitor....... Arachidonic acid rescued cells treated with antibodies to the FGF receptor or the PLC inhibitor, but not cells in which the activity of PKC, p59(fyn), FAK, Ras, or MEK was inhibited. Interaction of NCAM with a synthetic NCAM peptide ligand, known to induce neurite outgrowth, was shown to stimulate...

  15. Nerve growth factor (NGF) and pro-NGF increase low-density lipoprotein (LDL) receptors in neuronal cells partly by different mechanisms: role of LDL in neurite outgrowth.

    Science.gov (United States)

    Do, Hai Thi; Bruelle, Céline; Pham, Dan Duc; Jauhiainen, Matti; Eriksson, Ove; Korhonen, Laura T; Lindholm, Dan

    2016-01-01

    Low-density lipoprotein receptors (LDLRs) mediate the uptake of lipoprotein particles into cells, as studied mainly in peripheral tissues. Here, we show that nerve growth factor (NGF) increases LDLR levels in PC6.3 cells and in cultured septal neurons from embryonic rat brain. Study of the mechanisms showed that NGF enhanced transcription of the LDLR gene, acting mainly via Tropomyosin receptor kinase A receptors. Simvastatin, a cholesterol-lowering drug, also increased the LDLR expression in PC6.3 cells. In addition, pro-NGF and pro-brain-derived neurotrophic factor, acting via the p75 neurotrophin receptor (p75NTR) also increased LDLRs. We further observed that Myosin Regulatory Light Chain-Interacting Protein/Inducible Degrader of the LDLR (Mylip/Idol) was down-regulated by pro-NGF, whereas the other LDLR regulator, proprotein convertase subtilisin kexin 9 (PCSK9) was not significantly changed. On the functional side, NGF and pro-NGF increased lipoprotein uptake by neuronal cells as shown using diacetyl-labeled LDL. The addition of serum-derived lipoprotein particles in conjunction with NGF or simvastatin enhanced neurite outgrowth. Collectively, these results show that NGF and simvastatin are able to stimulate lipoprotein uptake by neurons with a positive effect on neurite outgrowth. Increases in LDLRs and lipoprotein particles in neurons could play a functional role during brain development, in neuroregeneration and after brain injuries. Nerve growth factor (NGF) and pro-NGF induce the expression of low-density lipoprotein receptors (LDLRs) in neuronal cells leading to increased LDLR levels. Pro-NGF also down-regulated myosin regulatory light chain-interacting protein/inducible degrader of the LDLR (Mylip/Idol) that is involved in the degradation of LDLRs. NGF acts mainly via Tropomyosin receptor kinase A (TrkA) receptors, whereas pro-NGF stimulates p75 neurotrophin receptor (p75NTR). Elevated LDLRs upon NGF and pro-NGF treatments enhanced lipoprotein uptake

  16. Melanopsin retinal ganglion cells are not labeled in Thy-1YFP-16 transgenic mice.

    Science.gov (United States)

    Grillo, Stephanie L; Stella, Salvatore L

    2018-01-17

    Retinal ganglion cells (RGCs) that express the photopigment melanopsin (mRGCs) are photosensitive and initiate the non-image-forming pathway, where the majority of their axons terminate in the suprachiasmatic nucleus (SCN). RGCs only make up approximately half of the cells in the ganglion cell layer of the retina; therefore, it is important to be able to distinguish them from other cell types. The transgenic Thy-1 YFP mouse line 16 (Thy-1 YFP-16) expresses yellow-fluorescent protein (YFP) in projection neurons, including RGCs. Our objective was to determine whether mRGCs are labeled with YFP in Thy-1 YFP-16 transgenic mice. Paraformaldehyde-fixed retinal wholemounts and frozen vertical sections were prepared from Thy-1 YFP-16 mice and fluorescently labeled with rabbit anti-melanopsin and guinea-pig anti-RNA binding protein with multiple splicing to identify mRGCs and total RGCs, respectively. Thy-1 YFP-16 mouse brains were sectioned coronally and imaged to view RGC axonal projections to the SCN. Confocal images of retinal preparations show that the majority (∼89%) of mRGCs are not YFP-positive in Thy-1 YFP-16 mice, where ∼11% expressed a weak fluorescent signal. In addition, there are almost no YFP-positive axons present in the SCN of coronal brain sections. We conclude that the majority of mRGC somas and axons are not labeled with YFP in the transgenic Thy-1 YFP-16 mouse line; therefore, this mouse model may not suitable for research involving mRGC visual pathways.

  17. Spiral Ganglion Cells and Macrophages Initiate Neuro-inflammation and Scarring Following Cochlear Implantation

    Directory of Open Access Journals (Sweden)

    Esperanza eBas

    2015-08-01

    Full Text Available Conservation of a patient’s residual hearing and prevention of fibrous tissue/new bone formation around an electrode array are some of the major challenges in cochlear implant (CI surgery. Although it is well known that fibrotic tissue formation around the electrode array can interfere with hearing performance in implanted patients, and that associated intracochlear inflammation can initiate loss of residual hearing, little is known about the molecular and cellular mechanisms that promote this response in the cochlea. In vitro studies in neonatal rats and in vivo studies in adult mice were performed to gain insight into the pro-inflammatory, proliferative, and remodeling phases of pathological wound healing that occur in the cochlea following an electrode analogue insertion. Resident Schwann cells, macrophages/microglia, and fibroblasts had a prominent role in the inflammatory process in the cochlea. Leukocytes were recruited to the cochlea following insertion of a nylon filament in adult mice, where contributed to the inflammatory response. The reparative stages in wound healing are characterized by persistent neuro-inflammation of spiral ganglion neurons and expression of regenerative macrophages in the cochlea. Accordingly, genes involved in extracellular matrix deposition and remodeling were up-regulated in implanted cochleae.Maturation of scar tissue occurs in the remodeling phase of wound healing in the cochlea. Similar to other damaged peripheral nerves, M2 macrophages and de-differentiated Schwann cells were observed in damaged cochleae and may play a role in cell survival and axonal regeneration. In conclusion, the insertion of an electrode analogue into the cochlea is associated with robust early and chronic inflammatory responses characterized by recruitment of leukocytes and expression of pro-inflammatory cytokines that promote intracochlear fibrosis and loss of auditory hair cells and spiral ganglion neurons important for hearing

  18. Extremely Low-Frequency Electromagnetic Fields Promote In Vitro Neuronal Differentiation and Neurite Outgrowth of Embryonic Neural Stem Cells via Up-Regulating TRPC1

    Science.gov (United States)

    Ma, Qinlong; Chen, Chunhai; Deng, Ping; Zhu, Gang; Lin, Min; Zhang, Lei; Xu, Shangcheng; He, Mindi; Lu, Yonghui; Duan, Weixia; Pi, Huifeng; Cao, Zhengwang; Pei, Liping; Li, Min; Liu, Chuan; Zhang, Yanwen; Zhong, Min; Zhou, Zhou; Yu, Zhengping

    2016-01-01

    Exposure to extremely low-frequency electromagnetic fields (ELF-EMFs) can enhance hippocampal neurogenesis in adult mice. However, little is focused on the effects of ELF-EMFs on embryonic neurogenesis. Here, we studied the potential effects of ELF-EMFs on embryonic neural stem cells (eNSCs). We exposed eNSCs to ELF-EMF (50 Hz, 1 mT) for 1, 2, and 3 days with 4 hours per day. We found that eNSC proliferation and maintenance were significantly enhanced after ELF-EMF exposure in proliferation medium. ELF-EMF exposure increased the ratio of differentiated neurons and promoted the neurite outgrowth of eNSC-derived neurons without influencing astrocyes differentiation and the cell apoptosis. In addition, the expression of the proneural genes, NeuroD and Ngn1, which are crucial for neuronal differentiation and neurite outgrowth, was increased after ELF-EMF exposure. Moreover, the expression of transient receptor potential canonical 1 (TRPC1) was significantly up-regulated accompanied by increased the peak amplitude of intracellular calcium level induced by ELF-EMF. Furthermore, silencing TRPC1 expression eliminated the up-regulation of the proneural genes and the promotion of neuronal differentiation and neurite outgrowth induced by ELF-EMF. These results suggest that ELF-EMF exposure promotes the neuronal differentiation and neurite outgrowth of eNSCs via up-regulation the expression of TRPC1 and proneural genes (NeuroD and Ngn1). These findings also provide new insights in understanding the effects of ELF-EMF exposure on embryonic brain development. PMID:26950212

  19. Isolated dorsal root ganglion neurones inhibit receptor-dependent adenylyl cyclase activity in associated glial cells

    Science.gov (United States)

    Ng, KY; Yeung, BHS; Wong, YH; Wise, H

    2013-01-01

    Background and Purpose Hyper-nociceptive PGE2 EP4 receptors and prostacyclin (IP) receptors are present in adult rat dorsal root ganglion (DRG) neurones and glial cells in culture. The present study has investigated the cell-specific expression of two other Gs-protein coupled hyper-nociceptive receptor systems: β-adrenoceptors and calcitonin gene-related peptide (CGRP) receptors in isolated DRG cells and has examined the influence of neurone–glial cell interactions in regulating adenylyl cyclase (AC) activity. Experimental Approach Agonist-stimulated AC activity was determined in mixed DRG cell cultures from adult rats and compared with activity in DRG neurone-enriched cell cultures and pure DRG glial cell cultures. Key Results Pharmacological analysis showed the presence of Gs-coupled β2-adrenoceptors and CGRP receptors, but not β1-adrenoceptors, in all three DRG cell preparations. Agonist-stimulated AC activity was weakest in DRG neurone-enriched cell cultures. DRG neurones inhibited IP receptor-stimulated glial cell AC activity by a process dependent on both cell–cell contact and neurone-derived soluble factors, but this is unlikely to involve purine or glutamine receptor activation. Conclusions and Implications Gs-coupled hyper-nociceptive receptors are readily expressed on DRG glial cells in isolated cell cultures and the activity of CGRP, EP4 and IP receptors, but not β2-adrenoceptors, in glial cells is inhibited by DRG neurones. Studies using isolated DRG cells should be aware that hyper-nociceptive ligands may stimulate receptors on glial cells in addition to neurones, and that variable numbers of neurones and glial cells will influence absolute measures of AC activity and affect downstream functional responses. PMID:22924655

  20. The effect of morphology upon electrophysiological responses of retinal ganglion cells: simulation results.

    Science.gov (United States)

    Maturana, Matias I; Kameneva, Tatiana; Burkitt, Anthony N; Meffin, Hamish; Grayden, David B

    2014-04-01

    Retinal ganglion cells (RGCs) display differences in their morphology and intrinsic electrophysiology. The goal of this study is to characterize the ionic currents that explain the behavior of ON and OFF RGCs and to explore if all morphological types of RGCs exhibit the phenomena described in electrophysiological data. We extend our previous single compartment cell models of ON and OFF RGCs to more biophysically realistic multicompartment cell models and investigate the effect of cell morphology on intrinsic electrophysiological properties. The membrane dynamics are described using the Hodgkin - Huxley type formalism. A subset of published patch-clamp data from isolated intact mouse retina is used to constrain the model and another subset is used to validate the model. Two hundred morphologically distinct ON and OFF RGCs are simulated with various densities of ionic currents in different morphological neuron compartments. Our model predicts that the differences between ON and OFF cells are explained by the presence of the low voltage activated calcium current in OFF cells and absence of such in ON cells. Our study shows through simulation that particular morphological types of RGCs are capable of exhibiting the full range of phenomena described in recent experiments. Comparisons of outputs from different cells indicate that the RGC morphologies that best describe recent experimental results are ones that have a larger ratio of soma to total surface area.

  1. Can Retinal Ganglion Cell Dipoles Seed Iso-Orientation Domains in the Visual Cortex?

    Science.gov (United States)

    Schottdorf, Manuel; Eglen, Stephen J.; Wolf, Fred; Keil, Wolfgang

    2014-01-01

    It has been argued that the emergence of roughly periodic orientation preference maps (OPMs) in the primary visual cortex (V1) of carnivores and primates can be explained by a so-called statistical connectivity model. This model assumes that input to V1 neurons is dominated by feed-forward projections originating from a small set of retinal ganglion cells (RGCs). The typical spacing between adjacent cortical orientation columns preferring the same orientation then arises via Moiré-Interference between hexagonal ON/OFF RGC mosaics. While this Moiré-Interference critically depends on long-range hexagonal order within the RGC mosaics, a recent statistical analysis of RGC receptive field positions found no evidence for such long-range positional order. Hexagonal order may be only one of several ways to obtain spatially repetitive OPMs in the statistical connectivity model. Here, we investigate a more general requirement on the spatial structure of RGC mosaics that can seed the emergence of spatially repetitive cortical OPMs, namely that angular correlations between so-called RGC dipoles exhibit a spatial structure similar to that of OPM autocorrelation functions. Both in cat beta cell mosaics as well as primate parasol receptive field mosaics we find that RGC dipole angles are spatially uncorrelated. To help assess the level of these correlations, we introduce a novel point process that generates mosaics with realistic nearest neighbor statistics and a tunable degree of spatial correlations of dipole angles. Using this process, we show that given the size of available data sets, the presence of even weak angular correlations in the data is very unlikely. We conclude that the layout of ON/OFF ganglion cell mosaics lacks the spatial structure necessary to seed iso-orientation domains in the primary visual cortex. PMID:24475081

  2. Rescuing axons from degeneration does not affect retinal ganglion cell death

    Directory of Open Access Journals (Sweden)

    S. de Lima

    2016-01-01

    Full Text Available After a traumatic injury to the central nervous system, the distal stumps of axons undergo Wallerian degeneration (WD, an event that comprises cytoskeleton and myelin breakdown, astrocytic gliosis, and overexpression of proteins that inhibit axonal regrowth. By contrast, injured neuronal cell bodies show features characteristic of attempts to initiate the regenerative process of elongating their axons. The main molecular event that leads to WD is an increase in the intracellular calcium concentration, which activates calpains, calcium-dependent proteases that degrade cytoskeleton proteins. The aim of our study was to investigate whether preventing axonal degeneration would impact the survival of retinal ganglion cells (RGCs after crushing the optic nerve. We observed that male Wistar rats (weighing 200-400 g; n=18 treated with an exogenous calpain inhibitor (20 mM administered via direct application of the inhibitor embedded within the copolymer resin Evlax immediately following optic nerve crush showed a delay in the onset of WD. This delayed onset was characterized by a decrease in the number of degenerated fibers (P<0.05 and an increase in the number of preserved fibers (P<0.05 4 days after injury. Additionally, most preserved fibers showed a normal G-ratio. These results indicated that calpain inhibition prevented the degeneration of optic nerve fibers, rescuing axons from the process of axonal degeneration. However, analysis of retinal ganglion cell survival demonstrated no difference between the calpain inhibitor- and vehicle-treated groups, suggesting that although the calpain inhibitor prevented axonal degeneration, it had no effect on RGC survival after optic nerve damage.

  3. Characterization of Ca2+ Signalling in Postnatal Mouse Retinal Ganglion Cells: Involvement of OPA1 in Ca2+ Clearance

    Czech Academy of Sciences Publication Activity Database

    Dayanithi, Govindan; Chen-Kuo-Chang, M.; Viero, C.; Hamel, C.; Muller, A.; Lenaers, G.

    2010-01-01

    Roč. 31, č. 2 (2010), s. 53-65 ISSN 1381-6810 Institutional research plan: CEZ:AV0Z50390703 Keywords : retinal ganglion cells * Ca2+ homeostasis * Ca2+ clearance Subject RIV: FH - Neurology Impact factor: 1.333, year: 2010

  4. Role of glial cell line-derived neurotrophic factor (GDNF)-neural cell adhesion molecule (NCAM) interactions in induction of neurite outgrowth and identification of a binding site for NCAM in the heel region of GDNF

    DEFF Research Database (Denmark)

    Nielsen, Janne; Gotfryd, Kamil; Li, Shizhong

    2009-01-01

    The formation of appropriate neuronal circuits is an essential part of nervous system development and relies heavily on the outgrowth of axons and dendrites and their guidance to their respective targets. This process is governed by a large array of molecules, including glial cell line......-derived neurotrophic factor (GDNF) and the neural cell adhesion molecule (NCAM), the interaction of which induce neurite outgrowth. In the present study the requirements for NCAM-mediated GDNF-induced neurite outgrowth were investigated in cultures of hippocampal neurons, which do not express Ret. We demonstrate...

  5. Electrical activity of ON and OFF retinal ganglion cells: a modelling study

    Science.gov (United States)

    Guo, Tianruo; Tsai, David; Morley, John W.; Suaning, Gregg J.; Kameneva, Tatiana; Lovell, Nigel H.; Dokos, Socrates

    2016-04-01

    Objective. Retinal ganglion cells (RGCs) demonstrate a large range of variation in their ionic channel properties and morphologies. Cell-specific properties are responsible for the unique way RGCs process synaptic inputs, as well as artificial electrical signals such as that from a visual prosthesis. A cell-specific computational modelling approach allows us to examine the functional significance of regional membrane channel expression and cell morphology. Approach. In this study, an existing RGC ionic model was extended by including a hyperpolarization activated non-selective cationic current as well as a T-type calcium current identified in recent experimental findings. Biophysically-defined model parameters were simultaneously optimized against multiple experimental recordings from ON and OFF RGCs. Main results. With well-defined cell-specific model parameters and the incorporation of detailed cell morphologies, these models were able to closely reconstruct and predict ON and OFF RGC response properties recorded experimentally. Significance. The resulting models were used to study the contribution of different ion channel properties and spatial structure of neurons to RGC activation. The techniques of this study are generally applicable to other excitable cell models, increasing the utility of theoretical models in accurately predicting the response of real biological neurons.

  6. Effect of Extracellular Zinc Chelator on Rat Retinal Ganglion Cell Number, and Taurine and Zinc Transporters in These Cells

    Directory of Open Access Journals (Sweden)

    Asarí Márquez García

    2017-05-01

    Full Text Available Zinc deficiency in humans causes decreased antioxidants in the retina and is related with abnormal darkness adaptation, cataracts, blindness, and macular degeneration. There is little information about the effects of zinc on the taurine system in mammalian retinal cells. Therefore, we studied the effect of zinc on the taurine transporter (TAUT and zinc transporters (ZnT-1 and 3 using the extracellular zinc chelator, diethylenetriaminepentaacetic acid (DTPA by fluorescence immunocytochemistry and immunohistochemistry in the ganglion cells (CG and cell layers of the retina of rats. Three days after administration of DTPA (10µM primary antibodies and secondary antibodies conjugated with rhodamine or fluorescein isothiocyanate (FITC were used as required. For immunocytochemical labeling approximately three hundred cells per condition were counted. For immunohistochemical labeling, the fluorescence intensity was measured as integrated optical density (DOI in four areas for each layer of tissue. DTPA produced a decrease of 32 % and 29 % in GC of the total cells labeled with antibody against glycoprotein Thy 1.1 and γ-synuclein, respectively. It also produced a significant decrease in TAUT localization in 27 and 28 % compared to controls. DTPA produced a decrease in the localization of ZnT-1 and ZnT-3 in the retina layers (ganglion cells, GCC and the outer and inner plexiform, CEP and CIP. The study of these molecules in the retina is relevant to understanding the interactions of taurine and zinc in this structure.

  7. Effect of eye NGF administration on two animal models of retinal ganglion cells degeneration

    Directory of Open Access Journals (Sweden)

    Valeria Colafrancesco

    2011-01-01

    Full Text Available The aim of this study was to investigate the effect of nerve growth factor (NGF administration on retinal ganglion cells (RGCs in experimentally induced glaucoma (GL and diabetic retinopathy (DR. GL was induced in adult rats by injection of hypertonic saline into the episcleral vein of the eye and diabetes (DT was induced by administration of streptozoticin. Control and experimental rats were treated daily with either ocular application of NGF or vehicle solution. We found that both animal models present a progressive degeneration of RGCs and changing NGF and VEGF levels in the retina and optic nerve. We then proved that NGF eye drop administration exerts a protective effect on these models of retinal degeneration. In brief, our findings indicate that NGF can play a protective role against RGC degeneration occurring in GL and DR and suggest that ocular NGF administration might be an effective pharmacological approach.

  8. Synchronized Firings in Retinal Ganglion Cells in Response to Natural Stimulation

    International Nuclear Information System (INIS)

    Zhang Ying-Ying; Xiao Lei; Liu Wen-Zhong; Gong Hai-Qing; Liang Pei-Ji

    2011-01-01

    The response of synchronously firing groups of population retinal ganglion cells (RGCs) to natural movies (NMs) and pseudo-random white-noise checker-board flickering (CB, as control) are investigated using an information-theoretic algorithm. The main results are: (1) the population RGCs tend to fire in synchrony far more frequently than expected by chance during both NM and CB stimulation; (2) more synchronous groups could be formed and each group contains more neurons under NM than CB stimulation; (3) the individual neurons also participate in more groups and have more distinct partners in NM than CB stimulation. All these results suggest that the synchronized firings in RGCs are more extensive and diverse, which may account for more effective information processing in representing the natural visual environment. (cross-disciplinary physics and related areas of science and technology)

  9. Human adipose tissue-derived multilineage progenitor cells exposed to oxidative stress induce neurite outgrowth in PC12 cells through p38 MAPK signaling

    Directory of Open Access Journals (Sweden)

    Moriyama Mariko

    2012-08-01

    Full Text Available Abstract Background Adipose tissues contain populations of pluripotent mesenchymal stem cells that also secrete various cytokines and growth factors to support repair of damaged tissues. In this study, we examined the role of oxidative stress on human adipose-derived multilineage progenitor cells (hADMPCs in neurite outgrowth in cells of the rat pheochromocytoma cell line (PC12. Results We found that glutathione depletion in hADMPCs, caused by treatment with buthionine sulfoximine (BSO, resulted in the promotion of neurite outgrowth in PC12 cells through upregulation of bone morphogenetic protein 2 (BMP2 and fibroblast growth factor 2 (FGF2 transcription in, and secretion from, hADMPCs. Addition of N-acetylcysteine, a precursor of the intracellular antioxidant glutathione, suppressed the BSO-mediated upregulation of BMP2 and FGF2. Moreover, BSO treatment caused phosphorylation of p38 MAPK in hADMPCs. Inhibition of p38 MAPK was sufficient to suppress BMP2 and FGF2 expression, while this expression was significantly upregulated by overexpression of a constitutively active form of MKK6, which is an upstream molecule from p38 MAPK. Conclusions Our results clearly suggest that glutathione depletion, followed by accumulation of reactive oxygen species, stimulates the activation of p38 MAPK and subsequent expression of BMP2 and FGF2 in hADMPCs. Thus, transplantation of hADMPCs into neurodegenerative lesions such as stroke and Parkinson’s disease, in which the transplanted hADMPCs are exposed to oxidative stress, can be the basis for simple and safe therapies.

  10. Topographic prominence discriminator for the detection of short-latency spikes of retinal ganglion cells

    Science.gov (United States)

    Choi, Myoung-Hwan; Ahn, Jungryul; Park, Dae Jin; Lee, Sang Min; Kim, Kwangsoo; Cho, Dong-il Dan; Senok, Solomon S.; Koo, Kyo-in; Goo, Yong Sook

    2017-02-01

    Objective. Direct stimulation of retinal ganglion cells in degenerate retinas by implanting epi-retinal prostheses is a recognized strategy for restoration of visual perception in patients with retinitis pigmentosa or age-related macular degeneration. Elucidating the best stimulus-response paradigms in the laboratory using multielectrode arrays (MEA) is complicated by the fact that the short-latency spikes (within 10 ms) elicited by direct retinal ganglion cell (RGC) stimulation are obscured by the stimulus artifact which is generated by the electrical stimulator. Approach. We developed an artifact subtraction algorithm based on topographic prominence discrimination, wherein the duration of prominences within the stimulus artifact is used as a strategy for identifying the artifact for subtraction and clarifying the obfuscated spikes which are then quantified using standard thresholding. Main results. We found that the prominence discrimination based filters perform creditably in simulation conditions by successfully isolating randomly inserted spikes in the presence of simple and even complex residual artifacts. We also show that the algorithm successfully isolated short-latency spikes in an MEA-based recording from degenerate mouse retinas, where the amplitude and frequency characteristics of the stimulus artifact vary according to the distance of the recording electrode from the stimulating electrode. By ROC analysis of false positive and false negative first spike detection rates in a dataset of one hundred and eight RGCs from four retinal patches, we found that the performance of our algorithm is comparable to that of a generally-used artifact subtraction filter algorithm which uses a strategy of local polynomial approximation (SALPA). Significance. We conclude that the application of topographic prominence discrimination is a valid and useful method for subtraction of stimulation artifacts with variable amplitudes and shapes. We propose that our algorithm

  11. Coding properties of three intrinsically distinct retinal ganglion cells under periodic stimuli: a computational study

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    Lei Wang

    2016-09-01

    Full Text Available As the sole output neurons in the retina, ganglion cells play significant roles in transforming visual information into spike trains, and then transmitting them to the higher visual centers. However, coding strategies that retinal ganglion cells (RGCs adopt to accomplish these processes are not completely clear yet. To clarify these issues, we investigate the coding properties of three types of RGCs (repetitive spiking, tonic firing, and phasic firing by two different measures (spike-rate and spike-latency. Model results show that for periodic stimuli, repetitive spiking RGC and tonic RGC exhibit similar spike-rate patterns. Their spike-rates decrease gradually with increased stimulus frequency, moreover, variation of stimulus amplitude would change the two RGCs’ spike-rate patterns. For phasic RGC, it activates strongly at medium levels of frequency when the stimulus amplitude is low. While if high stimulus amplitude is applied, phasic RGC switches to respond strongly at low frequencies. These results suggest that stimulus amplitude is a prominent factor in regulating RGCs in encoding periodic signals. Similar conclusions can be drawn when analyzes spike-latency patterns of the three RGCs. More importantly, the above phenomena can be accurately reproduced by Hodgkin’s three classes of neurons, indicating that RGCs can perform the typical three classes of firing dynamics, depending on the distinctions of ion channel densities. Consequently, model results from the three RGCs may be not specific, but can also applicable to neurons in other brain regions which exhibit part(s or all of the Hodgkin’s three excitabilities.

  12. Ganglion cell loss in relation to visual disability in multiple sclerosis.

    Science.gov (United States)

    Walter, Scott D; Ishikawa, Hiroshi; Galetta, Kristin M; Sakai, Reiko E; Feller, Daniel J; Henderson, Sam B; Wilson, James A; Maguire, Maureen G; Galetta, Steven L; Frohman, Elliot; Calabresi, Peter A; Schuman, Joel S; Balcer, Laura J

    2012-06-01

    We used high-resolution spectral-domain optical coherence tomography (SD-OCT) with retinal segmentation to determine how ganglion cell loss relates to history of acute optic neuritis (ON), retinal nerve fiber layer (RNFL) thinning, visual function, and vision-related quality of life (QOL) in multiple sclerosis (MS). Cross-sectional study. A convenience sample of patients with MS (n = 122; 239 eyes) and disease-free controls (n = 31; 61 eyes). Among MS eyes, 87 had a history of ON before enrollment. The SD-OCT images were captured using Macular Cube (200×200 or 512×128) and ONH Cube 200×200 protocols. Retinal layer segmentation was performed using algorithms established for glaucoma studies. Thicknesses of the ganglion cell layer/inner plexiform layer (GCL+IPL), RNFL, outer plexiform/inner nuclear layers (OPL+INL), and outer nuclear/photoreceptor layers (ONL+PRL) were measured and compared in MS versus control eyes and MS ON versus non-ON eyes. The relation between changes in macular thickness and visual disability was also examined. The OCT measurements of GCL+IPL and RNFL thickness; high contrast visual acuity (VA); low-contrast letter acuity (LCLA) at 2.5% and 1.25% contrast; on the 25-item National Eye Institute Visual Function Questionnaire (NEI-VFQ-25) and 10-Item Neuro-Ophthalmic Supplement composite score. Macular RNFL and GCL+IPL were significantly decreased in MS versus control eyes (Pvisual function and vision-specific QOL in MS, and may serve as a useful structural marker of disease. Our findings parallel those of magnetic resonance imaging studies that show gray matter disease is a marker of neurologic disability in MS. Proprietary or commercial disclosure may be found after the references. Copyright © 2012 American Academy of Ophthalmology. Published by Elsevier Inc. All rights reserved.

  13. Endothelin B receptors contribute to retinal ganglion cell loss in a rat model of glaucoma.

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    Alena Z Minton

    Full Text Available Glaucoma is an optic neuropathy, commonly associated with elevated intraocular pressure (IOP characterized by optic nerve degeneration, cupping of the optic disc, and loss of retinal ganglion cells which could lead to loss of vision. Endothelin-1 (ET-1 is a 21-amino acid vasoactive peptide that plays a key role in the pathogenesis of glaucoma; however, the receptors mediating these effects have not been defined. In the current study, endothelin B (ET(B receptor expression was assessed in vivo, in the Morrison's ocular hypertension model of glaucoma in rats. Elevation of IOP in Brown Norway rats produced increased expression of ET(B receptors in the retina, mainly in retinal ganglion cells (RGCs, nerve fiber layer (NFL, and also in the inner plexiform layer (IPL and inner nuclear layer (INL. To determine the role of ET(B receptors in neurodegeneration, Wistar-Kyoto wild type (WT and ET(B receptor-deficient (KO rats were subjected to retrograde labeling with Fluoro-Gold (FG, following which IOP was elevated in one eye while the contralateral eye served as control. IOP elevation for 4 weeks in WT rats caused an appreciable loss of RGCs, which was significantly attenuated in KO rats. In addition, degenerative changes in the optic nerve were greatly reduced in KO rats compared to those in WT rats. Taken together, elevated intraocular pressure mediated increase in ET(B receptor expression and its activation may contribute to a decrease in RGC survival as seen in glaucoma. These findings raise the possibility of using endothelin receptor antagonists as neuroprotective agents for the treatment of glaucoma.

  14. Asymmetric inheritance of the apical domain and self-renewal of retinal ganglion cell progenitors depend on Anillin function.

    Science.gov (United States)

    Paolini, Alessio; Duchemin, Anne-Laure; Albadri, Shahad; Patzel, Eva; Bornhorst, Dorothee; González Avalos, Paula; Lemke, Steffen; Machate, Anja; Brand, Michael; Sel, Saadettin; Di Donato, Vincenzo; Del Bene, Filippo; Zolessi, Flavio R; Ramialison, Mirana; Poggi, Lucia

    2015-03-01

    Divisions that generate one neuronal lineage-committed and one self-renewing cell maintain the balance of proliferation and differentiation for the generation of neuronal diversity. The asymmetric inheritance of apical domains and components of the cell division machinery has been implicated in this process, and might involve interactions with cell fate determinants in regulatory feedback loops of an as yet unknown nature. Here, we report the dynamics of Anillin - an essential F-actin regulator and furrow component - and its contribution to progenitor cell divisions in the developing zebrafish retina. We find that asymmetrically dividing retinal ganglion cell progenitors position the Anillin-rich midbody at the apical domain of the differentiating daughter. anillin hypomorphic conditions disrupt asymmetric apical domain inheritance and affect daughter cell fate. Consequently, the retinal cell type composition is profoundly affected, such that the ganglion cell layer is dramatically expanded. This study provides the first in vivo evidence for the requirement of Anillin during asymmetric neurogenic divisions. It also provides insights into a reciprocal regulation between Anillin and the ganglion cell fate determinant Ath5, suggesting a mechanism whereby the balance of proliferation and differentiation is accomplished during progenitor cell divisions in vivo. © 2015. Published by The Company of Biologists Ltd.

  15. The ciliary margin zone of the mammalian retina generates retinal ganglion cells

    Science.gov (United States)

    Marcucci, Florencia; Murcia-Belmonte, Veronica; Coca, Yaiza; Ferreiro-Galve, Susana; Wang, Qing; Kuwajima, Takaaki; Khalid, Sania; Ross, M. Elizabeth; Herrera, Eloisa; Mason, Carol

    2016-01-01

    Summary The retina of lower vertebrates grows continuously by integrating new neurons generated from progenitors in the ciliary margin zone (CMZ). Whether the mammalian CMZ provides the neural retina with retinal cells is controversial. Live-imaging of embryonic retina expressing eGFP in the CMZ shows that cells migrate laterally from the CMZ to the neural retina where differentiated retinal ganglion cells (RGCs) reside. As Cyclin D2, a cell-cycle regulator, is enriched in ventral CMZ, we analyzed Cyclin D2−/− mice to test whether the CMZ is a source of retinal cells. Neurogenesis is diminished in Cyclin D2 mutants, leading to a reduction of RGCs in the ventral retina. In line with these findings, in the albino retina, the decreased production of ipsilateral RGCs is correlated with fewer Cyclin D2+ cells. Together, these results implicate the mammalian CMZ as a neurogenic site that produces RGCs and whose proper generation depends on Cyclin D2 activity. PMID:28009286

  16. Dose-dependent effects of ouabain on spiral ganglion neurons and Schwann cells in mouse cochlea.

    Science.gov (United States)

    Zhang, Zhi-Jian; Guan, Hong-Xia; Yang, Kun; Xiao, Bo-Kui; Liao, Hua; Jiang, Yang; Zhou, Tao; Hua, Qing-Quan

    2017-10-01

    This study aimed in fully investigating the toxicities of ouabain to mouse cochlea and the related cellular environment, and providing an optimal animal model system for cell transplantation in the treatment of auditory neuropathy (AN) and sensorineural hearing loss (SNHL). Different dosages of ouabain were applied to mouse round window. The auditory brainstem responses and distortion product otoacoustic emissions were used to evaluate the cochlear function. The immunohistochemical staining and cochlea surface preparation were performed to detect the spiral ganglion neurons (SGNs), Schwann cells and hair cells. Ouabain at the dosages of 0.5 mM, 1 mM and 3 mM selectively and permanently destroyed SGNs and their functions, while leaving the hair cells relatively intact. Ouabain at 3 mM resulted in the most severe SGNs loss and induced significant loss of Schwann cells started as early as 7 days and with further damages at 14 and 30 days after ouabain exposure. The application of ouabain to mouse round window induces damages of SGNs and Schwann cells in a dose- and time-dependent manner, this study established a reliable and accurate animal model system of AN and SNHL.

  17. The Ciliary Margin Zone of the Mammalian Retina Generates Retinal Ganglion Cells.

    Science.gov (United States)

    Marcucci, Florencia; Murcia-Belmonte, Veronica; Wang, Qing; Coca, Yaiza; Ferreiro-Galve, Susana; Kuwajima, Takaaki; Khalid, Sania; Ross, M Elizabeth; Mason, Carol; Herrera, Eloisa

    2016-12-20

    The retina of lower vertebrates grows continuously by integrating new neurons generated from progenitors in the ciliary margin zone (CMZ). Whether the mammalian CMZ provides the neural retina with retinal cells is controversial. Live imaging of embryonic retina expressing eGFP in the CMZ shows that cells migrate laterally from the CMZ to the neural retina where differentiated retinal ganglion cells (RGCs) reside. Because Cyclin D2, a cell-cycle regulator, is enriched in ventral CMZ, we analyzed Cyclin D2 -/- mice to test whether the CMZ is a source of retinal cells. Neurogenesis is diminished in Cyclin D2 mutants, leading to a reduction of RGCs in the ventral retina. In line with these findings, in the albino retina, the decreased production of ipsilateral RGCs is correlated with fewer Cyclin D2 + cells. Together, these results implicate the mammalian CMZ as a neurogenic site that produces RGCs and whose proper generation depends on Cyclin D2 activity. Copyright © 2016 The Authors. Published by Elsevier Inc. All rights reserved.

  18. Loss of Aβ-nerve endings associated with the Merkel cell-neurite complex in the lesional oral mucosa epithelium of lichen planus and hyperkeratosis.

    Science.gov (United States)

    Carrión, Daniela Calderón; Korkmaz, Yüksel; Cho, Britta; Kopp, Marion; Bloch, Wilhelm; Addicks, Klaus; Niedermeier, Wilhelm

    2016-03-30

    The Merkel cell-neurite complex initiates the perception of touch and mediates Aβ slowly adapting type I responses. Lichen planus is a chronic inflammatory autoimmune disease with T-cell-mediated inflammation, whereas hyperkeratosis is characterized with or without epithelial dysplasia in the oral mucosa. To determine the effects of lichen planus and hyperkeratosis on the Merkel cell-neurite complex, healthy oral mucosal epithelium and lesional oral mucosal epithelium of lichen planus and hyperkeratosis patients were stained by immunohistochemistry (the avidin-biotin-peroxidase complex and double immunofluorescence methods) using pan cytokeratin, cytokeratin 20 (K20, a Merkel cell marker), and neurofilament 200 (NF200, a myelinated Aβ- and Aδ-nerve fibre marker) antibodies. NF200-immunoreactive (ir) nerve fibres in healthy tissues and in the lesional oral mucosa epithelium of lichen planus and hyperkeratosis were counted and statistically analysed. In the healthy oral mucosa, K20-positive Merkel cells with and without close association to the intraepithelial NF200-ir nerve fibres were detected. In the lesional oral mucosa of lichen planus and hyperkeratosis patients, extremely rare NF200-ir nerve fibres were detected only in the lamina propria. Compared with healthy tissues, lichen planus and hyperkeratosis tissues had significantly decreased numbers of NF200-ir nerve fibres in the oral mucosal epithelium. Lichen planus and hyperkeratosis were associated with the absence of Aβ-nerve endings in the oral mucosal epithelium. Thus, we conclude that mechanosensation mediated by the Merkel cell-neurite complex in the oral mucosal epithelium is impaired in lichen planus and hyperkeratosis.

  19. Neurite outgrowth in human induced pluripotent stem cell-derived neurons as a high-throughput screen for developmental neurotoxicity or neurotoxicity.

    Science.gov (United States)

    Ryan, Kristen R; Sirenko, Oksana; Parham, Fred; Hsieh, Jui-Hua; Cromwell, Evan F; Tice, Raymond R; Behl, Mamta

    2016-03-01

    Due to the increasing prevalence of neurological disorders and the large number of untested compounds in the environment, there is a need to develop reliable and efficient screening tools to identify environmental chemicals that could potentially affect neurological development. Herein, we report on a library of 80 compounds screened for their ability to inhibit neurite outgrowth, a process by which compounds may elicit developmental neurotoxicity, in a high-throughput, high-content assay using human neurons derived from induced pluripotent stem cells (iPSC). The library contains a diverse set of compounds including those that have been known to be associated with developmental neurotoxicity (DNT) and/or neurotoxicity (NT), environmental compounds with unknown neurotoxic potential (e.g., polycyclic aromatic hydrocarbons (PAHs) and flame retardants (FRs)), as well as compounds with no documented neurotoxic potential. Neurons were treated for 72h across a 6-point concentration range (∼0.3-100μM) in 384-well plates. Effects on neurite outgrowth were assessed by quantifying total outgrowth, branches, and processes. We also assessed the number ofviable cells per well. Concentration-response profiles were evaluated using a Hill model to derive benchmark concentration (BMC) values. Assay performance was evaluated using positive and negative controls and test replicates. Compounds were ranked by activity and selectivity (i.e., specific effects on neurite outgrowth in the absence of concomitant cytotoxicity) and repeat studies were conducted to confirm selectivity. Among the 80 compounds tested, 38 compounds were active, of which 16 selectively inhibited neurite outgrowth. Of these 16 compounds, 12 were known to cause DNT/NT and the remaining 4 compounds included 3 PAHs and 1 FR. In independent repeat studies, 14/16 selective compounds were reproducibly active in the assay, of which only 6 were selective for inhibition of neurite outgrowth. These 6 compounds were

  20. Hepatocyte growth factor promotes long-term survival and axonal regeneration of retinal ganglion cells after optic nerve injury: comparison with CNTF and BDNF.

    Science.gov (United States)

    Wong, Wai-Kai; Cheung, Anny Wan-Suen; Yu, Sau-Wai; Sha, Ou; Cho, Eric Yu Pang

    2014-10-01

    Different trophic factors are known to promote retinal ganglion cell survival and regeneration, but each had their own limitations. We report that hepatocyte growth factor (HGF) confers distinct advantages in supporting ganglion cell survival and axonal regeneration, when compared to two well-established trophic factors ciliary neurotrophic factor (CNTF) and brain-derived neurotrophic factor (BDNF). Ganglion cells in adult hamster were injured by cutting the optic nerve. HGF, CNTF, or BDNF was injected at different dosages intravitreally after injury. Ganglion cell survival was quantified at 7, 14, or 28 days postinjury. Peripheral nerve (PN) grafting to the cut optic nerve of the growth factor-injected eye was performed either immediately after injury or delayed until 7 days post-injury. Expression of heat-shock protein 27 and changes in microglia numbers were quantified in different growth factor groups. The cellular distribution of c-Met in the retina was examined by anti-c-Met immunostaining. Hepatocyte Growth Factor (HGF) was equally potent as BDNF in promoting short-term survival (up to 14 days post-injury) and also supported survival at 28 days post-injury when ganglion cells treated by CNTF or BDNF failed to be sustained. When grafting was performed without delay, HGF stimulated twice the number of axons to regenerate compared with control but was less potent than CNTF. However, in PN grafting delayed for 7 days after optic nerve injury, HGF maintained a better propensity of ganglion cells to regenerate than CNTF. Unlike CNTF, HGF application did not increase HSP27 expression in ganglion cells. Microglia proliferation was prolonged in HGF-treated retinas compared with CNTF or BDNF. C-Met was localized to both ganglion cells and Muller cells, suggesting HGF could be neuroprotective via interacting with both neurons and glia. Compared with CNTF or BDNF, HGF is advantageous in sustaining long-term ganglion cell survival and their propensity to respond to

  1. Effects of sub-lethal neurite outgrowth inhibitory concentrations of chlorpyrifos oxon on cytoskeletal proteins and acetylcholinesterase in differentiating N2a cells

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    Flaskos, J., E-mail: flaskos@vet.auth.gr [Laboratory of Biochemistry and Toxicology, School of Veterinary Medicine, Aristotle University of Thessaloniki, 54124 Thessaloniki (Greece); Nikolaidis, E. [Laboratory of Biochemistry and Toxicology, School of Veterinary Medicine, Aristotle University of Thessaloniki, 54124 Thessaloniki (Greece); Harris, W. [School of Science and Technology, Nottingham Trent University, Clifton Lane, Nottingham NG11 8NS (United Kingdom); Sachana, M. [Laboratory of Biochemistry and Toxicology, School of Veterinary Medicine, Aristotle University of Thessaloniki, 54124 Thessaloniki (Greece); Hargreaves, A.J., E-mail: alan.hargreaves@ntu.ac.uk [School of Science and Technology, Nottingham Trent University, Clifton Lane, Nottingham NG11 8NS (United Kingdom)

    2011-11-15

    Previous work in our laboratory has shown that sub-lethal concentrations (1-10 {mu}M) of chlorpyrifos (CPF), diazinon (DZ) and diazinon oxon (DZO) inhibit the outgrowth of axon-like neurites in differentiating mouse N2a neuroblastoma cells concomitant with altered levels and/or phosphorylation state of axonal cytoskeleton and growth-associated proteins. The aim of the present work was to determine whether chlorpyrifos oxon (CPO) was capable of inhibiting N2a cell differentiation in a similar manner. Using experimental conditions similar to our previous work, sub-lethal concentrations (1-10 {mu}M) of CPO were found to inhibit N2a cell differentiation. However, unlike previous studies with DZ and DZO, there was a high level of sustained inhibition of acetylcholinesterase (AChE) in CPO treated cells. Impairment of neurite outgrowth was also associated with reduced levels of growth associated protein-43 and neurofilament heavy chain (NFH), and the distribution of NFH in cells stained by indirect immunofluorescence was disrupted. However, in contrast to previous findings for DZO, the absolute level of phosphorylated NFH was unaffected by CPO exposure. Taken together, the findings suggest that sub-lethal concentrations of CPO inhibit axon outgrowth in differentiating N2a cells and that this effect involves reduced levels of two proteins that play key roles in axon outgrowth and maintenance. Although the inhibition of neurite outgrowth is unlikely to involve AChE inhibition directly, further work will help to determine whether the persistent inhibition of AChE by CPO can account for the different effects induced by CPO and DZO on the levels of total and phosphorylated NFH. -- Highlights: Black-Right-Pointing-Pointer Sub-lethal levels of chlorpyrifos oxon inhibit neurite outgrowth in N2a cells Black-Right-Pointing-Pointer Acetylcholinesterase exhibits sustained inhibition throughout exposure Black-Right-Pointing-Pointer The levels of neurofilament heavy chain and GAP-43

  2. Imaging of single retinal ganglion cell with differential interference contrast microscopy (Conference Presentation)

    Science.gov (United States)

    Oh, Juyeong; Kim, Yu Jeong; Kim, Chul-Ki; Lee, Taik Jin; Seo, Mina; Lee, Seok; Woo, Deok Ha; Jun, Seong Chan; Park, Ki-Ho; Kim, Seok Hwan; Kim, Jae Hun

    2017-02-01

    Glaucoma is a progressive optic neuropathy, characterized by the selective loss of retinal ganglion cells (RGCs). Therefore, monitoring the change of number or morphology of RGC is essential for the early detection as well as investigation of pathophysiology of glaucoma. Since RGC layer is transparent and hyporeflective, the direct optical visualization of RGCs has not been successful so far. Therefore, glaucoma evaluation mostly depends on indirect diagnostic methods such as the evaluation of optic disc morphology or retinal nerve fiber layer thickness measurement by optical coherence tomography. We have previously demonstrated single photoreceptor cell imaging with differential interference contrast (DIC) microscopy. Herein, we successfully visualized single RGC using DIC microscopy. Since RGC layer is much less reflective than photoreceptor layer, various techniques including the control of light wavelength and bandwidth using a tunable band pass filter were adopted to reduce the chromatic aberration in z-axis for higher and clearer resolution. To verify that the imaged cells were the RGCs, the flat-mounted retina of Sprague-Dawley rat, in which the RGCs were retrogradely labeled with fluorescence, was observed by both fluorescence and DIC microscopies for direct comparison. We have confirmed that the cell images obtained by fluorescence microscopy were perfectly matched with cell images by DIC microscopy. As conclusion, we have visualized single RGC with DIC microscopy, and confirmed with fluorescence microscopy.

  3. Effects of L-carnitine on high glucose-induced oxidative stress in retinal ganglion cells.

    Science.gov (United States)

    Cao, Yu; Li, Xin; Shi, Ping; Wang, Le-xin; Sui, Zhong-guo

    2014-01-01

    Oxidative stress plays a role in diabetic retinopathy. L-Carnitine is an endogenous mitochondrial membrane compound. To elucidate the protective effects of L-carnitine on high glucose-induced oxidative stress in retinal ganglion cells (RGCs). Hoechst 33258 staining was used to estimate cell loss. Mitochondrial function was predicted by mitochondrial membrane potential (ΔΨm) measurement. The expression of apoptosis-related protein was measured by Western blotting. Assays for reactive oxygen species (ROS) accumulation, lipid peroxidation, total antioxidative capacity (T-AOC) and antioxidant defense enzymes were completed to explain the antioxidative capacity of L-carnitine. L-Carnitine (12 h) inhibited high glucose-mediated cell loss and restored mitochondrial function including a reversion of ΔΨm loss and cytochrome c release. Cell apoptosis triggered by high glucose was also inhibited by L-carnitine, characterized by the downregulation of caspase-9, caspase-3 and Bax/Bcl-2. Furthermore, L-carnitine inhibited high glucose-induced ROS production and lipid peroxidation and promoted endogenous antioxidant defense components including superoxide dismutase, glutathione peroxidase, catalase and T-AOC in a concentration-dependent manner. L-Carnitine may protect RGCs from high glucose-induced injury through the inhibition of oxidative damage, mitochondrial dysfunction and, ultimately, cell apoptosis. © 2014 S. Karger AG, Basel.

  4. Imipramine protects retinal ganglion cells from oxidative stress through the tyrosine kinase receptor B signaling pathway

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    Ming-lei Han

    2016-01-01

    Full Text Available Retinal ganglion cell (RGC degeneration is irreversible in glaucoma and tyrosine kinase receptor B (TrkB-associated signaling pathways have been implicated in the process. In this study, we attempted to examine whether imipramine, a tricyclic antidepressant, may protect hydrogen peroxide (H 2 O 2 -induced RGC degeneration through the activation of the TrkB pathway in RGC-5 cell lines. RGC-5 cell lines were pre-treated with imipramine 30 minutes before exposure to H 2 O 2 . Western blot assay showed that in H 2 O 2 -damaged RGC-5 cells, imipramine activated TrkB pathways through extracellular signal-regulated protein kinase/TrkB phosphorylation. TUNEL staining assay also demonstrated that imipramine ameliorated H 2 O 2 -induced apoptosis in RGC-5 cells. Finally, TrkB-IgG intervention was able to reverse the protective effect of imipramine on H 2 O 2 -induced RGC-5 apoptosis. Imipramine therefore protects RGCs from oxidative stress-induced apoptosis through the TrkB signaling pathway.

  5. The role of NgR-Rhoa-Rock signal pathway in retinal ganglion cell apoptosis of early diabetic rats

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    Yun-Jie Fu

    2014-09-01

    Full Text Available AIM: To study the function and mechanism of the NgR-Rhoa-Rock signal pathways which exists in the retinal ganglion cells apoptosis in diabetes mellitus(DMrats. METHODS: Some healthy SD rats were operated by means of single intraperitoneal injection of 1% streptozotocin based on the standard of 50mg/kg wight, after that the blood sugar value was greater than 16.7mmol/L as DM model, then randomly divided into 3 groups, each group was 10 rats. In addition to take 10 healthy SD rats as control group. Four groups of rats were bilaterally eyeball intravitreal injection in turn with NgR-siRNA virus 10μL(siRNA group, NgR-siRNA virus diluted 10μL(DM group, NgR-siRNA virus-negative-control solution 10μL(siRNA blank group, NgR-siRNA virus diluted 10μL(normal control group, and fed normally. During that time, some life indexes like blood glucose, body mass, etc. were measured and recorded. After 12wk, the expression of NgR and Rhoa, HE staining, and TUNNEL staining were detected by Western blot analysis. RESULTS: Western blot analysis: compared with normal control group, the expression of NgR and Rhoa in DM group and siRNA blank group increased significantly(PP>0.05; compared with DM group and siRNA blank group, the expression of those proteins significantly lowered in siRNA group. HE staining: compared with normal control group, some extent ganglion cells arranged disorder, irregular shape, spacing not consistent were all found in three groups of model rats; compared with DM group and siRNA blank group, there was some improvement in siRNA group of ganglion cells about the order and shape size. TUNEL staining: compared with normal control group, there were retinal ganglion cells apoptosis in all of three groups of model rats. Compared with DM group and siRNA blank group, the number of retinal ganglion cells apoptotic cells was less, and the shape of cells had improved significantly in siRNA group. CONCLUSION: In the DM phase, the expression of NgR and

  6. Neurite outgrowth stimulatory effects of myco­synthesized AuNPs from Hericium erinaceus (Bull.: Fr. Pers. on pheochromocytoma (PC-12 cells

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    Raman J

    2015-09-01

    Full Text Available Jegadeesh Raman,1 Hariprasath Lakshmanan,1 Priscilla A John,1,2 Chan Zhijian,3 Vengadesh Periasamy,3 Pamela David,1,4 Murali Naidu,1,4 Vikineswary Sabaratnam1,2 1Mushroom Research Centre, 2Institute of Biological Sciences, Faculty of Science, University of Malaya, 3Low Dimensional Materials Research Center (LDMRC, Department of Physics, Faculty of Science, 4Department of Anatomy, Faculty of Medicine, University of Malaya, Kuala Lumpur, Malaysia Background: Hericium erinaceus has been reported to have a wide range of medicinal properties such as stimulation of neurite outgrowth, promotion of functional recovery of axonotmetic peroneal nerve injury, antioxidant, antihypertensive, and antidiabetic properties. In recent years, the green synthesis of gold nanoparticles (AuNPs has attracted intense interest due to the potential use in biomedical applications. The aim of this study was to investigate the effects of AuNPs from aqueous extract of H. erinaceus on neurite outgrowth of rat pheochromocytoma (PC-12 cells. Methods: The formation of AuNPs was characterized by UV–visible spectrum, energy dispersive X-ray (EDX, field-emission scanning electron microscope (FESEM, transmission electron microscopy (TEM, particle size distribution, and Fourier transform-infrared spectroscopy (FTIR. Furthermore, the neurite extension study of synthesized AuNPs was evaluated by in vitro assay. Results: The AuNPs exhibited maximum absorbance between 510 and 600 nm in UV–visible spectrum. FESEM and TEM images showed the existence of nanoparticles with sizes of 20–40 nm. FTIR measurements were carried out to identify the possible biomolecules responsible for capping and efficient stabilization of the nanoparticles. The purity and the crystalline properties were confirmed by EDX diffraction analysis, which showed strong signals with energy peaks in the range of 2–2.4 keV, indicating the existence of gold atoms. The synthesized AuNPs showed significant neurite

  7. Distribution of 5-HT1FReceptors in Monkey Vestibular and Trigeminal Ganglion Cells.

    Science.gov (United States)

    Usman, Habiba O; Balaban, Carey D

    2016-01-01

    Evidence of serotonergic involvement in vestibular pathway contributions to migraine and balance disorders is compelling. Serotonergic 5-HT 1B and 5-HT 1D receptors are expressed extensively in inner ear ganglia of monkeys and rats. The serotonergic 5-HT 1F receptor is also a target of triptans. This study describes its distribution in vestibular and trigeminal ganglia of monkeys. Using primary polyclonal antibodies raised against oligopeptides specific for the human 5-HT 1F receptor, neuronal somatic area and intensity of immunoreactive vestibular and trigeminal ganglia were quantified. Virtually all vestibular and considerable trigeminal ganglia showed positive 5-HT 1F receptor immunoreactivity. Inferior and superior vestibular ganglia staining appeared confined to distinct cell regions, varying considerably among cells of different sizes: more intense in small, punctate in some medium and regionally polarized in some large cells. Analyses of average somatic vestibular neuronal immunoreactive intensity identified mainly medium sized cells with high standard deviation of intensity corresponding to punctately stained cells. Less variability occurred in somatic intensity staining and cellular distribution among 5-HT 1F receptor immunopositive trigeminal ganglia. Most exhibited similar punctate staining patterns, higher mean somatic immunoreactive intensity and larger neuronal somatic size proportions per size distribution subpopulation compared to vestibular ganglia size distribution populations. Centrally directed vestibular ganglion neuronal processes, cochlear inner hair cells, vestibular hair cells and blood vessels in vestibular maculae and cristae were immunoreactive. The 5-HT 1F receptor expression in vestibular ganglia shows complex variable staining intensity patterns associated with cell size of immunopositive neurons, not seen in immunopositive trigeminal ganglia and not previously evident with 5-HT 1B and 5-HT 1D receptor subtype immunoreactivity in

  8. Differential responses to high-frequency electrical stimulation in ON and OFF retinal ganglion cells

    Science.gov (United States)

    Twyford, Perry; Cai, Changsi; Fried, Shelley

    2014-04-01

    Objective. The field of retinal prosthetics for artificial vision has advanced considerably in recent years, however clinical outcomes remain inconsistent. The performance of retinal prostheses is likely limited by the inability of electrical stimuli to preferentially activate different types of retinal ganglion cell (RGC). Approach. Here we examine the response of rabbit RGCs to high-frequency stimulation, using biphasic pulses applied at 2000 pulses per second. Responses were recorded using cell-attached patch clamp methods, and stimulation was applied epiretinally via a small cone electrode. Main results. When prolonged stimulus trains were applied to OFF-brisk transient (BT) RGCs, the cells exhibited a non-monotonic relationship between response strength and stimulus amplitude; this response pattern was different from those elicited previously by other electrical stimuli. When the amplitude of the stimulus was modulated transiently from a non-zero baseline amplitude, ON-BT and OFF-BT cells exhibited different activity patterns: ON cells showed an increase in activity while OFF cells exhibited a decrease in activity. Using a different envelope to modulate the amplitude of the stimulus, we observed the opposite effect: ON cells exhibited a decrease in activity while OFF cells show an increase in activity. Significance. As ON and OFF RGCs often exhibit opposing activity patterns in response to light stimulation, this work suggests that high-frequency electrical stimulation of RGCs may be able to elicit responses that are more physiological than traditional pulsatile stimuli. Additionally, the prospect of an electrical stimulus capable of cell-type specific selective activation has broad applications throughout the fields of neural stimulation and neuroprostheses.

  9. Nonselective Wiring Accounts for Red-Green Opponency in Midget Ganglion Cells of the Primate Retina.

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    Wool, Lauren E; Crook, Joanna D; Troy, John B; Packer, Orin S; Zaidi, Qasim; Dacey, Dennis M

    2018-02-07

    In primate retina, "red-green" color coding is initiated when signals originating in long (L) and middle (M) wavelength-sensitive cone photoreceptors interact antagonistically. The center-surround receptive field of "midget" ganglion cells provides the neural substrate for L versus M cone-opponent interaction, but the underlying circuitry remains unsettled, centering around the longstanding question of whether specialized cone wiring is present. To address this question, we measured the strength, sign, and spatial tuning of L- and M-cone input to midget receptive fields in the peripheral retina of macaque primates of either sex. Consistent with previous work, cone opponency arose when one of the cone types showed a stronger connection to the receptive field center than to the surround. We implemented a difference-of-Gaussians spatial receptive field model, incorporating known biology of the midget circuit, to test whether physiological responses we observed in real cells could be captured entirely by anatomical nonselectivity. When this model sampled nonselectively from a realistic cone mosaic, it accurately reproduced key features of a cone-opponent receptive field structure, and predicted both the variability and strength of cone opponency across the retina. The model introduced here is consistent with abundant anatomical evidence for nonselective wiring, explains both local and global properties of the midget population, and supports a role in their multiplexing of spatial and color information. It provides a neural basis for human chromatic sensitivity across the visual field, as well as the maintenance of normal color vision despite significant variability in the relative number of L and M cones across individuals. SIGNIFICANCE STATEMENT Red-green color vision is a hallmark of the human and nonhuman primate that starts in the retina with the presence of long (L)- and middle (M)-wavelength sensitive cone photoreceptor types. Understanding the underlying retinal

  10. Dp71Δ78-79 dystrophin mutant stimulates neurite outgrowth in PC12 cells via upregulation and phosphorylation of HspB1.

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    Merino-Jiménez, Candelaria; Aragón, Jorge; Ceja, Víctor; Rodríguez-Martínez, Griselda; Cázares-Raga, Febe E; Chardonnet, Solenne; Pionneau, Cédric; Rendon, Alvaro; Montañez, Cecilia

    2016-05-01

    PC12 cells acquire a neuronal phenotype in response to nerve growth factor (NGF). However, this phenotype is more efficiently achieved when the Dp71Δ78-79 dystrophin mutant is stably expressed in PC12-C11 cells. To investigate the effect of Dp71Δ78-79 overexpression on the protein profile of PC12-C11 cells, we compared the expression profiles of undifferentiated and NGF-differentiated PC12-C11 and PC12 cells by 2DE. In undifferentiated cultures, one protein was downregulated, and five were upregulated. Dp71Δ78-79 overexpression had a greater effect on differentiated cultures, with ten proteins downregulated and seven upregulated. The protein with the highest upregulation was HspB1. Changes in HspB1 expression were validated by Western blot and immunofluorescence analyses. Interestingly, the neurite outgrowth in PC12-C11 cells was affected by a polyclonal antibody against HspB1, and the level of HspB1 and HspB1Ser86 decreased, suggesting an important role for this protein in this cellular process. Our results show that Dp71Δ78-79 affects the expression level of some proteins and that the stimulated neurite outgrowth produced by this mutant is mainly through upregulation and phosphorylation of HspB1. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  11. Differential calcium signaling mediated by voltage-gated calcium channels in rat retinal ganglion cells and their unmyelinated axons.

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    Allison Sargoy

    Full Text Available Aberrant calcium regulation has been implicated as a causative factor in the degeneration of retinal ganglion cells (RGCs in numerous injury models of optic neuropathy. Since calcium has dual roles in maintaining homeostasis and triggering apoptotic pathways in healthy and injured cells, respectively, investigation of voltage-gated Ca channel (VGCC regulation as a potential strategy to reduce the loss of RGCs is warranted. The accessibility and structure of the retina provide advantages for the investigation of the mechanisms of calcium signalling in both the somata of ganglion cells as well as their unmyelinated axons. The goal of the present study was to determine the distribution of VGCC subtypes in the cell bodies and axons of ganglion cells in the normal retina and to define their contribution to calcium signals in these cellular compartments. We report L-type Ca channel α1C and α1D subunit immunoreactivity in rat RGC somata and axons. The N-type Ca channel α1B subunit was in RGC somata and axons, while the P/Q-type Ca channel α1A subunit was only in the RGC somata. We patch clamped isolated ganglion cells and biophysically identified T-type Ca channels. Calcium imaging studies of RGCs in wholemounted retinas showed that selective Ca channel antagonists reduced depolarization-evoked calcium signals mediated by L-, N-, P/Q- and T-type Ca channels in the cell bodies but only by L-type Ca channels in the axons. This differential contribution of VGCC subtypes to calcium signals in RGC somata and their axons may provide insight into the development of target-specific strategies to spare the loss of RGCs and their axons following injury.

  12. The Traditional Japanese Herbal Medicine Hachimijiogan Elicits Neurite Outgrowth Effects in PC12 Cells and Improves Cognitive in AD Model Rats via Phosphorylation of CREB

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    Kaori Kubota

    2017-11-01

    Full Text Available Hachimijiogan (HJG is a traditional herbal medicine that improves anxiety disorders in patients with dementia. In this study, we tested the hypothesis that HJG exerts neurotrophic factor-like effects to ameliorate memory impairment in Alzheimer disease (AD model rats. First, we describe that HJG acts to induce neurite outgrowth in PC12 cells (a rat pheochromocytoma cell line like nerve growth factor (NGF in a concentration-dependent manner (3 μg/ml HJG, p < 0.05; 10–500 μg/ml HJG, p < 0.001. While six herbal constituents of HJG, Rehmannia root, Dioscorea rhizome, Rhizoma Alismatis, Poria sclerotium, Moutan bark, and Cinnamon bark, could induce neurite outgrowth effects, the effect was strongest with HJG (500 μg/ml. Second, we demonstrated that HJG-induced neurite outgrowth was blocked by an inhibitor of cAMP response element binding protein (CREB, KG-501 (10 μM, p < 0.001. Moreover, HJG was observed to induce CREB phosphorylation 20–90 min after treatment (20 min, 2.50 ± 0.58-fold and CRE-mediated transcription in cultured PC12 cells (500 μg/ml, p < 0.01; 1000 μg/ml, p < 0.001. These results suggest a CREB-dependent mechanism underlies the neurotrophic effects of HJG. Finally, we examined improvements of memory impairment following HJG treatment using a Morris water maze in AD model animals (CI + Aβ rats. Repeated oral administration of HJG improved memory impairment (300 mg/kg, p < 0.05; 1000 mg/kg, p < 0.001 and induced CREB phosphorylation within the hippocampus (1000 mg/kg, p < 0.01. Together, our results suggest that HJG possesses neurotrophic effects similar to those of NGF, and can ameliorate cognitive dysfunction in a rat dementia model via CREB activation. Thus, HJG could potentially be a substitute for neurotrophic factors as a treatment for dementia.

  13. Isolation of Primary Murine Retinal Ganglion Cells (RGCs) by Flow Cytometry.

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    Chintalapudi, Sumana R; Patel, Need N; Goldsmith, Zachary K; Djenderedjian, Levon; Wang, Xiang Di; Marion, Tony N; Jablonski, Monica M; Morales-Tirado, Vanessa M

    2017-07-05

    Neurodegenerative diseases often have a devastating impact on those affected. Retinal ganglion cell (RGC) loss is implicated in an array of diseases, including diabetic retinopathy and glaucoma, in addition to normal aging. Despite their importance, RGCs have been extremely difficult to study until now due in part to the fact that they comprise only a small percentage of the wide variety of cells in the retina. In addition, current isolation methods use intracellular markers to identify RGCs, which produce non-viable cells. These techniques also involve lengthy isolation protocols, so there is a lack of practical, standardized, and dependable methods to obtain and isolate RGCs. This work describes an efficient, comprehensive, and reliable method to isolate primary RGCs from mice retinae using a protocol based on both positive and negative selection criteria. The presented methods allow for the future study of RGCs, with the goal of better understanding the major decline in visual acuity that results from the loss of functional RGCs in neurodegenerative diseases.

  14. Loss of Melanopsin-Expressing Retinal Ganglion Cells in Severely Staged Glaucoma Patients

    DEFF Research Database (Denmark)

    Obara, Elisabeth Anne; Hannibal, Jens; Heegaard, Steffen

    2016-01-01

    Purpose: Multiple studies have shown overwhelming evidence supporting the impairment of melanopsin function due to glaucoma. However, few studies have been carried out in humans analyzing the histology of melanopsin-expressing retinal ganglion cells (mRGCs) in retinas with glaucoma. The aim...... of this study was to analyze the pattern of expression of mRGCs relative to RGCs in the normal retina and retinas harboring varying stages of glaucoma. Methods: Paraffin-embedded human donor eyes with glaucoma (n = 11) and age-matched controls (n = 10) were obtained from Department of Pathology at Rigshospital...... difference was observed in mRGC expression in the normal retinas and mild-staged retinas with glaucoma; the densities of mRGCs were 3.08 ± 0.47 and 3.00 ± 0.13 cell counts/mm2, respectively. However, the severely staged retinas with glaucoma showed a significant loss in mRGCs density, 1.09 ± 0.35 cell counts...

  15. Methane rescues retinal ganglion cells and limits retinal mitochondrial dysfunction following optic nerve crush.

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    Wang, Ruobing; Sun, Qinglei; Xia, Fangzhou; Chen, Zeli; Wu, Jiangchun; Zhang, Yuelu; Xu, Jiajun; Liu, Lin

    2017-06-01

    Secondary degeneration is a common event in traumatic central nervous system disorders, which involves neuronal apoptosis and mitochondrial dysfunction. Exogenous methane exerts the therapeutic effects in many organ injury. Our study aims to investigate the potential neuroprotection of methane in a rat model of optic nerve crush (ONC). Adult male Sprague-Dawley rats were subjected to ONC and administrated intraperitoneally with methane-saturated or normal saline (10 ml/kg) once per day for one week after ONC. The retinal ganglion cells (RGCs) density was assessed by hematoxylin and eosin staining and Fluoro-Gold retrogradely labeling. Visual function was evaluated by flash visual evoked potentials (FVEP). The retinal apoptosis was measured by terminal-deoxy-transferase-mediated dUTP nick end labeling (TUNEL) assay and the expression of apoptosis-related factors, such as phosphorylated Bcl-2-associated death promoter (pBAD), phosphorylated glycogen synthase kinase-3β (pGSK-3β), Bcl-2 associated X protein (Bax) and Bcl-2 extra large (Bcl-xL). Retinal mitochondrial function was assessed by the mRNA expressions of peroxisome proliferator-activated receptor gamma coactivator-1α (PGC-1α), nuclear respiratory factor 1 (NRF1) and mitochondrial transcription factor A (TFAM), the mitochondrial DNA (mtDNA) copy number, citrate synthase activity and ATP content. Methane treatment significantly improved the RGC loss and visual dysfunction following ONC. As expected, methane also remarkably inhibited the retinal neural apoptosis, such as the fewer TUNEL-positive cells in ganglion cell layer, accompanied by the up-regulations of anti-apoptotic factors (pGSK-3β, pBAD, Bcl-xL) and the down-regulation of pro-apoptotic factor (Bax). Furthermore, methane treatment suppressed up-regulations of critical mitochondrial components (PGC-1α, NRF1 and TFAM) mRNA and mtDNA copy number, as well as improved the reduction of functional mitochondria markers, including citrate synthase

  16. Chagas' disease: selective affinity and cytotoxicity of Trypanosoma cruzi-immune lymphocytes to parasympathetic ganglion cells

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    Maria Lúcia Teixeira

    1980-12-01

    Full Text Available The megaesophagus and megacolon endemic in South America are related , to Chagas' disease. These mega conditions are found in patients with chronic Chagas's infection, when the parasite is not demonstrable in the lesions. These are characterized by depopulation of parasympathetic ganglion cells, dilation and hypertrophy of the viscera. In the experiments described here we deminstrate a selective affinity and adherence of Trypanosoma cruzi-immune lymphocytes to myenteric, parasympathetic ganglion cells, leading to neuronolysis. None of these features are observed when non-immune lymphocytes from control rabbits are used, or when the immune lymphocytes are allowed to react with CNS neurons. This demonstration is an indication of the high degree of specificity of the destruction of parasympathetic neurons in Chagas' disease. We postulate that the T. cruzi-immune lymphocyte rejection of parasympathetic neurons, but not of CNS neurons, might be related to recognition of a cross-reacting antigenic determinant secreted only by the target neurons. In favor of this interpretation is the observation of lymphocytic infiltrates and parasympathetic ganglion cell destruction in chronic Chagas' infection in the absence of encephalitis.O megaesôfago e o megacolon endêmicos na América do Sul estão relacionados á doença de Chagas. Estas condições clínicas são encontradas em pacientes com infecção chagásica crônica, quando o parasito não é demonstrado nas lesões caracterizadas por povoamento de células neuronais parassimpáticas. Nos experimentos descritos aqui nós demonstramos uma afinidade seletiva de linfócitos imunes, sensibilizados pelo T. cruzi, para neurônios de gânglios mioentéricos. Os linfócitos imunes citotóxicos aderem nas células ganglionares, produzindo neuronólise. Isto não se observa quando linfócitos não-imunes são usados, ou quando os linfócitos imunes são colocados na presença de neurônios do sistema nervoso

  17. Neuroprotective effect of peroxiredoxin 6 against hypoxia-induced retinal ganglion cell damage

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    Kumar Anil

    2010-10-01

    Full Text Available Abstract Background The ability to respond to changes in the extra-intracellular environment is prerequisite for cell survival. Cellular responses to the environment include elevating defense systems, such as the antioxidant defense system. Hypoxia-evoked reactive oxygen species (ROS-driven oxidative stress is an underlying mechanism of retinal ganglion cell (RGC death that leads to blinding disorders. The protein peroxiredoxin 6 (PRDX6 plays a pleiotropic role in negatively regulating death signaling in response to stressors, and thereby stabilizes cellular homeostasis. Results We have shown that RGCs exposed to hypoxia (1% or hypoxia mimetic cobalt chloride display reduced expression of PRDX6 with higher ROS expression and activation of NF-κB. These cells undergo apoptosis, while cells with over-expression of PRDX6 demonstrate resistance against hypoxia-driven RGC death. The RGCs exposed to hypoxia either with 1% oxygen or cobalt chloride (0-400 μM, revealed ~30%-70% apoptotic cell death after 48 and 72 h of exposure. Western analysis and real-time PCR showed elevated expression of PRDX6 during hypoxia at 24 h, while PRDX6 protein and mRNA expression declined from 48 h onwards following hypoxia exposure. Concomitant with this, RGCs showed increased ROS expression and activation of NF-κB with IkB phosphorylation/degradation, as examined with H2DCF-DA and transactivation assays. These hypoxia-induced adverse reactions could be reversed by over-expression of PRDX6. Conclusion Because an abundance of PRDX6 in cells was able to attenuate hypoxia-induced RGC death, the protein could possibly be developed as a novel therapeutic agent acting to postpone RGC injury and delay the progression of glaucoma and other disorders caused by the increased-ROS-generated death signaling related to hypoxia.

  18. ERK5 activity is required for nerve growth factor-induced neurite outgrowth and stabilization of tyrosine hydroxylase in PC12 cells.

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    Obara, Yutaro; Yamauchi, Arata; Takehara, Shin; Nemoto, Wataru; Takahashi, Maho; Stork, Philip J S; Nakahata, Norimichi

    2009-08-28

    Extracellular signal-regulated kinases (ERKs) play important physiological roles in proliferation, differentiation, and gene expression. ERK5 is approximately twice the size of ERK1/2, and its amino-terminal half contains the kinase domain that shares homology with ERK1/2 and TEY activation motif, whereas the carboxyl-terminal half is unique. In this study, we examined a physiological role of ERK5 in rat pheochromocytoma cells (PC12), comparing it with ERK1/2. Nerve growth factor (NGF) induced phosphorylation of both ERK5 and ERK1/2, whereas the cAMP analog dibutyryl cAMP (Bt(2)cAMP) caused only ERK1/2 phosphorylation. U0126, at 30 mum, that blocks ERK1/2 signaling selectively attenuated neurite outgrowth induced by NGF and Bt(2)cAMP, but BIX02188 and BIX02189, at 30 mum, that block ERK5 signaling and an ERK5 dominant-negative mutant suppressed only NGF-induced neurite outgrowth. Next, we examined the expression of tyrosine hydroxylase, a rate-limiting enzyme of catecholamine biosynthesis. Both NGF and Bt(2)cAMP increased tyrosine hydroxylase gene promoter activity in an ERK1/2-dependent manner but was ERK5-independent. However, when both ERK5 and ERK1/2 signalings were inhibited, tyrosine hydroxylase protein up-regulation by NGF and Bt(2)cAMP was abolished, because of the loss of stabilization of tyrosine hydroxylase protein by ERK5. Taking these results together, ERK5 is involved in neurite outgrowth and stabilization of tyrosine hydroxylase in PC12 cells, and ERK5, along with ERK1/2, plays essential roles in the neural differentiation process.

  19. The inverse F-BAR domain protein srGAP2 acts through srGAP3 to modulate neuronal differentiation and neurite outgrowth of mouse neuroblastoma cells.

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    Yue Ma

    Full Text Available The inverse F-BAR (IF-BAR domain proteins srGAP1, srGAP2 and srGAP3 are implicated in neuronal development and may be linked to mental retardation, schizophrenia and seizure. A partially overlapping expression pattern and highly similar protein structures indicate a functional redundancy of srGAPs in neuronal development. Our previous study suggests that srGAP3 negatively regulates neuronal differentiation in a Rac1-dependent manner in mouse Neuro2a cells. Here we show that exogenously expressed srGAP1 and srGAP2 are sufficient to inhibit valporic acid (VPA-induced neurite initiation and growth in the mouse Neuro2a cells. While ectopic- or over-expression of RhoGAP-defective mutants, srGAP1(R542A and srGAP2(R527A exert a visible inhibitory effect on neuronal differentiation. Unexpectedly, knockdown of endogenous srGAP2 fails to facilitate the neuronal differentiation induced by VPA, but promotes neurite outgrowth of differentiated cells. All three IF-BAR domains from srGAP1-3 can induce filopodia formation in Neuro2a, but the isolated IF-BAR domain from srGAP2, not from srGAP1 and srGAP3, can promote VPA-induced neurite initiation and neuronal differentiation. We identify biochemical and functional interactions of the three srGAPs family members. We propose that srGAP3-Rac1 signaling may be required for the effect of srGAP1 and srGAP2 on attenuating neuronal differentiation. Furthermore, inhibition of Slit-Robo interaction can phenocopy a loss-of-function of srGAP3, indicating that srGAP3 may be dedicated to the Slit-Robo pathway. Our results demonstrate the interplay between srGAP1, srGAP2 and srGAP3 regulates neuronal differentiation and neurite outgrowth. These findings may provide us new insights into the possible roles of srGAPs in neuronal development and a potential mechanism for neurodevelopmental diseases.

  20. APP upregulation contributes to retinal ganglion cell degeneration via JNK3.

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    Liu, Chao; Zhang, Cheng-Wu; Zhou, Yi; Wong, Wan Qing; Lee, Liying Corinne; Ong, Wei Yi; Yoon, Sung Ok; Hong, Wanjin; Fu, Xin-Yuan; Soong, Tuck Wah; Koo, Edward H; Stanton, Lawrence W; Lim, Kah-Leong; Xiao, Zhi-Cheng; Dawe, Gavin S

    2018-03-01

    Axonal injury is a common feature of central nervous system insults. Upregulation of amyloid precursor protein (APP) is observed following central nervous system neurotrauma and is regarded as a marker of central nervous system axonal injury. However, the underlying mechanism by which APP mediates neuronal death remains to be elucidated. Here, we used mouse optic nerve axotomy (ONA) to model central nervous system axonal injury replicating aspects of retinal ganglion cell (RGC) death in optic neuropathies. APP and APP intracellular domain (AICD) were upregulated in retina after ONA and APP knockout reduced Tuj1 + RGC loss. Pathway analysis of microarray data combined with chromatin immunoprecipitation and a luciferase reporter assay demonstrated that AICD interacts with the JNK3 gene locus and regulates JNK3 expression. Moreover, JNK3 was found to be upregulated after ONA and to contribute to Tuj1 + RGC death. APP knockout reduced the ONA-induced enhanced expression of JNK3 and phosphorylated JNK (pJNK). Gamma-secretase inhibitors prevented production of AICD, reduced JNK3 and pJNK expression similarly, and protected Tuj1 + RGCs from ONA-induced cell death. Together these data indicate that ONA induces APP expression and that gamma-secretase cleavage of APP releases AICD, which upregulates JNK3 leading to RGC death. This pathway may be a novel target for neuronal protection in optic neuropathies and other forms of neurotrauma.

  1. Gap junctions are essential for generating the correlated spike activity of neighboring retinal ganglion cells.

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    Béla Völgyi

    Full Text Available Neurons throughout the brain show spike activity that is temporally correlated to that expressed by their neighbors, yet the generating mechanism(s remains unclear. In the retina, ganglion cells (GCs show robust, concerted spiking that shapes the information transmitted to central targets. Here we report the synaptic circuits responsible for generating the different types of concerted spiking of GC neighbors in the mouse retina. The most precise concerted spiking was generated by reciprocal electrical coupling of GC neighbors via gap junctions, whereas indirect electrical coupling to a common cohort of amacrine cells generated the correlated activity with medium precision. In contrast, the correlated spiking with the lowest temporal precision was produced by shared synaptic inputs carrying photoreceptor noise. Overall, our results demonstrate that different synaptic circuits generate the discrete types of GC correlated activity. Moreover, our findings expand our understanding of the roles of gap junctions in the retina, showing that they are essential for generating all forms of concerted GC activity transmitted to central brain targets.

  2. Can Variability of Pattern ERG Signal Help to Detect Retinal Ganglion Cells Dysfunction in Glaucomatous Eyes?

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    Alberto Mavilio

    2015-01-01

    Full Text Available Objective. To evaluate variability of steady-state pattern electroretinogram (SS-PERG signal in normal, suspected, and glaucomatous eyes. Methods. Twenty-one subjects with suspected glaucoma due to disc abnormalities (GS, 37 patients with early glaucoma (EG, and 24 normal control (NC were tested with spectral-domain optical coherence tomography (SD-OCT, standard automated perimetry (SAP, and SS-PERG. Mean deviation (MD, pattern standard deviation (PSD, retinal nerve fiber layer (RNFL, and ganglionar complex cells (GCC were evaluated. The SS-PERG was recorded five consecutive times and the amplitude and phase of second harmonic were measured. PERG amplitude and coefficient of variation of phase (CVphase were recorded, and correlation with structural and functional parameters of disease, by means of one-way ANOVA and Pearson’s correlation, was analysed. Results. PERG amplitude was reduced, as expression of retinal ganglion cells (RGCs dysfunction, in EG patients and GS subjects compared to NC patients (P<0.0001. CVphase was significantly increased in EG patients and GS subjects, compared to healthy (P<0.0001, and it was also correlated with PSD (P=0.0009, GCC (P=0.028, and RNFL (P=0.0078 only in EG patients. Conclusions. Increased intrasession variability of phase in suspected glaucomatous eyes may be a sign of RGCs dysfunction.

  3. Retinal ganglion cell survival and axon regeneration after optic nerve injury in naked mole-rats.

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    Park, Kevin K; Luo, Xueting; Mooney, Skyler J; Yungher, Benjamin J; Belin, Stephane; Wang, Chen; Holmes, Melissa M; He, Zhigang

    2017-02-01

    In the adult mammalian central nervous system (CNS), axonal damage often triggers neuronal cell death and glial activation, with very limited spontaneous axon regeneration. In this study, we performed optic nerve injury in adult naked mole-rats, the longest living rodent, with a maximum life span exceeding 30 years, and found that injury responses in this species are quite distinct from those in other mammalian species. In contrast to what is seen in other mammals, the majority of injured retinal ganglion cells (RGCs) survive with relatively high spontaneous axon regeneration. Furthermore, injured RGCs display activated signal transducer and activator of transcription-3 (STAT3), whereas astrocytes in the optic nerve robustly occupy and fill the lesion area days after injury. These neuron-intrinsic and -extrinsic injury responses are reminiscent of those in "cold-blooded" animals, such as fish and amphibians, suggesting that the naked mole-rat is a powerful model for exploring the mechanisms of neuronal injury responses and axon regeneration in mammals. J. Comp. Neurol. 525:380-388, 2017. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

  4. A Pixel-Encoder Retinal Ganglion Cell with Spatially Offset Excitatory and Inhibitory Receptive Fields

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    Keith P. Johnson

    2018-02-01

    Full Text Available The spike trains of retinal ganglion cells (RGCs are the only source of visual information to the brain. Here, we genetically identify an RGC type in mice that functions as a pixel encoder and increases firing to light increments (PixON-RGC. PixON-RGCs have medium-sized dendritic arbors and non-canonical center-surround receptive fields. From their receptive field center, PixON-RGCs receive only excitatory input, which encodes contrast and spatial information linearly. From their receptive field surround, PixON-RGCs receive only inhibitory input, which is temporally matched to the excitatory center input. As a result, the firing rate of PixON-RGCs linearly encodes local image contrast. Spatially offset (i.e., truly lateral inhibition of PixON-RGCs arises from spiking GABAergic amacrine cells. The receptive field organization of PixON-RGCs is independent of stimulus wavelength (i.e., achromatic. PixON-RGCs project predominantly to the dorsal lateral geniculate nucleus (dLGN of the thalamus and likely contribute to visual perception.

  5. Neuroprotective effects of bis(7-tacrine against glutamate-induced retinal ganglion cells damage

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    Xu Zhi

    2010-03-01

    Full Text Available Abstract Background Glutamate-mediated excitotoxicity, primarily through N-methyl-D-aspartate (NMDA receptors, may be an important cause of retinal ganglion cells (RGCs death in glaucoma and several other retinal diseases. Bis(7-tacrine is a noncompetitive NMDA receptors antagonist that can prevent glutamate-induced hippocampal neurons damage. We tested the effects of bis(7-tacrine against glutamate-induced rat RGCs damage in vitro and in vivo. Results In cultured neonatal rats RGCs, the MTT assay showed that glutamate induced a concentration- and time-dependent toxicity. Bis(7-tacrine and memantine prevented glutamate-induced cell death in a concentration-dependent manner with IC50 values of 0.028 μM and 0.834 μM, respectively. The anti-apoptosis effects of bis(7-tacrine were confirmed by annexin V-FITC/PI staining. In vivo, TUNEL analysis and retrograde labeling analysis found that pretreatment with bis(7-tacrine(0.2 mg/kg induced a significant neuroprotective effect against glutamate-induced RGCs damage. Conclusions Our results showed that bis(7-tacrine had neuroprotective effects against glutamate-induced RGCs damage in vitro and in vivo, possibly through the drug's anti-NMDA receptor effects. These findings make bis(7-tacrine potentially useful for treating a variety of ischemic or traumatic retinopathies inclusive of glaucoma.

  6. Inner nuclear layer thickening is inversley proportional to retinal ganglion cell loss in optic neuritis.

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    Megha Kaushik

    Full Text Available AIM: To examine the relationship between retinal ganglion cell loss and changes in the inner nuclear layer (INL in optic neuritis (ON. METHODS: 36 multiple sclerosis (MS patients with a history of ON and 36 age and sex-matched controls underwent Optical Coherence Tomography. The paramacular retinal nerve fiber layer (RNFL, combined ganglion cell and inner plexiform layers (GCL/IPL and inner nuclear layer (INL thickness were measured at 36 points around the fovea. To remove inter-subject variability, the difference in thickness of each layer between the ON and fellow eye of each patient was calculated. A topographic analysis was conducted. RESULTS: The INL of the ON patients was thicker than the controls (42.9µm versus 39.6µm, p=0.002. ON patients also had a thinner RNFL (27.8µm versus 32.2µm, p<0.001 and GCL/IPL (69.3µm versus 98.1µm, p<0.001. Among the controls, there was no correlation between RNFL and GCL/IPL as well as RNFL and INL, but a positive correlation was seen between GCL/IPL and INL (r=0.65, p<0.001. In the ON group, there was a positive correlation between RNFL and GCL/IPL (r=0.80, p<0.001 but a negative correlation between RNFL and INL (r=-0.61, p<0.001 as well as GCL/IPL and INL (r=-0.44, p=0.007. The negative correlation between GCL/IPL and INL strengthened in the ON group when inter-subject variability was removed (r=-0.75, p<0.001. Microcysts within the INL were present in 5 ON patients, mainly in the superior and infero-nasal paramacular regions. While patients with microcysts lay at the far end of the correlation curve between GCL/IPL and INL (i.e. larger INL and smaller GCL/IPL compared to other patients, their exclusion did not affect the correlation (r= -0.76, p<0.001. CONCLUSIONS: INL enlargement in MS-related ON is associated with the severity of GCL loss. This is a continuous relationship and patients with INL microcysts may represent the extreme end of the scale.

  7. c-Jun N-terminal kinase 3 expression in the retina of ocular hypertension mice: a possible target to reduce ganglion cell apoptosis

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    Yue He

    2015-01-01

    Full Text Available Glaucoma, a type of optic neuropathy, is characterized by the loss of retinal ganglion cells. It remains controversial whether c-Jun N-terminal kinase (JNK participates in the apoptosis of retinal ganglion cells in glaucoma. This study sought to explore a possible mechanism of action of JNK signaling pathway in glaucoma-induced retinal optic nerve damage. We established a mouse model of chronic ocular hypertension by reducing the aqueous humor followed by photocoagulation using the laser ignition method. Results showed significant pathological changes in the ocular tissues after the injury. Apoptosis of retinal ganglion cells increased with increased intraocular pressure, as did JNK3 mRNA expression in the retina. These data indicated that the increased expression of JNK3 mRNA was strongly associated with the increase in intraocular pressure in the retina, and correlated positively with the apoptosis of retinal ganglion cells.

  8. Gene transfection mediated by polyethyleneimine-polyethylene glycol nanocarrier prevents cisplatin-induced spiral ganglion cell damage

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    Guan-gui Chen

    2015-01-01

    Full Text Available Polyethyleneimine-polyethylene glycol (PEI-PEG, a novel nanocarrier, has been used for transfection and gene therapy in a variety of cells. In our previous study, we successfully carried out PEI-PEG-mediated gene transfer in spiral ganglion cells. It remains unclear whether PEI-PEG could be used for gene therapy with X-linked inhibitor of apoptosis protein (XIAP in the inner ear. In the present study, we performed PEI-PEG-mediated XIAP gene transfection in the cochlea of Sprague-Dawley rats, via scala tympani fenestration, before daily cisplatin injections. Auditory brainstem reflex tests demonstrated the protective effects of XIAP gene therapy on auditory function. Immunohistochemical staining revealed XIAP protein expression in the cytoplasm of cells in the spiral ganglion, the organ of Corti and the stria vascularis. Reverse transcription-PCR detected high levels of XIAP mRNA expression in the cochlea. The present findings suggest that PEI-PEG nanocarrier-mediated XIAP gene transfection results in XIAP expression in the cochlea, prevents damage to cochlear spiral ganglion cells, and protects hearing.

  9. Retinal Ganglion Cell Loss in X-linked Adrenoleukodystrophy with anABCD1Mutation (Gly266Arg).

    Science.gov (United States)

    Ohkuma, Yasuhiro; Hayashi, Takaaki; Yoshimine, Syouyou; Tsuneoka, Hiroshi; Terao, Yoko; Akiyama, Masaharu; Ida, Hiroyuki; Ohashi, Toya; Okumura, Akihisa; Ebihara, Nobuyuki; Murakami, Akira; Shimozawa, Nobuyuki

    2014-01-01

    The authors here report a single case of a 10-year-old male patient who presented with severe vision loss associated with progressive demyelination. The patient was diagnosed with X-linked childhood cerebral adrenoleukodystrophy (ALD). Genetic analysis demonstrated a missense mutation (Gly266Arg) in exon 1 of the ABCD1 gene. His corrected visual acuity confirmed the absolute lack of light perception in both eyes. Funduscopy revealed severe pallor of the optic disc in both eyes. Spectral-domain optical coherence tomography showed thinning of the retinal ganglion cell and inner plexiform layers (GCL and IPL). Thinning of the GCL and IPL may be due to transneuronal retrograde degeneration of ganglion cells secondary to optic tract demyelination.

  10. Cortical Proteins are Chemokinetic to Cells from the Medial Ganglionic Eminence

    Science.gov (United States)

    2011-05-28

    Neuroscience Program Director During embryonic development, a majority of neocortical interneurons originate from the medial ganglionic eminence (MGE...day vaginal plug is seen) EGF Epidermal growth factor ErbB EGF receptor GABA Gamma-aminobutyric acid GE Ganglionic eminence HGF Hepatocyte...species, all mammals have GABAergic precursors residing in the GE producing neurons that migrate along the same tangential route to the neocortex

  11. Neural cell adhesion molecule-stimulated neurite outgrowth depends on activation of protein kinase C and the Ras-mitogen-activated protein kinase pathway

    DEFF Research Database (Denmark)

    Kolkova, K; Novitskaya, V; Pedersen, N

    2000-01-01

    transfected with expression plasmids encoding constitutively active forms of Ras, Raf, MAP kinase kinases MEK1 and 2, dominant negative forms of Ras and Raf, and the FAK-related nonkinase. Alternatively, PC12-E2 cells were submitted to treatment with antibodies to the fibroblast growth factor (FGF) receptor......, inhibitors of the nonreceptor tyrosine kinase p59(fyn), PLC, PKC and MEK and an activator of PKC, phorbol-12-myristate-13-acetate (PMA). MEK2 transfection rescued cells treated with all inhibitors. The same was found for PMA treatment, except when cells concomitantly were treated with the MEK inhibitor....... Arachidonic acid rescued cells treated with antibodies to the FGF receptor or the PLC inhibitor, but not cells in which the activity of PKC, p59(fyn), FAK, Ras, or MEK was inhibited. Interaction of NCAM with a synthetic NCAM peptide ligand, known to induce neurite outgrowth, was shown to stimulate...

  12. Electrical receptive fields of retinal ganglion cells: Influence of presynaptic neurons.

    Science.gov (United States)

    Maturana, Matias I; Apollo, Nicholas V; Garrett, David J; Kameneva, Tatiana; Cloherty, Shaun L; Grayden, David B; Burkitt, Anthony N; Ibbotson, Michael R; Meffin, Hamish

    2018-02-01

    Implantable retinal stimulators activate surviving neurons to restore a sense of vision in people who have lost their photoreceptors through degenerative diseases. Complex spatial and temporal interactions occur in the retina during multi-electrode stimulation. Due to these complexities, most existing implants activate only a few electrodes at a time, limiting the repertoire of available stimulation patterns. Measuring the spatiotemporal interactions between electrodes and retinal cells, and incorporating them into a model may lead to improved stimulation algorithms that exploit the interactions. Here, we present a computational model that accurately predicts both the spatial and temporal nonlinear interactions of multi-electrode stimulation of rat retinal ganglion cells (RGCs). The model was verified using in vitro recordings of ON, OFF, and ON-OFF RGCs in response to subretinal multi-electrode stimulation with biphasic pulses at three stimulation frequencies (10, 20, 30 Hz). The model gives an estimate of each cell's spatiotemporal electrical receptive fields (ERFs); i.e., the pattern of stimulation leading to excitation or suppression in the neuron. All cells had excitatory ERFs and many also had suppressive sub-regions of their ERFs. We show that the nonlinearities in observed responses arise largely from activation of presynaptic interneurons. When synaptic transmission was blocked, the number of sub-regions of the ERF was reduced, usually to a single excitatory ERF. This suggests that direct cell activation can be modeled accurately by a one-dimensional model with linear interactions between electrodes, whereas indirect stimulation due to summated presynaptic responses is nonlinear.

  13. Correspondence between visual and electrical input filters of ON and OFF mouse retinal ganglion cells

    Science.gov (United States)

    Sekhar, S.; Jalligampala, A.; Zrenner, E.; Rathbun, D. L.

    2017-08-01

    Objective. Over the past two decades retinal prostheses have made major strides in restoring functional vision to patients blinded by diseases such as retinitis pigmentosa. Presently, implants use single pulses to activate the retina. Though this stimulation paradigm has proved beneficial to patients, an unresolved problem is the inability to selectively stimulate the on and off visual pathways. To this end our goal was to test, using white noise, voltage-controlled, cathodic, monophasic pulse stimulation, whether different retinal ganglion cell (RGC) types in the wild type retina have different electrical input filters. This is an important precursor to addressing pathway-selective stimulation. Approach. Using full-field visual flash and electrical and visual Gaussian noise stimulation, combined with the technique of spike-triggered averaging (STA), we calculate the electrical and visual input filters for different types of RGCs (classified as on, off or on-off based on their response to the flash stimuli). Main results. Examining the STAs, we found that the spiking activity of on cells during electrical stimulation correlates with a decrease in the voltage magnitude preceding a spike, while the spiking activity of off cells correlates with an increase in the voltage preceding a spike. No electrical preference was found for on-off cells. Comparing STAs of wild type and rd10 mice revealed narrower electrical STA deflections with shorter latencies in rd10. Significance. This study is the first comparison of visual cell types and their corresponding temporal electrical input filters in the retina. The altered input filters in degenerated rd10 retinas are consistent with photoreceptor stimulation underlying visual type-specific electrical STA shapes in wild type retina. It is therefore conceivable that existing implants could target partially degenerated photoreceptors that have only lost their outer segments, but not somas, to selectively activate the on and off

  14. Analysis the macular ganglion cell complex thickness in monocular strabismic amblyopia patients by Fourier-domain OCT

    Directory of Open Access Journals (Sweden)

    Hong-Wei Deng

    2014-11-01

    Full Text Available AIM: To detect the macular ganglion cell complex thickness in monocular strabismus amblyopia patients, in order to explore the relationship between the degree of amblyopia and retinal ganglion cell complex thickness, and found out whether there is abnormal macular ganglion cell structure in strabismic amblyopia. METHODS: Using a fourier-domain optical coherence tomography(FD-OCTinstrument iVue®(Optovue Inc, Fremont, CA, Macular ganglion cell complex(mGCCthickness was measured and statistical the relation rate with the best vision acuity correction was compared Gman among 26 patients(52 eyesincluded in this study. RESULTS: The mean thickness of the mGCC in macular was investigated into three parts: centrial, inner circle(3mmand outer circle(6mm. The mean thicknesses of mGCC in central, inner and outer circle was 50.74±21.51μm, 101.4±8.51μm, 114.2±9.455μm in the strabismic amblyopia eyes(SAE, and 43.79±11.92μm,92.47±25.01μm, 113.3±12.88μm in the contralateral sound eyes(CSErespectively. There was no statistically significant difference among the eyes(P>0.05. But the best corrected vision acuity had a good correlation rate between mGcc thicknesses, which was better relative for the lower part than the upper part.CONCLUSION:There is a relationship between the amblyopia vision acuity and the mGCC thickness. Although there has not statistically significant difference of the mGCC thickness compared with the SAE and CSE. To measure the macular center mGCC thickness in clinic may understand the degree of amblyopia.

  15. Lentiviral-mediated transfer of CNTF to schwann cells within reconstructed peripheral nerve grafts enhances adult retinal ganglion cell survival and axonal regeneration

    NARCIS (Netherlands)

    Hu, Ying; Leaver, Simone G; Plant, Giles W; Hendriks, William T J; Niclou, Simone P; Verhaagen, J.; Harvey, Alan R; Cui, Qi

    We recently described a method for reconstituting peripheral nerve (PN) sheaths using adult Schwann cells (SCs). Reconstructed PN tissue grafted onto the cut optic nerve supports the regeneration of injured adult rat retinal ganglion cell (RGC) axons. To determine whether genetic manipulation of

  16. Developmental markers of ganglion cells in the enteric nervous system and their application for evaluation of Hirschsprung disease.

    Science.gov (United States)

    Kawai, Hitomi; Satomi, Kaishi; Morishita, Yukio; Murata, Yoshihiko; Sugano, Masato; Nakano, Noriyuki; Noguchi, Masayuki

    2014-09-01

    Hirschsprung disease (HSCR) is a congenital disease resulting from failure of neural crest-derived ganglion cells to colonize the colon. Conventional diagnostic methods are insufficient for evaluating the 'functional' prognosis of HSCR. In order to elucidate the maturation of ganglion cells, 17 immunohistochemical markers were examined. We examined the digestive tracts of 2 human early delivery patients, 2 miniature swine fetuses, 4 little infants, 3 infants, 3 children, 6 adults, and 3 aged individuals. With increasing age, the labeling index (LI) for both calretinin and tyrosine hydroxylase (TH) increased, whereas that for SOX10 decreased. We then examined the 'transitional zone' of HSCR in 21 affected patients and 18 controls for these three markers. The LI of calretinin and TH were significantly lower than in the controls (median: 3.7 in HSCR and 8.2 in controls, P ganglion cells are present in the transitional zone of HSCR, and that HSCR may have two different pathophysiological processes. © 2014 Japanese Society of Pathology and Wiley Publishing Asia Pty Ltd.

  17. Ezh2 does not mediate retinal ganglion cell homeostasis or their susceptibility to injury.

    Directory of Open Access Journals (Sweden)

    Lin Cheng

    Full Text Available Epigenetic predisposition is thought to critically contribute to adult-onset disorders, such as retinal neurodegeneration. The histone methyltransferase, enhancer of zeste homolog 2 (Ezh2, is transiently expressed in the perinatal retina, particularly enriched in retinal ganglion cells (RGCs. We previously showed that embryonic deletion of Ezh2 from retinal progenitors led to progressive photoreceptor degeneration throughout life, demonstrating a role for embryonic predisposition of Ezh2-mediated repressive mark in maintaining the survival and function of photoreceptors in the adult. Enrichment of Ezh2 in RGCs leads to the question if Ezh2 also mediates gene expression and function in postnatal RGCs, and if its deficiency changes RGC susceptibility to cell death under injury or disease in the adult. To test this, we generated mice carrying targeted deletion of Ezh2 from RGC progenitors driven by Math5-Cre (mKO. mKO mice showed no detectable defect in RGC development, survival, or cell homeostasis as determined by physiological analysis, live imaging, histology, and immunohistochemistry. Moreover, RGCs of Ezh2 deficient mice revealed similar susceptibility against glaucomatous and acute optic nerve trauma-induced neurodegeneration compared to littermate floxed or wild-type control mice. In agreement with the above findings, analysis of RNA sequencing of RGCs purified from Ezh2 deficient mice revealed few gene changes that were related to RGC development, survival and function. These results, together with our previous report, support a cell lineage-specific mechanism of Ezh2-mediated gene repression, especially those critically involved in cellular function and homeostasis.

  18. Rac1 selective activation improves retina ganglion cell survival and regeneration.

    Directory of Open Access Journals (Sweden)

    Erika Lorenzetto

    Full Text Available In adult mammals, after optic nerve injury, retinal ganglion cells (RGCs do not regenerate their axons and most of them die by apoptosis within a few days. Recently, several strategies that activate neuronal intracellular pathways were proposed to prevent such degenerative processes. The rho-related small GTPase Rac1 is part of a complex, still not fully understood, intracellular signaling network, mediating in neurons many effects, including axon growth and cell survival. However, its role in neuronal survival and regeneration in vivo has not yet been properly investigated. To address this point we intravitreally injected selective cell-penetrating Rac1 mutants after optic nerve crush and studied the effect on RGC survival and axonal regeneration. We injected two well-characterized L61 constitutively active Tat-Rac1 fusion protein mutants, in which a second F37A or Y40C mutation confers selectivity in downstream signaling pathways. Results showed that, 15 days after crush, both mutants were able to improve survival and to prevent dendrite degeneration, while the one harboring the F37A mutation also improved axonal regeneration. The treatment with F37A mutant for one month did not improve the axonal elongation respect to 15 days. Furthermore, we found an increase of Pak1 T212 phosphorylation and ERK1/2 expression in RGCs after F37A treatment, whereas ERK1/2 was more activated in glial cells after Y40C administration. Our data suggest that the selective activation of distinct Rac1-dependent pathways could represent a therapeutic strategy to counteract neuronal degenerative processes in the retina.

  19. Neuroprotective Effect of Lutein on NMDA-Induced Retinal Ganglion Cell Injury in Rat Retina.

    Science.gov (United States)

    Zhang, Chanjuan; Wang, Zhen; Zhao, Jiayi; Li, Qin; Huang, Cuiqin; Zhu, Lihong; Lu, Daxiang

    2016-05-01

    Lutein injection is a possible therapeutic approach for retinal diseases, but the molecular mechanism of its neuroprotective effect remains to be elucidated. The aim of this study was to investigate its protective effects in retinal ganglion cells (RGCs) against N-methyl-D-aspartate (NMDA)-induced retinal damage in vivo. Retinal damage was induced by intravitreal NMDA injection in rats. Each animal was given five daily intraperitoneal injections of Lutein or vehicle along with intravitreal NMDA injections. Electroretinograms were recorded. The number of viable RGCs was quantified using the retinal whole-mount method by immunofluorescence. Proteins were measured by Western blot assays. Lutein reduced the retinal damage and improved the response to light, as shown by an animal behavior assay (the black-and-white box method) in rats. Furthermore, Lutein treatment prevented the NMDA-induced reduction in phNR wave amplitude. Lutein increased RGC number after NMDA-induced retina damage. Most importantly, Bax, cytochrome c, p-p38 MAPK, and p-c-Jun were all upregulated in rats injected with NMDA, but these expression patterns were reversed by continuous Lutein uptake. Bcl-2, p-GSK-3β, and p-Akt in the Lutein-treated eyes were increased compared with the NMDA group. Lutein has neuroprotective effects against retinal damage, its protective effects may be partly mediated by its anti-excitability neurotoxicity, through MAPKs and PI3K/Akt signaling, suggesting a potential approach for suppressing retinal neural damage.

  20. Rasagiline-induced delay of retinal ganglion cell death in experimental glaucoma in rats.

    Science.gov (United States)

    Levkovitch-Verbin, Hani; Vander, Shelly; Melamed, Shlomo

    2011-01-01

    To evaluate the neuroprotective effect of rasagiline (N-propargyl-1 (R)-aminoindan), a selective monoamine oxidase inhibitor, on the survival of retinal ganglion cells (RGCs) in glaucomatous rat eyes. Rasagiline is an FDA approved anti-Parkinson disease drug with neuroprotective capabilities that were shown in many models of brain damage. The neuroprotective effect of daily intraperitoneal (IP) injections of rasagiline (0.5 mg/kg and 3 mg/kg) was evaluated and compared with saline injections using the translimbal photocoagulation model of experimental glaucoma in Wistar rats. Intraocular pressure (IOP) was measured before and immediately after the laser treatment, and then weekly. Seven weeks after the induction of glaucoma, the animals were killed, the eyes were enucleated and the retinas were prepared as whole mounts. Fluoro-gold had been injected into the superior colliculus 10 days before enucleation, and RGC survival was evaluated by counting the surviving labeled RGCs in a masked way. All rats (n=29) displayed significant IOP elevation and RGC damage. Seven weeks after the induction of glaucoma, the mean RGC survival was 43±8% in the rasagiline 3 mg/kg-treated group and 43±9% in the rasagiline 0.5 mg/kg-treated group compared with 23%±4% in the saline-treated (control) group (P=0.01 and P=0.02, respectively). Systemic treatment with rasagiline significantly enhances the survival of RGCs in experimental glaucoma.

  1. Trimetazidine protects retinal ganglion cells from acute glaucoma via the Nrf2/Ho-1 pathway.

    Science.gov (United States)

    Wan, Peixing; Su, Wenru; Zhang, Yingying; Li, Zhidong; Deng, Caibin; Zhuo, Yehong

    2017-09-15

    Acute glaucoma is one of the leading causes of irreversible vision impairment characterized by the rapid elevation of intraocular pressure (IOP) and consequent retinal ganglion cell (RGC) death. Oxidative stress and neuroinflammation have been considered critical for the pathogenesis of RGC death in acute glaucoma. Trimetazidine (TMZ), an anti-ischemic drug, possesses antioxidative and anti-inflammatory properties, contributing to its therapeutic potential in tissue damage. However, the role of TMZ in acute glaucoma and the underlying molecular mechanisms remain elusive. Here, we report that treatment with TMZ significantly attenuated retinal damage and RGC death in mice with acute glaucoma, with a significant decrease in reactive oxygen species (ROS) and inflammatory cytokine production in the retina. Furthermore, TMZ treatment directly decreased ROS production and rebalanced the intracellular redox state, thus contributing to the survival of RGCs in vitro TMZ treatment also reduced the production of inflammatory cytokines in vitro Mechanistically, the TMZ-mediated inhibition of apoptosis and inflammatory cytokine production in RGCs occurred via the regulation of the nuclear factor erythroid 2-related factor 2/heme oxygenase 1/caspase-8 pathway. Moreover, the TMZ-mediated neuroprotection in acute glaucoma was abrogated when an HO-1 inhibitor, SnPP, was used. Our findings identify potential mechanisms of RGC apoptosis and propose a novel therapeutic agent, TMZ, which exerts a precise neuroprotective effect against acute glaucoma. © 2017 The Author(s).

  2. Losartan Treatment Protects Retinal Ganglion Cells and Alters Scleral Remodeling in Experimental Glaucoma

    Science.gov (United States)

    Pitha, Ian F.; Nguyen, Cathy; Steinhart, Matthew R.; Nguyen, Thao D.; Pease, Mary Ellen; Oglesby, Ericka N.; Berlinicke, Cynthia A.; Mitchell, Katherine L.; Kim, Jessica; Jefferys, Joan J.

    2015-01-01

    Purpose To determine if oral losartan treatment decreases the retinal ganglion cell (RGC) death caused by experimental intraocular pressure (IOP) elevation in mice. Methods We produced IOP increase in CD1 mice and performed unilateral optic nerve crush. Mice received oral losartan, spironolactone, enalapril, or no drug to test effects of inhibiting angiotensin receptors. IOP was monitored by Tonolab, and blood pressure was monitored by tail cuff device. RGC loss was measured in masked axon counts and RGC bodies by β-tubulin labeling. Scleral changes that could modulate RGC injury were measured including axial length, scleral thickness, and retinal layer thicknesses, pressure-strain behavior in inflation testing, and study of angiotensin receptors and pathways by reverse transcription polymerase chain reaction, Western blot, and immunohistochemistry. Results Losartan treatment prevented significant RGC loss (median loss = 2.5%, p = 0.13), while median loss with water, spironolactone, and enalapril treatments were 26%, 28% and 43%; p glaucoma eyes (p = 0.007). Losartan inhibited effects of glaucoma, including reduction in extracellular signal-related kinase activity and modification of glaucoma-related changes in scleral thickness and creep under controlled IOP. Conclusions The neuroprotective effect of losartan in mouse glaucoma is associated with adaptive changes in the sclera expressed at the optic nerve head. PMID:26505191

  3. The identification of ganglion cells in Hirschsprung disease by the immunohistochemical detection of ret oncoprotein.

    Science.gov (United States)

    Karim, Sadiqa; Hession, Colleen; Marconi, Sharon; Gang, David L; Otis, Christopher N

    2006-07-01

    The absence of ganglion cells (GCs) is the primary anatomic abnormality in Hirschsprung disease. Light microscopy is the mainstay in establishing this diagnosis. However, establishing a condition of aganglionosis may be challenging on routine H&E-stained sections of colonic biopsies and resections. We studied the identification of GCs by retinoblastoma oncoprotein (ret) immunoreactivity and routine H&E light microscopy by evaluating 53 blocks from 34 patients demonstrating GCs on original H&E-stained sections and 55 blocks from 38 patients lacking GCs on original H&E-stained sections. All blocks demonstrating GCs on H&E-stained sections also were positive for GCs on ret staining (100%). In 3 blocks that were negative for GCs by H&E staining (5%), GCs were shown on ret-stained sections. Immunoreactivity for ret has comparable specificity but slightly higher sensitivity to routine light microscopic evaluation in identifying GCs. GCs are identified more readily by ret immunoreactivity than by routine morphologic examination.

  4. Elevated intracranial pressure causes optic nerve and retinal ganglion cell degeneration in mice.

    Science.gov (United States)

    Nusbaum, Derek M; Wu, Samuel M; Frankfort, Benjamin J

    2015-07-01

    The purpose of this study was to develop a novel experimental system for the modulation and measurement of intracranial pressure (ICP), and to use this system to assess the impact of elevated ICP on the optic nerve and retinal ganglion cells (RGCs) in CD1 mice. This system involved surgical implantation of an infusion cannula and a radiowave based pressure monitoring probe through the skull and into the subarachnoid space. The infusion cannula was used to increase ICP, which was measured by the probe and transmitted to a nearby receiver. The system provided robust and consistent ICP waveforms, was well tolerated, and was stable over time. ICP was elevated to approximately 30 mmHg for one week, after which we assessed changes in optic nerve structure with transmission electron microscopy in cross section and RGC numbers with antibody staining in retinal flat mounts. ICP elevation resulted in optic nerve axonal loss and disorganization, as well as RGC soma loss. We conclude that the controlled manipulation of ICP in active, awake mice is possible, despite their small size. Furthermore, ICP elevation results in visual system phenotypes of optic nerve and RGC degeneration, suggesting that this model can be used to study the impact of ICP on the visual system. Potentially, this model can also be used to study the relationship between ICP and IOP, as well diseases impacted by ICP variation such as glaucoma, idiopathic intracranial hypertension, and the spaceflight-related visual impairment intracranial pressure syndrome. Copyright © 2015 Elsevier Ltd. All rights reserved.

  5. Sox11 Expression Promotes Regeneration of Some Retinal Ganglion Cell Types but Kills Others.

    Science.gov (United States)

    Norsworthy, Michael W; Bei, Fengfeng; Kawaguchi, Riki; Wang, Qing; Tran, Nicholas M; Li, Yi; Brommer, Benedikt; Zhang, Yiming; Wang, Chen; Sanes, Joshua R; Coppola, Giovanni; He, Zhigang

    2017-06-21

    At least 30 types of retinal ganglion cells (RGCs) send distinct messages through the optic nerve to the brain. Available strategies of promoting axon regeneration act on only some of these types. Here we tested the hypothesis that overexpressing developmentally important transcription factors in adult RGCs could reprogram them to a "youthful" growth-competent state and promote regeneration of other types. From a screen of transcription factors, we identified Sox11 as one that could induce substantial axon regeneration. Transcriptome profiling indicated that Sox11 activates genes involved in cytoskeletal remodeling and axon growth. Remarkably, α-RGCs, which preferentially regenerate following treatments such as Pten deletion, were killed by Sox11 overexpression. Thus, Sox11 promotes regeneration of non-α-RGCs, which are refractory to Pten deletion-induced regeneration. We conclude that Sox11 can reprogram adult RGCs to a growth-competent state, suggesting that different growth-promoting interventions promote regeneration in distinct neuronal types. Copyright © 2017 Elsevier Inc. All rights reserved.

  6. Segregation of ipsilateral retinal ganglion cell axons at the optic chiasm requires the Shh receptor Boc.

    Science.gov (United States)

    Fabre, Pierre J; Shimogori, Tomomi; Charron, Frédéric

    2010-01-06

    The pattern of contralaterally and ipsilaterally projecting retinal ganglion cell (RGC) axons at the optic chiasm is essential for the establishment of binocular vision. Contralateral axons cross the chiasm midline as they progress from the optic nerve to the optic tract. In contrast, ipsilateral axons deviate from the chiasm and continue in the ipsilateral optic tract, avoiding the chiasm midline. The molecular mechanism underlying this phenomenon is not completely understood. Here we show that the Sonic Hedgehog (Shh) receptor Boc is enriched in ipsilateral RGCs of the developing retina. Together with the presence of Shh at the midline, this complementary expression pattern led us to hypothesize that Shh might repel ipsilateral RGC axons at the chiasm. Consistent with this hypothesis, we found that only Boc-positive RGC axons retract in vitro in response to Shh and that this response is lost in Boc mutant RGCs. In vivo, we show that Boc is required for the normal segregation of ipsilateral axons at the optic chiasm and, conversely, that Boc expression in contralateral RGCs prevents their axons from crossing the optic chiasm. Together, these results suggest that Shh repels ipsilateral RGC axons at the optic chiasm via its receptor Boc. This work identifies a novel molecular pathway required for the segregation of axons at the optic chiasm.

  7. Distinctive effect on nerve growth factor-induced PC12 cell neurite outgrowth by two unique neolignan enantiomers from Illicium merrillianum

    Science.gov (United States)

    Tian, Xinhui; Yue, Rongcai; Zeng, Huawu; Li, Honglin; Shan, Lei; He, Weiwei; Shen, Yunheng; Zhang, Weidong

    2015-11-01

    Merrillianoid (1), a racemic neolignan possessing the characteristic benzo-2,7-dioxabicyclo[3.2.1]octane moiety, was isolated from the branches and leaves of Illicium merrillianum. Chiral separation of 1 gave two enantiomers (+)-1 and (-)-1. The structure of 1 was established by comprehensive spectroscopic analysis and single crystal X-ray diffraction. The absolute configurations of enantiomers were determined by quantum mechanical calculation. Compound (+)-1 exhibited a better neurotrophic activity than racemate 1 by promoting nerve growth factor (NGF) induced PC12 cell neurite outgrowth, while (-)-1 showed a distinctive inhibitory effect. Furthermore, a mechanism study indicated that the two enantiomers influenced NGF-induced neurite outgrowth of PC12 cells possibly by interacting with the trkA receptor, and extracellular signal regulated kinases 1/2 (ERK1/2) and mitogen-activated protein kinase (MEK) in Ras/ERK signal cascade. But the phosphorylation level of serine/threonine kinase Akt1 and Akt2 in PI3K/Akt signal pathway showed no significant difference between (+)-1 and (-)-1.

  8. Hydrogel Design for Supporting Neurite Outgrowth and Promoting Gene Delivery to Maximize Neurite Extension

    Science.gov (United States)

    Shepard, Jaclyn A.; Stevans, Alyson C.; Holland, Samantha; Wang, Christine E.; Shikanov, Ariella; Shea, Lonnie D.

    2012-01-01

    Hydrogels capable of gene delivery provide a combinatorial approach for nerve regeneration, with the hydrogel supporting neurite outgrowth and gene delivery inducing the expression of inductive factors. This report investigates the design of hydrogels that balance the requirements for supporting neurite growth with those requirements for promoting gene delivery. Enzymatically-degradable PEG hydrogels encapsulating dorsal root ganglia explants, fibroblasts, and lipoplexes encoding nerve growth factor were gelled within channels that can physically guide neurite outgrowth. Transfection of fibroblasts increased with increasing concentration of Arg-Gly-Asp (RGD) cell adhesion sites and decreasing PEG content. The neurite length increased with increasing RGD concentration within 10% PEG hydrogels, yet was maximal within 7.5% PEG hydrogels at intermediate RGD levels. Delivering lipoplexes within the gel produced longer neurites than culture in NGF-supplemented media or co-culture with cells exposed to DNA prior to encapsulation. Hydrogels designed to support neurite outgrowth and deliver gene therapy vectors locally may ultimately be employed to address multiple barriers that limit regeneration. PMID:22038654

  9. Protection of pattern electroretinogram and retinal ganglion cells by oncostatin M after optic nerve injury.

    Directory of Open Access Journals (Sweden)

    Xin Xia

    Full Text Available Injury to retinal ganglion cell (RGC axons leads to selective loss of RGCs and vision. Previous studies have shown that exogenous neurotrophic factors promote RGC survival. We investigated the neuroprotective effects of oncostatin M (OSM, a member of the IL-6 family of cytokines, on pattern electroretinogram (PERG and RGC survival after optic nerve crush (ON-crush in the mouse. BALB/C mice received ON-crush in the left eyes for either 4-second or 1-second duration (4-s or 1-s. Fluoro-gold retrograde labeling was used to identify RGCs. RGC function was assessed by PERG measurement. OSM or CNTF protein was injected intravitreally immediately after ON-crush. OSM responsive cells were identified by localization of increased STAT3 phosphorylation. Significant higher RGC survival (46% of untreated control was seen in OSM-treated eyes when assessed 2 weeks after 4-s ON-crush as compared to that (14% of untreated control of the PBS-treated eyes (P<0.001. In addition, PERG amplitude was significantly higher in eyes treated with OSM or CNTF 1 week after 1-s ON-crush (36% of baseline as compared with the amplitude of PBS-treated eyes (19% of the baseline, P = 0.003. An increase in STAT3 phosphorylation was localized in Müller layer after OSM treatment, suggesting that Müller cells mediate the effect of OSM. Our results demonstrate that one single injection of either OSM or CNTF after ON-crush improves RGC survival together with their electrophysiological activity. These data provide proof-of-concept for using neurotrophic factors OSM and CNTF for RGC degenerative diseases, including glaucoma and acute optic nerve trauma.

  10. Effect of Gonadectomy on the Androgen-Dependent Behavior of Ganglion Cell-Like Cells in Djungarian Hamsters (Phodopus sungorus).

    Science.gov (United States)

    Nakahira, Rei; Yoshida, Rumika; Michishita, Masaki; Ohkusu-Tsukada, Kozo; Takahashi, Kimimasa

    2016-02-01

    Ganglion cell-like (GL) cells reside in the dermis of the ventral skin of mature male Djungarian hamsters (Phodopus sugorus) and express androgen receptor (AR). To assess whether GL cells have androgen-dependent behavior, we evaluated the histologic changes of GL cells after gonadectomy. Five male and 5 female hamsters were gonadectomized at the age of 4 wk and necropsied 14 wk later. The number, distribution, and proliferative activity of GL cells in the thoracoabdominal and dorsal skins were evaluated histologically and compared with those of corresponding intact animals. GL cells were more numerous, were distributed throughout the skin more widely, and had higher proliferative activity in the intact male hamsters than in their gonadectomized counterparts. Similar trends regarding these 3 parameters were seen in ovariectomized compared with intact female hamsters and between intact male and intact female hamsters. These results suggest that the GL cells of Djungarian hamsters demonstrate sex-associated differences in their distribution and proliferative activity and that androgen may be involved in the development of these cells.

  11. Characterization of retinal ganglion cell, horizontal cell, and amacrine cell types expressing the neurotrophic receptor tyrosine kinase Ret.

    Science.gov (United States)

    Parmhans, Nadia; Sajgo, Szilard; Niu, Jingwen; Luo, Wenqin; Badea, Tudor Constantin

    2018-03-01

    We report the retinal expression pattern of Ret, a receptor tyrosine kinase for the glial derived neurotrophic factor (GDNF) family ligands (GFLs), during development and in the adult mouse. Ret is initially expressed in retinal ganglion cells (RGCs), followed by horizontal cells (HCs) and amacrine cells (ACs), beginning with the early stages of postmitotic development. Ret expression persists in all three classes of neurons in the adult. Using RNA sequencing, immunostaining and random sparse recombination, we show that Ret is expressed in at least three distinct types of ACs, and ten types of RGCs. Using intersectional genetics, we describe the dendritic arbor morphologies of RGC types expressing Ret in combination with each of the three members of the POU4f/Brn3 family of transcription factors. Ret expression overlaps with Brn3a in 4 RGC types, with Brn3b in 5 RGC types, and with Brn3c in one RGC type, respectively. Ret + RGCs project to the lateral geniculate nucleus (LGN), pretectal area (PTA) and superior colliculus (SC), and avoid the suprachiasmatic nucleus and accessory optic system. Brn3a + Ret + and Brn3c + Ret + RGCs project preferentially to contralateral retinorecipient areas, while Brn3b + Ret + RGCs shows minor ipsilateral projections to the olivary pretectal nucleus and the LGN. Our findings establish intersectional genetic approaches for the anatomic and developmental characterization of individual Ret + RGC types. In addition, they provide necessary information for addressing the potential interplay between GDNF neurotrophic signaling and transcriptional regulation in RGC type specification. © 2017 Wiley Periodicals, Inc.

  12. Comparative sensitivity of human and rat neural cultures to chemical-induced inhibition of neurite outgrowth

    International Nuclear Information System (INIS)

    Harrill, Joshua A.; Freudenrich, Theresa M.; Robinette, Brian L.; Mundy, William R.

    2011-01-01

    There is a need for rapid, efficient and cost-effective alternatives to traditional in vivo developmental neurotoxicity testing. In vitro cell culture models can recapitulate many of the key cellular processes of nervous system development, including neurite outgrowth, and may be used as screening tools to identify potential developmental neurotoxicants. The present study compared primary rat cortical cultures and human embryonic stem cell-derived neural cultures in terms of: 1) reproducibility of high content image analysis based neurite outgrowth measurements, 2) dynamic range of neurite outgrowth measurements and 3) sensitivity to chemicals which have been shown to inhibit neurite outgrowth. There was a large increase in neurite outgrowth between 2 and 24 h in both rat and human cultures. Image analysis data collected across multiple cultures demonstrated that neurite outgrowth measurements in rat cortical cultures were more reproducible and had higher dynamic range as compared to human neural cultures. Human neural cultures were more sensitive than rat cortical cultures to chemicals previously shown to inhibit neurite outgrowth. Parallel analysis of morphological (neurite count, neurite length) and cytotoxicity (neurons per field) measurements were used to detect selective effects on neurite outgrowth. All chemicals which inhibited neurite outgrowth in rat cortical cultures did so at concentrations which did not concurrently affect the number of neurons per field, indicating selective effects on neurite outgrowth. In contrast, more than half the chemicals which inhibited neurite outgrowth in human neural cultures did so at concentrations which concurrently decreased the number of neurons per field, indicating that effects on neurite outgrowth were secondary to cytotoxicity. Overall, these data demonstrate that the culture models performed differently in terms of reproducibility, dynamic range and sensitivity to neurite outgrowth inhibitors. While human neural

  13. [Effects on survival of shRNA mediated APE/Ref1 gene silencing in rat spiral ganglion cells in oxidative stress].

    Science.gov (United States)

    Jiang, Zhendong; Zhong, Cheng; Li, Taijun; Xiang, Zhaolan; Zhang, Xueyuan

    2014-02-01

    To investigate the effects of reducing APE/Ref1 expression in the cultures of rat spiral ganglion cells with oxidative damage induced by H(2)O(2). Primary cultured rat spiral ganglion cells were infected with small interfering RNA to APE/Ref1 (Ape1siRNA) for 72 h, followed by treating with H(2)O(2) (0, 10, 25, 50, 100 and 300 µmol/L) for 1 h , and then cultured in normal medium for 24 h. Western blot were used to detect the level of APE/Ref1 protein and phosphorylation of histone protein H2AX in the infected cells. The caspase3 activation was tested by spectrophotometric method . The cell viability was determined by MTT and the apoptosis of spiral ganglion cells was determined by terminal-deoxynucleotidyl transferase mediated nick and labeling (TUNEL). Western blot showed that infection with Ape1siRNA resulted in APE/Ref1 reduced expression in the spiral ganglion cells. Exposing spiral ganglion cultures with reduced expression of APE/Ref1 to H(2)O(2) (50, 100, 300 µmol/L) for 1 h resulted in increasing in the phosphorylation of histone protein H2AX. The reduction in APE/Ref1 significantly reduced cell viability in cultures 24 h after 1 h expression to 50-300 µmol/L H(2)O(2). The apoptosis of cells and caspase 3 activity was detected significantly improved. The induced of APE/Ref1 results in significantly decrease in spiral ganglion cells viability in oxidative stress. The repairing function of APE/Ref1 is necessary for optimal levels of neuronal rat spiral ganglion cells survival.

  14. ["Point by point" approach to structure-function correlation of glaucoma on the ganglion cell complex in the posterior pole].

    Science.gov (United States)

    Zeitoun, M

    2017-01-01

    To try to establish a "point by point" relationship between the local thickness of the retinal ganglion cell complex and its sensitivity. In total, 104 glaucomatous eyes of 89 patients with a confirmed 24-2 visual field, were measured by superimposing the visual field, using imaging software, with the Wide 40° by 30° measurements of retinal ganglion cell complex obtained from the Topcon © 3D 2000 OCT, after upward adjustment, inversion and scaling. Visual fields were classified into two groups according to the extent of the disease: 58 mild to moderate (MD up to -12dB), and 46 severe (MD beyond -12dB). The 6mm by 6mm central region, equipped with a normative database, was studied, corresponding to 16 points in the visual field. These points were individually matched one by one to the local ganglion cell complex, which was classified into 2 groups depending on whether it was greater or less than 70 microns. The normative database confirmed the pathological nature of the thin areas, with a significance of 95 to 99%. Displacement of central retinal ganglion cells was compensated for. Of 1664 points (16 central points for 104 eyes), 283 points were found to be "borderline" and excluded. Of the 1381 analyzed points, 727 points were classified as "over 70 microns" and 654 points "under 70 microns". (1) For all stages combined, 85.8% of the 727 points which were greater than 70 microns had a deviation between -3 and +3dB: areas above 70 microns had no observable loss of light sensitivity. (2) In total, 92.5% of the 428 points having a gap ranging from -6 to -35dB were located on ganglion cell complex areas below 70 microns: functional visual loss was identified in thin areas, which were less than 70 microns. (3) Areas which were less than 70 microns, that is 654 points, had quite variable sensitivity and can be divided into three groups: the first with preserved sensitivity, another with obliterated sensitivity, and an intermediate group connecting

  15. Random Wiring, Ganglion Cell Mosaics, and the Functional Architecture of the Visual Cortex.

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    Schottdorf, Manuel; Keil, Wolfgang; Coppola, David; White, Leonard E; Wolf, Fred

    2015-11-01

    The architecture of iso-orientation domains in the primary visual cortex (V1) of placental carnivores and primates apparently follows species invariant quantitative laws. Dynamical optimization models assuming that neurons coordinate their stimulus preferences throughout cortical circuits linking millions of cells specifically predict these invariants. This might indicate that V1's intrinsic connectome and its functional architecture adhere to a single optimization principle with high precision and robustness. To validate this hypothesis, it is critical to closely examine the quantitative predictions of alternative candidate theories. Random feedforward wiring within the retino-cortical pathway represents a conceptually appealing alternative to dynamical circuit optimization because random dimension-expanding projections are believed to generically exhibit computationally favorable properties for stimulus representations. Here, we ask whether the quantitative invariants of V1 architecture can be explained as a generic emergent property of random wiring. We generalize and examine the stochastic wiring model proposed by Ringach and coworkers, in which iso-orientation domains in the visual cortex arise through random feedforward connections between semi-regular mosaics of retinal ganglion cells (RGCs) and visual cortical neurons. We derive closed-form expressions for cortical receptive fields and domain layouts predicted by the model for perfectly hexagonal RGC mosaics. Including spatial disorder in the RGC positions considerably changes the domain layout properties as a function of disorder parameters such as position scatter and its correlations across the retina. However, independent of parameter choice, we find that the model predictions substantially deviate from the layout laws of iso-orientation domains observed experimentally. Considering random wiring with the currently most realistic model of RGC mosaic layouts, a pairwise interacting point process, the

  16. Random Wiring, Ganglion Cell Mosaics, and the Functional Architecture of the Visual Cortex.

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    Manuel Schottdorf

    2015-11-01

    Full Text Available The architecture of iso-orientation domains in the primary visual cortex (V1 of placental carnivores and primates apparently follows species invariant quantitative laws. Dynamical optimization models assuming that neurons coordinate their stimulus preferences throughout cortical circuits linking millions of cells specifically predict these invariants. This might indicate that V1's intrinsic connectome and its functional architecture adhere to a single optimization principle with high precision and robustness. To validate this hypothesis, it is critical to closely examine the quantitative predictions of alternative candidate theories. Random feedforward wiring within the retino-cortical pathway represents a conceptually appealing alternative to dynamical circuit optimization because random dimension-expanding projections are believed to generically exhibit computationally favorable properties for stimulus representations. Here, we ask whether the quantitative invariants of V1 architecture can be explained as a generic emergent property of random wiring. We generalize and examine the stochastic wiring model proposed by Ringach and coworkers, in which iso-orientation domains in the visual cortex arise through random feedforward connections between semi-regular mosaics of retinal ganglion cells (RGCs and visual cortical neurons. We derive closed-form expressions for cortical receptive fields and domain layouts predicted by the model for perfectly hexagonal RGC mosaics. Including spatial disorder in the RGC positions considerably changes the domain layout properties as a function of disorder parameters such as position scatter and its correlations across the retina. However, independent of parameter choice, we find that the model predictions substantially deviate from the layout laws of iso-orientation domains observed experimentally. Considering random wiring with the currently most realistic model of RGC mosaic layouts, a pairwise interacting point

  17. Retinal ganglion cell neuroprotection in a rat model of glaucoma following brimonidine, latanoprost or combined treatments.

    Science.gov (United States)

    Hernández, María; Urcola, J Haritz; Vecino, Elena

    2008-05-01

    The aim of the present study is to evaluate the neuroprotective effect of two antiglaucomatous substances, regardless of their hypotensive effect in the eye. Brimonidine, which does not reduce IOP when administered intraperitoneally, and latanoprost, which has a renowned hypotensive effect topically. We examined rat retinal ganglion cell (RGC) survival and size distribution in experimental glaucoma in response to different glaucomatous agents. IOP was elevated by episcleral vein cauterization (EVC) prior to the application of different treatments: (I) PBS application (control group), (II) intraperitoneal administration of brimonidine (a general hypotensive agent), (III) topical application of latanoprost (an ocular hypotensive agent), and (IV) latanoprost combined with brimonidine. After 12 weeks, RGCs were retrogradely labeled with fluorogold and RGC density was analyzed. EVC caused a significant increase (42%) in IOP in each group before drug treatment. After 12weeks of EVC, RGC survival in control vs. EVC rats was 78.9+/-3.2%. No IOP reduction was observed in brimonidine injected rats, but RGC survival at 12 weeks was total (103.7+/-2.7%). In latanoprost treated rats, IOP dropped by around 22% and 94.7+/-3.7% of the RGC population survived. Finally in the latanoprost+brimonidine combined group, IOP was significantly reduced by 25% and 94.4+/-2.2% of RGCs survived. Surprisingly, whereas EVC led to a 6% increase in RGC soma size, brimonidine treatment was associated with a 9% reduction in the soma size of RGCs at 12 weeks. We conclude that brimonidine exerts a neuroprotective effect via a mechanism which is independent of IOP reduction. These findings indicate that cell survival in glaucoma may be enhanced by neuroprotective strategies which are independent of IOP reduction. No synergistic neuroprotective effect was observed when both treatments were applied simultaneously.

  18. Losartan Treatment Protects Retinal Ganglion Cells and Alters Scleral Remodeling in Experimental Glaucoma.

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    Harry A Quigley

    Full Text Available To determine if oral losartan treatment decreases the retinal ganglion cell (RGC death caused by experimental intraocular pressure (IOP elevation in mice.We produced IOP increase in CD1 mice and performed unilateral optic nerve crush. Mice received oral losartan, spironolactone, enalapril, or no drug to test effects of inhibiting angiotensin receptors. IOP was monitored by Tonolab, and blood pressure was monitored by tail cuff device. RGC loss was measured in masked axon counts and RGC bodies by β-tubulin labeling. Scleral changes that could modulate RGC injury were measured including axial length, scleral thickness, and retinal layer thicknesses, pressure-strain behavior in inflation testing, and study of angiotensin receptors and pathways by reverse transcription polymerase chain reaction, Western blot, and immunohistochemistry.Losartan treatment prevented significant RGC loss (median loss = 2.5%, p = 0.13, while median loss with water, spironolactone, and enalapril treatments were 26%, 28% and 43%; p < 0.0001. The lower RGC loss with losartan was significantly less than the loss with spironolactone or enalapril (regression model p = 0.001; drug treatment group term p = 0.01. Both losartan and enalapril significantly lowered blood pressure (p< 0.001, but losartan was protective, while enalapril led to worse than water-treated RGC loss. RGC loss after crush injury was unaffected by losartan treatment (difference from control p = 0.9. Survival of RGC in cell culture was not prolonged by sartan treatment. Axonal transport blockade after 3 day IOP elevations was less in losartan-treated than in control glaucoma eyes (p = 0.007. Losartan inhibited effects of glaucoma, including reduction in extracellular signal-related kinase activity and modification of glaucoma-related changes in scleral thickness and creep under controlled IOP.The neuroprotective effect of losartan in mouse glaucoma is associated with adaptive changes in the sclera expressed at

  19. The role of intrinsically photosensitive retinal ganglion cells in nonimage-forming responses to light

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    Warthen DM

    2012-09-01

    Full Text Available Daniel M Warthen,1,2 Ignacio Provencio11Department of Biology, University of Virginia, Charlottesville, VA, USA; 2Department of Pharmacology, University of Virginia School of Medicine, Charlottesville, VA, USAAbstract: Light exerts many effects on behavior and physiology. These effects can be characterized as either image-forming or nonimage-forming (NIF visual processes. Image-forming vision refers to the process of detecting objects and organisms in the environment and distinguishing their physical characteristics, such as size, shape, and direction of motion. NIF vision, in contrast, refers to effects of light that are independent of fine spatiotemporal vision. NIF effects are many and varied, ranging from modulation of basal physiology, such as heart rate and body temperature, to changes in higher functions, such as mood and cognitive performance. In mammals, many NIF effects of light are dependent upon the inner retinal photopigment melanopsin and the cells in which melanopsin is expressed, the intrinsically photosensitive retinal ganglion cells (ipRGCs. The ipRGCs project broadly throughout the brain. Many of these projections terminate in areas known to mediate NIF effects, while others terminate in regions whose link to photoreception remains to be established. Additionally, the presence of ipRGC projections to areas of the brain with no known link to photoreception suggests the existence of additional ipRGC-mediated NIF effects. This review summarizes the known NIF effects of light and the role of melanopsin and ipRGCs in driving these effects, with an eye toward stimulating further investigation of the many and varied effects of light on physiology and behavior.Keywords: amygdala, bed nucleus of the stria terminalis, melanopsin, opsin, optic nerve, retina

  20. Molecular mechanisms of retinal ganglion cell degeneration in glaucoma and future prospects for cell body and axonal protection

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    Yasunari eMunemasa

    2013-01-01

    Full Text Available Glaucoma, which affects more than 70 million people worldwide, is a heterogeneous group of disorders with a resultant common denominator; optic neuropathy, eventually leading to irreversible blindness. The clinical manifestations of primary open-angle glaucoma (POAG, the most common subtype of glaucoma, include excavation of the optic disc and progressive loss of visual field. Axonal degeneration of retinal ganglion cells (RGCs and apoptotic death of their cell bodies are observed in glaucoma, in which the reduction of intraocular pressure is known to slow progression of the disease. A pattern of localized retinal nerve fiber layer defects in glaucoma patients indicates that axonal degeneration may precede RGC body death in this condition. The mechanisms of degeneration of neuronal cell bodies and their axons may differ. In this review, we addressed the molecular mechanisms of cell body death and axonal degeneration in glaucoma and proposed axonal protection in addition to cell body protection. The concept of axonal protection may become a new therapeutic strategy to prevent further axonal degeneration or revive dying axons in patients with preperimetric glaucoma. Further study will be needed to clarify whether the combination therapy of axonal protection and cell body protection will have greater protective effects in early or progressive glaucomatous optic neuropathy.

  1. Retinal Astrocytes and GABAergic Wide-Field Amacrine Cells Express PDGFRα: Connection to Retinal Ganglion Cell Neuroprotection by PDGF-AA.

    Science.gov (United States)

    Takahama, Shokichi; Adetunji, Modupe O; Zhao, Tantai; Chen, Shan; Li, Wei; Tomarev, Stanislav I

    2017-09-01

    Our previous experiments demonstrated that intravitreal injection of platelet-derived growth factor-AA (PDGF-AA) provides retinal ganglion cell (RGC) neuroprotection in a rodent model of glaucoma. Here we used PDGFRα-enhanced green fluorescent protein (EGFP) mice to identify retinal cells that may be essential for RGC protection by PDGF-AA. PDGFRα-EGFP mice expressing nuclear-targeted EGFP under the control of the PDGFRα promoter were used. Localization of PDGFRα in the neural retina was investigated by confocal imaging of EGFP fluorescence and immunofluorescent labeling with a panel of antibodies recognizing different retinal cell types. Primary cultures of mouse RGCs were produced by immunopanning. Neurobiotin injection of amacrine cells in a flat-mounted retina was used for the identification of EGFP-positive amacrine cells in the inner nuclear layer. In the mouse neural retina, PDGFRα was preferentially localized in the ganglion cell and inner nuclear layers. Immunostaining of the retina demonstrated that astrocytes in the ganglion cell layer and a subpopulation of amacrine cells in the inner nuclear layer express PDGFRα, whereas RGCs (in vivo or in vitro) did not. PDGFRα-positive amacrine cells are likely to be Type 45 gamma-aminobutyric acidergic (GABAergic) wide-field amacrine cells. These data indicate that the neuroprotective effect of PDGF-AA in a rodent model of glaucoma could be mediated by astrocytes and/or a subpopulation of amacrine cells. We suggest that after intravitreal injection of PDGF-AA, these cells secrete factors protecting RGCs.

  2. Vasoactive intestinal peptide-induced neurite remodeling in human neuroblastoma SH-SY5Y cells implicates the Cdc42 GTPase and is independent of Ras-ERK pathway

    International Nuclear Information System (INIS)

    Alleaume, Celine; Eychene, Alain; Harnois, Thomas; Bourmeyster, Nicolas; Constantin, Bruno; Caigneaux, Evelyne; Muller, Jean-Marc; Philippe, Michel

    2004-01-01

    Vasoactive intestinal peptide (VIP) is known to regulate proliferation or differentiation in normal and tumoral cells. SH-SY5Y is a differentiated cell subclone derived from the SK-N-SH human neuroblastoma cell line and possess all the components for an autocrine action of VIP. In the present study, we investigated the morphological changes and intracellular signaling pathways occurring upon VIP treatment of SH-SY5Y cells. VIP induced an early remodeling of cell projections: a branched neurite network spread out and prominent varicosities developed along neurites. Although activated by VIP, the Ras/ERK pathway was not required for the remodeling process. In contrast, pull-down experiments revealed a strong Cdc42 activation by VIP while expression of a dominant-negative Cdc42 prevented the VIP-induced neurite changes, suggesting an important role for this small GTPase in the process. These data provide the first evidence for a regulation of the activity of Rho family GTPases by VIP and bring new insights in the signaling pathways implicated in neurite remodeling process induced by VIP in neuroblastoma cells

  3. Mouse ganglion-cell photoreceptors are driven by the most sensitive rod pathway and by both types of cones.

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    Shijun Weng

    Full Text Available Intrinsically photosensitive retinal ganglion cells (iprgcs are depolarized by light by two mechanisms: directly, through activation of their photopigment melanopsin; and indirectly through synaptic circuits driven by rods and cones. To learn more about the rod and cone circuits driving ipRGCs, we made multielectrode array (MEA and patch-clamp recordings in wildtype and genetically modified mice. Rod-driven ON inputs to ipRGCs proved to be as sensitive as any reaching the conventional ganglion cells. These signals presumably pass in part through the primary rod pathway, involving rod bipolar cells and AII amacrine cells coupled to ON cone bipolar cells through gap junctions. Consistent with this interpretation, the sensitive rod ON input to ipRGCs was eliminated by pharmacological or genetic disruption of gap junctions, as previously reported for conventional ganglion cells. A presumptive cone input was also detectable as a brisk, synaptically mediated ON response that persisted after disruption of rod ON pathways. This was roughly three log units less sensitive than the rod input. Spectral analysis revealed that both types of cones, the M- and S-cones, contribute to this response and that both cone types drive ON responses. This contrasts with the blue-OFF, yellow-ON chromatic opponency reported in primate ipRGCs. The cone-mediated response was surprisingly persistent during steady illumination, echoing the tonic nature of both the rod input to ipRGCs and their intrinsic, melanopsin-based phototransduction. These synaptic inputs greatly expand the dynamic range and spectral bandpass of the non-image-forming visual functions for which ipRGCs provide the principal retinal input.

  4. Light-induced fos expression in intrinsically photosensitive retinal ganglion cells in melanopsin knockout (opn4 mice.

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    Gary E Pickard

    Full Text Available Retinal ganglion cells that express the photopigment melanopsin are intrinsically photosensitive (ipRGCs and exhibit robust synaptically driven ON-responses to light, yet they will continue to depolarize in response to light when all synaptic input from rod and cone photoreceptors is removed. The light-evoked increase in firing of classical ganglion cells is determined by synaptic input from ON-bipolar cells in the proximal sublamina of the inner plexiform layer. OFF-bipolar cells synapse with ganglion cell dendrites in the distal sublamina of the inner plexiform layer. Of the several types of ipRGC that have been described, M1 ipRGCs send dendrites exclusively into the OFF region of the inner plexiform layer where they stratify near the border of the inner nuclear layer. We tested whether M1 ipRGCs with dendrites restricted to the OFF sublamina of the inner plexiform layer receive synaptic ON-bipolar input by examining light-induced gene expression in vivo using melanopsin knockout mice. Mice in which both copies of the melanopsin gene (opn4 have been replaced with the tau-lacZ gene (homozygous tau-lacZ(+/+ knockin mice are melanopsin knockouts (opn4(-/- but M1 ipRGCs are specifically identified by their expression of beta-galactosidase. Approximately 60% of M1 ipRGCs in Opn4(-/- mice exposed to 3 hrs of light expressed c-Fos; no beta-galactosidase-positive RGCs expressed c-Fos in the dark. Intraocular application of L-AP4, a compound which blocks transmission of visual signals between photoreceptors and ON-bipolar cells significantly reduced light-evoked c-Fos expression in M1 ipRGCs compared to saline injected eyes (66% saline vs 27% L-AP4. The results are the first description of a light-evoked response in an ipRGC lacking melanopsin and provide in vivo confirmation of previous in vitro observations illustrating an unusual circuit in the retina in which ganglion cells sending dendrites to the OFF sublamina of the inner plexiform layer

  5. A dedicated circuit links direction-selective retinal ganglion cells to the primary visual cortex

    Science.gov (United States)

    Cruz-Martín, Alberto; El-Danaf, Rana N.; Osakada, Fumitaka; Sriram, Balaji; Dhande, Onkar S.; Nguyen, Phong L.; Callaway, Edward M.; Ghosh, Anirvan; Huberman, Andrew D.

    2014-03-01

    How specific features in the environment are represented within the brain is an important unanswered question in neuroscience. A subset of retinal neurons, called direction-selective ganglion cells (DSGCs), are specialized for detecting motion along specific axes of the visual field. Despite extensive study of the retinal circuitry that endows DSGCs with their unique tuning properties, their downstream circuitry in the brain and thus their contribution to visual processing has remained unclear. In mice, several different types of DSGCs connect to the dorsal lateral geniculate nucleus (dLGN), the visual thalamic structure that harbours cortical relay neurons. Whether direction-selective information computed at the level of the retina is routed to cortical circuits and integrated with other visual channels, however, is unknown. Here we show that there is a di-synaptic circuit linking DSGCs with the superficial layers of the primary visual cortex (V1) by using viral trans-synaptic circuit mapping and functional imaging of visually driven calcium signals in thalamocortical axons. This circuit pools information from several types of DSGCs, converges in a specialized subdivision of the dLGN, and delivers direction-tuned and orientation-tuned signals to superficial V1. Notably, this circuit is anatomically segregated from the retino-geniculo-cortical pathway carrying non-direction-tuned visual information to deeper layers of V1, such as layer 4. Thus, the mouse harbours several functionally specialized, parallel retino-geniculo-cortical pathways, one of which originates with retinal DSGCs and delivers direction- and orientation-tuned information specifically to the superficial layers of the primary visual cortex. These data provide evidence that direction and orientation selectivity of some V1 neurons may be influenced by the activation of DSGCs.

  6. Retinal ganglion cell complex changes using spectral domain optical coherence tomography in diabetic patients without retinopathy

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    Ahmed I. Hegazy

    2017-03-01

    Full Text Available AIM: To assess the ganglion cell complex (GCC thickness in diabetic eyes without retinopathy. METHODS: Two groups included 45 diabetic eyes without retinopathy and 21 non diabetic eyes. All subjects underwent full medical and ophthalmological history, full ophthalmological examination, measuring GCC thickness and central foveal thickness (CFT using the RTVue® spectral domain-optical coherence tomography (SD-OCT, and HbA1C level. RESULTS: GCC focal loss volume (FLV% was significantly more in diabetic eyes (22.2% below normal than normal eyes (P=0.024. No statistically significant difference was found between the diabetic group and the control group regarding GCC global loss volume (GLV% (P=0.160. CFT was positively correlated to the average, superior and inferior GCC (P=0.001, 0.000 and 0.001 respectively and negatively correlated to GLV% and FLV% (P=0.002 and 0.031 respectively in diabetic eyes. C/D ratio in diabetic eyes was negatively correlated to average, superior and inferior GCC (P=0.015, 0.007 and 0.017 respectively. The FLV% was negatively correlated to the refraction and level of HbA1c (P=0.019 and 0.013 respectively and positively correlated to the best corrected visual acuity (BCVA in logMAR in diabetic group (P=0.004. CONCLUSION: Significant GCC thinning in diabetes predates retinal vasculopathy, which is mainly focal rather than diffuse. It has no preference to either the superior or inferior halves of the macula. Increase of myopic error is significantly accompanied with increased focal GCC loss. GCC loss is accompanied with increased C/D ratio in diabetic eyes.

  7. Mice deficient of glutamatergic signaling from intrinsically photosensitive retinal ganglion cells exhibit abnormal circadian photoentrainment.

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    Nicole Purrier

    Full Text Available Several aspects of behavior and physiology, such as sleep and wakefulness, blood pressure, body temperature, and hormone secretion exhibit daily oscillations known as circadian rhythms. These circadian rhythms are orchestrated by an intrinsic biological clock in the suprachiasmatic nuclei (SCN of the hypothalamus which is adjusted to the daily environmental cycles of day and night by the process of photoentrainment. In mammals, the neuronal signal for photoentrainment arises from a small subset of intrinsically photosensitive retinal ganglion cells (ipRGCs that send a direct projection to the SCN. ipRGCs also mediate other non-image-forming (NIF visual responses such as negative masking of locomotor activity by light, and the pupillary light reflex (PLR via co-release of neurotransmitters glutamate and pituitary adenylate cyclase-activating peptide (PACAP from their synaptic terminals. The relative contribution of each neurotransmitter system for the circadian photoentrainment and other NIF visual responses is still unresolved. We investigated the role of glutamatergic neurotransmission for circadian photoentrainment and NIF behaviors by selective ablation of ipRGC glutamatergic synaptic transmission in mice. Mutant mice displayed delayed re-entrainment to a 6 h phase shift (advance or delay in the light cycle and incomplete photoentrainment in a symmetrical skeleton photoperiod regimen (1 h light pulses between 11 h dark periods. Circadian rhythmicity in constant darkness also was reduced in some mutant mice. Other NIF responses such as the PLR and negative masking responses to light were also partially attenuated. Overall, these results suggest that glutamate from ipRGCs drives circadian photoentrainment and negative masking responses to light.

  8. Effects of p-xylene inhalation on axonal transport in the rat retinal ganglion cells

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    Padilla, S.S.; Lyerly, D.P. (Environmental Protection Agency, Research Triangle Park, NC (USA))

    1989-12-01

    Although the solvent xylene is suspected of producing nervous system dysfunction in animals and humans, little is known regarding the neurochemical consequences of xylene inhalation. The intent of this study was to determine the effect of intermittent, acute, and subchronic p-xylene exposure on the axonal transport of proteins and glycoproteins within the rat retinofugal tract. A number of different exposure regimens were tested ranging from 50 ppm for a single 6-hr exposure to 1600 ppm 6 hr/day, 5 days/week, for a total of 8 exposure days. Immediately following removal from the inhalation chambers rats were injected intraocularly with (35S)methionine and (3H)fucose (to label retinal proteins and glycoproteins, respectively) and the axonal transport of labeled macromolecules to axons (optic nerve and optic tract) and nerve endings (lateral geniculate body and superior colliculus) was examined 20 hr after precursor injection. Only relatively severe exposure regimens (i.e., 800 or 1600 ppm 6 hr/day, 5 days/week, for 1.5 weeks) produced significant reductions in axonal transport; there was a moderate reduction in the axonal transport of 35S-labeled proteins in the 800-ppm-treated group which was more widespread in the 1600 ppm-treated group. Transport of 3H-labeled glycoproteins was less affected. Assessment of retinal metabolism immediately after isotope injection indicated that the rate of precursor uptake was not reduced in either treatment group. Furthermore, rapid transport was still substantially reduced in animals exposed to 1600 ppm p-xylene and allowed a 13-day withdrawal period. These data indicate that p-xylene inhalation decreases rapid axonal transport supplied to the projections of the rat retinal ganglion cells immediately after cessation of inhalation exposure and that this decreased transport is still apparent 13 days after the last exposure.

  9. Effects of p-xylene inhalation on axonal transport in the rat retinal ganglion cells

    International Nuclear Information System (INIS)

    Padilla, S.S.; Lyerly, D.P.

    1989-01-01

    Although the solvent xylene is suspected of producing nervous system dysfunction in animals and humans, little is known regarding the neurochemical consequences of xylene inhalation. The intent of this study was to determine the effect of intermittent, acute, and subchronic p-xylene exposure on the axonal transport of proteins and glycoproteins within the rat retinofugal tract. A number of different exposure regimens were tested ranging from 50 ppm for a single 6-hr exposure to 1600 ppm 6 hr/day, 5 days/week, for a total of 8 exposure days. Immediately following removal from the inhalation chambers rats were injected intraocularly with [35S]methionine and [3H]fucose (to label retinal proteins and glycoproteins, respectively) and the axonal transport of labeled macromolecules to axons (optic nerve and optic tract) and nerve endings (lateral geniculate body and superior colliculus) was examined 20 hr after precursor injection. Only relatively severe exposure regimens (i.e., 800 or 1600 ppm 6 hr/day, 5 days/week, for 1.5 weeks) produced significant reductions in axonal transport; there was a moderate reduction in the axonal transport of 35S-labeled proteins in the 800-ppm-treated group which was more widespread in the 1600 ppm-treated group. Transport of 3H-labeled glycoproteins was less affected. Assessment of retinal metabolism immediately after isotope injection indicated that the rate of precursor uptake was not reduced in either treatment group. Furthermore, rapid transport was still substantially reduced in animals exposed to 1600 ppm p-xylene and allowed a 13-day withdrawal period. These data indicate that p-xylene inhalation decreases rapid axonal transport supplied to the projections of the rat retinal ganglion cells immediately after cessation of inhalation exposure and that this decreased transport is still apparent 13 days after the last exposure

  10. Characterization of spontaneous excitatory synaptic currents in salamander retinal ganglion cells.

    Science.gov (United States)

    Taylor, W R; Chen, E; Copenhagen, D R

    1995-01-01

    1. Spontaneous excitatory postsynaptic currents (sEPSCs) were recorded under voltage-clamp conditions. Consistent with activation of non-NMDA-type glutamate receptors, the sEPSCs reversed at potentials above 0 mV, were blocked by 1 microM CNQX and prolonged by 2 mM aniracetam. 2. The peak conductance of the averaged sEPSCs (n = 70-400) was 130 +/- 60 pS (mean +/- S.D.; 17 cells, ranging from 70 to 290 pS). Amplitude distributions were skewed towards larger amplitudes. 3. The decay of individual and mean sEPSCs was exponential with a mean time constant (tau d) of 3.75 +/- 0.84 ms (n = 13), which was voltage independent. The 10-90% rise time of the sEPSCs was 1.30 +/- 0.44 ms (n = 13). There was no correlation between sEPSC rise time and tau d suggesting that dendritic filtering alone did not shape the time course of sEPSCs. 4. Light-evoked EPSCs in these retinal ganglion cells are mediated by concomitant activation of NMDA and non-NMDA receptors; however, no NMDA component was discerned in the sEPSCs, even when recording at -96 mV in Mg(2+)-free solutions. The decay time course was not altered by 20 microM AP7, an NMDA antagonist, nor was an NMDA component unmasked by adding glycine or D-serine. These results suggest that NMDA and non-NMDA receptors are not coactivated by a single vesicle of transmitter during spontaneous release, and thus are probably not colocalized in the postsynaptic membrane at the sites of spontaneous release. 5. The sEPSCs were an order of magnitude faster than the non-NMDA receptor-mediated EPSCs evoked by light stimuli, and it is proposed that the EPSC time course is determined largely by the extended time course of release of synaptic vesicles from bipolar cells. The quantal content of a light-evoked non-NMDA receptor-mediated EPSC in an on-off cell is about 200 quanta. Images Figure 6 PMID:7562636

  11. Hydrostatic pressure does not cause detectable changes in survival of human retinal ganglion cells.

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    Andrew Osborne

    Full Text Available PURPOSE: Elevated intraocular pressure (IOP is a major risk factor for glaucoma. One consequence of raised IOP is that ocular tissues are subjected to increased hydrostatic pressure (HP. The effect of raised HP on stress pathway signaling and retinal ganglion cell (RGC survival in the human retina was investigated. METHODS: A chamber was designed to expose cells to increased HP (constant and fluctuating. Accurate pressure control (10-100 mmHg was achieved using mass flow controllers. Human organotypic retinal cultures (HORCs from donor eyes (<24 h post mortem were cultured in serum-free DMEM/HamF12. Increased HP was compared to simulated ischemia (oxygen glucose deprivation, OGD. Cell death and apoptosis were measured by LDH and TUNEL assays, RGC marker expression by qRT-PCR (THY-1 and RGC number by immunohistochemistry (NeuN. Activated p38 and JNK were detected by Western blot. RESULTS: Exposure of HORCs to constant (60 mmHg or fluctuating (10-100 mmHg; 1 cycle/min pressure for 24 or 48 h caused no loss of structural integrity, LDH release, decrease in RGC marker expression (THY-1 or loss of RGCs compared with controls. In addition, there was no increase in TUNEL-positive NeuN-labelled cells at either time-point indicating no increase in apoptosis of RGCs. OGD increased apoptosis, reduced RGC marker expression and RGC number and caused elevated LDH release at 24 h. p38 and JNK phosphorylation remained unchanged in HORCs exposed to fluctuating pressure (10-100 mmHg; 1 cycle/min for 15, 30, 60 and 90 min durations, whereas OGD (3 h increased activation of p38 and JNK, remaining elevated for 90 min post-OGD. CONCLUSIONS: Directly applied HP had no detectable impact on RGC survival and stress-signalling in HORCs. Simulated ischemia, however, activated stress pathways and caused RGC death. These results show that direct HP does not cause degeneration of RGCs in the ex vivo human retina.

  12. Valproic acid prevents NMDA-induced retinal ganglion cell death via stimulation of neuronal TrkB receptor signaling.

    Science.gov (United States)

    Kimura, Atsuko; Namekata, Kazuhiko; Guo, Xiaoli; Noro, Takahiko; Harada, Chikako; Harada, Takayuki

    2015-03-01

    Valproic acid (VPA) is widely prescribed for treatment of epilepsy, mood disorders, migraines, and neuropathic pain. It exerts its therapeutic benefits through multiple mechanisms, including enhancement of GABAergic activity, activation of prosurvival protein kinases, and inhibition of histone deacetylase. Increasing evidence suggests that VPA possesses neuroprotective properties. We examined neuroprotective effects of VPA in an N-methyl-d-aspartate (NMDA) excitotoxicity model, which mimics some of the pathological features of glaucoma. In vivo retinal imaging using optical coherence tomography revealed that NMDA-induced retinal degeneration was suppressed in the VPA-treated retina, and histological analyses confirmed that VPA reduced retinal ganglion cell death. In vivo electrophysiological analyses demonstrated that visual impairment was prevented in the VPA-treated retina, clearly establishing both histological and functional effects of VPA. Brain-derived neurotrophic factor (BDNF) expression was up-regulated in Müller glial cells, and neuroprotective effects of VPA on retinal ganglion cells were significantly reduced in a conditional knockout mouse strain with deletion of tropomyosin receptor kinase B (TrkB), a receptor for BDNF from retinal ganglion cells. The results show that VPA stimulates BDNF up-regulation in Müller glial cells and provides direct evidence that neuronal TrkB is important in VPA-mediated neuroprotection. Also, VPA suppresses oxidative stress induced by NMDA in the retina. Our findings raise intriguing possibilities that the widely prescribed drug VPA may be useful for treatment of glaucoma. Copyright © 2015 American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.

  13. Dre - Cre sequential recombination provides new tools for retinal ganglion cell labeling and manipulation in mice.

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    Szilard Sajgo

    Full Text Available BACKGROUND: Genetic targeting methods have greatly advanced our understanding of many of the 20 Retinal Ganglion Cell (RGC types conveying visual information from the eyes to the brain. However, the complexity and partial overlap of gene expression patterns in RGCs call for genetic intersectional or sparse labeling strategies. Loci carrying the Cre recombinase in conjunction with conditional knock-out, reporter or other genetic tools can be used for targeted cell type ablation and functional manipulation of specific cell populations. The three members of the Pou4f family of transcription factors, Brn3a, Brn3b and Brn3c, expressed early during RGC development and in combinatorial pattern amongst RGC types are excellent candidates for such gene manipulations. METHODS AND FINDINGS: We generated conditional Cre knock-in alleles at the Brn3a and Brn3b loci, Brn3a(CKOCre and Brn3b(CKOCre. When crossed to mice expressing the Dre recombinase, the endogenous Brn3 gene expressed by Brn3a(CKOCre or Brn3b(CKOCre is removed and replaced with a Cre recombinase, generating Brn3a(Cre and Brn3b(Cre knock-in alleles. Surprisingly both Brn3a(Cre and Brn3b(Cre knock-in alleles induce early ubiquitous recombination, consistent with germline expression. However in later stages of development, their expression is limited to the expected endogenous pattern of the Brn3a and Brn3b genes. We use the Brn3a(Cre and Brn3b(Cre alleles to target a Cre dependent Adeno Associated Virus (AAV reporter to RGCs and demonstrate its use in morphological characterization, early postnatal gene delivery and tracing the expression of Brn3 genes in RGCs. CONCLUSIONS: Dre recombinase effectively recombines the Brn3a(CKOCre and Brn3b(CKOCre alleles containing its roxP target sites. Sequential Dre to Cre recombination reveals Brn3a and Brn3b expression in early mouse development. The generated Brn3a(Cre and Brn3b(Cre alleles are useful tools that can target exogenously delivered Cre dependent

  14. Endothelin-2/Vasoactive Intestinal Contractor: Regulation of Expression via Reactive Oxygen Species Induced by CoCl22, and Biological Activities Including Neurite Outgrowth in PC12 Cells

    Directory of Open Access Journals (Sweden)

    Eiichi Kotake-Nara

    2006-01-01

    Full Text Available This paper reviews the local hormone endothelin-2 (ET-2, or vasoactive intestinal contractor (VIC, a member of the vasoconstrictor ET peptide family, where ET-2 is the human orthologous peptide of the murine VIC. While ET-2/VIC gene expression has been observed in some normal tissues, ET-2 recently has been reported to act as a tumor marker and as a hypoxia-induced autocrine survival factor in tumor cells. A recently published study reported that the hypoxic mimetic agent CoCl2 at 200 µM increased expression of the ET-2/VIC gene, decreased expression of the ET-1 gene, and induced intracellular reactive oxygen species (ROS increase and neurite outgrowth in neuronal model PC12 cells. The ROS was generated by addition of CoCl2 to the culture medium, and the CoCl2-induced effects were completely inhibited by the antioxidant N-acetyl cysteine. Furthermore, interleukin-6 (IL-6 gene expression was up-regulated upon the differentiation induced by CoCl2. These results suggest that expression of ET-2/VIC and ET-1 mediated by CoCl2-induced ROS may be associated with neuronal differentiation through the regulation of IL-6 expression. CoCl2 acts as a pro-oxidant, as do Fe(II, III and Cu(II. However, some biological activities have been reported for CoCl2 that have not been observed for other metal salts such as FeCl3, CuSO4, and NiCl2. The characteristic actions of CoCl2 may be associated with the differentiation of PC12 cells. Further elucidation of the mechanism of neurite outgrowth and regulation of ET-2/VIC expression by CoCl2 may lead to the development of treatments for neuronal disorders.

  15. Baclofen Protects Primary Rat Retinal Ganglion Cells from Chemical Hypoxia-Induced Apoptosis through the Akt and PERK Pathways

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    Pingping Fu

    2016-11-01

    Full Text Available Retinal ganglion cells (RGCs consume large quantities of energy to convert light information into a neuronal signal, which makes them highly susceptible to hypoxic injury. This study aimed to investigate the potential protection by baclofen, a GABAB receptor agonist, of retinal ganglion cells against hypoxia-induced apoptosis. CoCl2 was applied to mimic hypoxia. Primary rat retinal ganglion cells (RGCs were subjected to CoCl2 with or without baclofen treatment, and RNA interference techniques were used to knock down the GABAB2 gene in the primary RGCs. The viability and apoptosis of RGCs were assessed using cell viability and TUNEL assays, Hoechst staining, and flow cytometry. The expression of cleaved caspase-3, bcl-2, bax, Akt, phospho-Akt, PERK, phospho-PERK, eIF2α, phospho-eIF2α, ATF-4, and CHOP were measured using western blotting. GABAB2 mRNA expression was determined using quantitative real-time polymerase chain reaction (qRT-PCR analysis. Our study revealed that CoCl2 significantly induced RGC apoptosis and that baclofen reversed these effects. CoCl2-induced reduction of Akt activity was also reversed by baclofen. Baclofen prevented the activation of the PERK pathway and the increase in CHOP expression induced by CoCl2. Knockdown of GABAB2 and the inactivation of the Akt pathway by inhibitors reduced the protective effect of baclofen on CoCl2-treated RGCs. Taken together, these results demonstrate that baclofen protects RGCs from CoCl2-induced apoptosis by increasing Akt activity and by suppressing the PERK pathway and CHOP activation.

  16. Developmental changes in NMDA receptor subunit composition at ON and OFF bipolar cell synapses onto direction-selective retinal ganglion cells.

    Science.gov (United States)

    Stafford, Benjamin K; Park, Silvia J H; Wong, Kwoon Y; Demb, Jonathan B

    2014-01-29

    In the developing mouse retina, spontaneous and light-driven activity shapes bipolar→ganglion cell glutamatergic synapse formation, beginning around the time of eye-opening (P12-P14) and extending through the first postnatal month. During this time, glutamate release can spill outside the synaptic cleft and possibly stimulate extrasynaptic NMDA-type glutamate receptors (NMDARs) on ganglion cells. Furthermore, the role of NMDARs during development may differ between ON and OFF bipolar synapses as in mature retina, where ON synapses reportedly include extrasynaptic NMDARs with GluN2B subunits. To better understand the function of glutamatergic synapses during development, we made whole-cell recordings of NMDAR-mediated responses, in vitro, from two types of genetically identified direction-selective ganglion cells (dsGCs): TRHR (thyrotropin-releasing hormone receptor) and Drd4 (dopamine receptor 4). Both dsGC types responded to puffed NMDA between P7 and P28; and both types exhibited robust light-evoked NMDAR-mediated responses at P14 and P28 that were quantified by conductance analysis during nicotinic and GABA(A) receptor blockade. For a given cell type and at a given age, ON and OFF bipolar cell inputs evoked similar NMDAR-mediated responses, suggesting that ON-versus-OFF differences in mature retina do not apply to the cell types or ages studied here. At P14, puff- and light-evoked NMDAR-mediated responses in both dsGCs were partially blocked by the GluN2B antagonist ifenprodil, whereas at P28 only TRHR cells remained ifenprodil-sensitive. NMDARs contribute at both ON and OFF bipolar cell synapses during a period of robust activity-dependent synaptic development, with declining GluN2B involvement over time in specific ganglion cell types.

  17. Role of retinal glial cell glutamate transporters in retinal ganglion cell survival following stimulation of NMDA receptor.

    Science.gov (United States)

    Furuya, Toshie; Pan, Zhiying; Kashiwagi, Kenji

    2012-03-01

    To investigate the role of glutamate transporters (GLTs)in retinal glial cells that were treated with N-methyl-D-aspartate (NMDA), in retinal ganglion cell (RGC) survival. Primary cultures of retinal glial cells or RGCs from 3-day-old Sprague-Dawley rats were employed in the present study. Retinal glial cells were treated with NMDA and changes in GLT mRNA and protein expression were analyzed. The effects of pretreating retinal glial cells with the GLAST-specific inhibitor, rottlerin (ROT), and the GLT-1-specific inhibitor, dihydrokainic acid (DHK), on RGC survival were investigated under exposure to NMDA. The amount of glutamate in the culture medium of retinal glial cells was measured by high-performance liquid chromatography. NMDA treatment increased GLAST and GLT-1 expression. GLAST and GLT-1 mRNA expression increased by 2.94-fold and 3.36-fold at 12 h after treatment with the highest concentration of NMDA (33 mM), and by 1.41-fold and 1.39-fold at 24 h, respectively. GLT-1 and GLAST protein expression also increased. MK801, an NMDA-receptor antagonist, inhibited the NMDA-induced upregulation of GLT mRNA expression. Co-culture with retinal glial cells increased the survival rate of RGCs. ROT decreased the survival rate of RGCs, whereas DHK significantly increased the survival rate of RGCs treated with 33 mM NMDA. NMDA treatment reduced the total amount of glutamate in the culture medium, particularly when 33 mM NMDA was added to the medium. ROT pretreatment increased the amount of glutamate in the culture medium, whereas DHK pretreatment decreased it. GLAST and GLT-1 may have different roles in the survival of RGCs mediated by retinal glial cells. These results suggest that the NMDA-associated induction of GLTs plays an important role in RGC survival.

  18. Orexin-A potentiates L-type calcium/barium currents in rat retinal ganglion cells.

    Science.gov (United States)

    Liu, F; Weng, S-J; Yang, X-L; Zhong, Y-M

    2015-10-01

    Two neuropeptides, orexin-A and orexin-B (also called hypocretin-1 and -2), have been implicated in sleep/wake regulation, feeding behaviors via the activation of two subtypes of G-protein-coupled receptors: orexin 1 and orexin 2 receptors (OX1R and OX2R). While the expression of orexins and orexin receptors is immunohistochemically revealed in retinal neurons, the function of these peptides in the retina is largely unknown. Using whole-cell patch-clamp recordings in rat retinal slices, we demonstrated that orexin-A increased L-type-like barium currents (IBa,L) in ganglion cells (GCs), and the effect was blocked by the selective OX1R antagonist SB334867, but not by the OX2R antagonist TCS OX2 29. The orexin-A effect was abolished by intracellular dialysis of GDP-β-S/GPAnt-2A, a Gq protein inhibitor, suggesting the mediation of Gq. Additionally, during internal dialysis of the phosphatidylinositol (PI)-phospholipase C (PLC) inhibitor U73122, orexin-A did not change the IBa,L of GCs, whereas the orexin-A effect persisted in the presence of the phosphatidylcholine (PC)-PLC inhibitor D609. The orexin-A-induced potentiation was not seen with internal infusion of Ca(2+)-free solution or when inositol 1,4,5-trisphosphate (IP3)-sensitive Ca(2+) release from intracellular stores was blocked by heparin/xestospongins-C. Moreover, the orexin-A effect was mimicked by the protein kinase C (PKC) activator phorbol 12-myristate 13-acetate, but was eliminated when PKC was inhibited by bisindolylmaleimide IV (Bis-IV)/Gö6976. Neither adenosine 3',5'-cyclic monophosphate (cAMP)-protein kinase A (PKA) nor guanosine 3',5'-cyclic monophosphate (cGMP)-protein kinase G (PKG) signaling pathway was likely involved, as orexin-A persisted to potentiate the IBa,L of GCs no matter these two pathways were activated or inhibited. These results suggest that, by activating OX1R, orexin-A potentiates the IBa,L of rat GCs through a distinct Gq/PI-PLC/IP3/Ca(2+)/PKC signaling pathway. Copyright

  19. Coatings of Different Carbon Nanotubes on Platinum Electrodes for Neuronal Devices: Preparation, Cytocompatibility and Interaction with Spiral Ganglion Cells.

    Science.gov (United States)

    Burblies, Niklas; Schulze, Jennifer; Schwarz, Hans-Christoph; Kranz, Katharina; Motz, Damian; Vogt, Carla; Lenarz, Thomas; Warnecke, Athanasia; Behrens, Peter

    2016-01-01

    Cochlear and deep brain implants are prominent examples for neuronal prostheses with clinical relevance. Current research focuses on the improvement of the long-term functionality and the size reduction of neural interface electrodes. A promising approach is the application of carbon nanotubes (CNTs), either as pure electrodes but especially as coating material for electrodes. The interaction of CNTs with neuronal cells has shown promising results in various studies, but these appear to depend on the specific type of neurons as well as on the kind of nanotubes. To evaluate a potential application of carbon nanotube coatings for cochlear electrodes, it is necessary to investigate the cytocompatibility of carbon nanotube coatings on platinum for the specific type of neuron in the inner ear, namely spiral ganglion neurons. In this study we have combined the chemical processing of as-delivered CNTs, the fabrication of coatings on platinum, and the characterization of the electrical properties of the coatings as well as a general cytocompatibility testing and the first cell culture investigations of CNTs with spiral ganglion neurons. By applying a modification process to three different as-received CNTs via a reflux treatment with nitric acid, long-term stable aqueous CNT dispersions free of dispersing agents were obtained. These were used to coat platinum substrates by an automated spray-coating process. These coatings enhance the electrical properties of platinum electrodes, decreasing the impedance values and raising the capacitances. Cell culture investigations of the different CNT coatings on platinum with NIH3T3 fibroblasts attest an overall good cytocompatibility of these coatings. For spiral ganglion neurons, this can also be observed but a desired positive effect of the CNTs on the neurons is absent. Furthermore, we found that the well-established DAPI staining assay does not function on the coatings prepared from single-wall nanotubes.

  20. The novel cyclophilin D inhibitor compound 19 protects retinal pigment epithelium cells and retinal ganglion cells from UV radiation.

    Science.gov (United States)

    Xie, Laiqing; Cheng, Long; Xu, Guoxu; Zhang, Ji; Ji, Xiaoyan; Song, E

    2017-06-10

    Excessive Ultra violet (UV) radiation induces injuries to retinal pigment epithelium (RPE) cells (RPEs) and retinal ganglion cells (RGCs), causing retinal degeneration. Cyclophilin D (Cyp-D)-dependent mitochondrial permeability transition pore (mPTP) opening mediates UV-induced cell death. In this study, we show that a novel Cyp-D inhibitor compound 19 efficiently protected RPEs and RGCs from UV radiation. Compound 19-mediated cytoprotection requires Cyp-D, as it failed to further protect RPEs/RGCs from UV when Cyp-D was silenced by targeted shRNAs. Compound 19 almost blocked UV-induced p53-Cyp-D mitochondrial association, mPTP opening and subsequent cytochrome C release. Further studies showed that compound 19 inhibited UV-induced reactive oxygen species (ROS) production, lipid peroxidation and DNA damage. Together, compound 19 protects RPEs and RGCs from UV radiation, possibly via silencing Cyp-D-regulated intrinsic mitochondrial death pathway. Compound 19 could a lead compound for treating UV-associated retinal degeneration diseases. Copyright © 2017 Elsevier Inc. All rights reserved.

  1. Isolation and Molecular Profiling of Primary Mouse Retinal Ganglion Cells: Comparison of Phenotypes from Healthy and Glaucomatous Retinas

    OpenAIRE

    Chintalapudi, Sumana R.; Djenderedjian, Levon; Stiemke, Andrew B.; Steinle, Jena J.; Jablonski, Monica M.; Morales-Tirado, Vanessa M.

    2016-01-01

    Loss of functional retinal ganglion cells (RGC) is an element of retinal degeneration that is poorly understood. This is in part due to the lack of a reliable and validated protocol for the isolation of primary RGCs. Here we optimize a feasible, reproducible, standardized flow cytometry-based protocol for the isolation and enrichment of homogeneous RGC with the Thy1.2hiCD48negCD15negCD57neg surface phenotype. A three-step validation process was performed by: (1) genomic profiling of 25-genes ...

  2. Retinal ganglion cells: mechanisms underlying depolarization block and differential responses to high frequency electrical stimulation of ON and OFF cells

    Science.gov (United States)

    Kameneva, T.; Maturana, M. I.; Hadjinicolaou, A. E.; Cloherty, S. L.; Ibbotson, M. R.; Grayden, D. B.; Burkitt, A. N.; Meffin, H.

    2016-02-01

    Objective. ON and OFF retinal ganglion cells (RGCs) are known to have non-monotonic responses to increasing amplitudes of high frequency (2 kHz) biphasic electrical stimulation. That is, an increase in stimulation amplitude causes an increase in the cell’s spike rate up to a peak value above which further increases in stimulation amplitude cause the cell to decrease its activity. The peak response for ON and OFF cells occurs at different stimulation amplitudes, which allows differential stimulation of these functional cell types. In this study, we investigate the mechanisms underlying the non-monotonic responses of ON and OFF brisk-transient RGCs and the mechanisms underlying their differential responses. Approach. Using in vitro patch-clamp recordings from rat RGCs, together with simulations of single and multiple compartment Hodgkin-Huxley models, we show that the non-monotonic response to increasing amplitudes of stimulation is due to depolarization block, a change in the membrane potential that prevents the cell from generating action potentials. Main results. We show that the onset for depolarization block depends on the amplitude and frequency of stimulation and reveal the biophysical mechanisms that lead to depolarization block during high frequency stimulation. Our results indicate that differences in transmembrane potassium conductance lead to shifts of the stimulus currents that generate peak spike rates, suggesting that the differential responses of ON and OFF cells may be due to differences in the expression of this current type. We also show that the length of the axon’s high sodium channel band (SOCB) affects non-monotonic responses and the stimulation amplitude that leads to the peak spike rate, suggesting that the length of the SOCB is shorter in ON cells. Significance. This may have important implications for stimulation strategies in visual prostheses.

  3. Overexpression of Pax6 results in microphthalmia, retinal dysplasia and defective retinal ganglion cell axon guidance

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    Jeffery Glen

    2008-05-01

    Full Text Available Abstract Background The transcription factor Pax6 is expressed by many cell types in the developing eye. Eyes do not form in homozygous loss-of-function mouse mutants (Pax6Sey/Sey and are abnormally small in Pax6Sey/+ mutants. Eyes are also abnormally small in PAX77 mice expressing multiple copies of human PAX6 in addition to endogenous Pax6; protein sequences are identical in the two species. The developmental events that lead to microphthalmia in PAX77 mice are not well-characterised, so it is not clear whether over- and under-expression of Pax6/PAX6 cause microphthalmia through similar mechanisms. Here, we examined the consequences of over-expression for the eye and its axonal connections. Results Eyes form in PAX77+/+ embryos but subsequently degenerate. At E12.5, we found no abnormalities in ocular morphology, retinal cell cycle parameters and the incidence of retinal cell death. From E14.5 on, we observed malformations of the optic disc. From E16.5 into postnatal life there is progressively more severe retinal dysplasia and microphthalmia. Analyses of patterns of gene expression indicated that PAX77+/+ retinae produce a normal range of cell types, including retinal ganglion cells (RGCs. At E14.5 and E16.5, quantitative RT-PCR with probes for a range of molecules associated with retinal development showed only one significant change: a slight reduction in levels of mRNA encoding the secreted morphogen Shh at E16.5. At E16.5, tract-tracing with carbocyanine dyes in PAX77+/+ embryos revealed errors in intraretinal navigation by RGC axons, a decrease in the number of RGC axons reaching the thalamus and an increase in the proportion of ipsilateral projections among those RGC axons that do reach the thalamus. A survey of embryos with different Pax6/PAX6 gene dosage (Pax6Sey/+, Pax6+/+, PAX77+ and PAX77+/+ showed that (1 the total number of RGC axons projected by the retina and (2 the proportions that are sorted into the ipsilateral and

  4. Delayed neurogenesis leads to altered specification of ventrotemporal retinal ganglion cells in albino mice

    Science.gov (United States)

    2014-01-01

    Background Proper binocular vision depends on the routing at the optic chiasm of the correct proportion of retinal ganglion cell (RGC) axons that project to the same (ipsilateral) and opposite (contralateral) side of the brain. The ipsilateral RGC projection is reduced in mammals with albinism, a congenital disorder characterized by deficient pigmentation in the skin, hair, and eyes. Compared to the pigmented embryonic mouse retina, the albino embryonic mouse retina has fewer RGCs that express the zinc-finger transcription factor, Zic2, which is transiently expressed by RGCs fated to project ipsilaterally. Here, using Zic2 as a marker of ipsilateral RGCs, Islet2 as a marker of contralateral RGCs, and birthdating, we investigate spatiotemporal dynamics of RGC production as they relate to the phenotype of diminished ipsilateral RGC number in the albino retina. Results At embryonic day (E)15.5, fewer Zic2-positive (Zic2+) RGCs are found in the albino ventrotemporal (VT) retina compared with the pigmented VT retina, as we previously reported. However, the reduction in Zic2+ RGCs in the albino is not accompanied by a compensatory increase in Zic2-negative (Zic2−) RGCs, resulting in fewer RGCs in the VT retina at this time point. At E17.5, however, the number of RGCs in the VT region is similar in pigmented and albino retinae, implicating a shift in the timing of RGC production in the albino. Short-term birthdating assays reveal a delay in RGC production in the albino VT retina between E13 and E15. Specifically, fewer Zic2+ RGCs are born at E13 and more Zic2− RGCs are born at E15. Consistent with an increase in the production of Zic2− RGCs born at later ages, more RGCs at E17.5 express the contralateral marker, Islet2, in the albino VT retina compared with the pigmented retina. Conclusions A delay in neurogenesis in the albino retina is linked to the alteration of RGC subtype specification and consequently leads to the reduced ipsilateral projection that

  5. Retinal ganglion cell-inner plexiform and nerve fiber layers in neuromyelitis optica.

    Science.gov (United States)

    Hu, Sai-Jing; Lu, Pei-Rong

    2018-01-01

    To determine the thickness of the retinal ganglion cell-inner plexiform layer (GCIPL) and the retinal nerve fiber layer (RNFL) in patients with neuromyelitis optica (NMO). We conducted a cross-sectional study that included 30 NMO patients with a total of 60 eyes. Based on the presence or absence of optic neuritis (ON), subjects were divided into either the NMO-ON group (30 eyes) or the NMO-ON contra group (10 eyes). A detailed ophthalmologic examination was performed for each group; subsequently, the GCIPL and the RNFL were measured using high-definition optical coherence tomography (OCT). In the NMO-ON group, the mean GCIPL thickness was 69.28±21.12 µm, the minimum GCIPL thickness was 66.02±10.02 µm, and the RNFL thickness were 109.33±11.23, 110.47±3.10, 64.92±12.71 and 71.21±50.22 µm in the superior, inferior, temporal and nasal quadrants, respectively. In the NMO-ON contra group, the mean GCIPL thickness was 85.12±17.09 µm, the minimum GCIPL thickness was 25.39±25.1 µm, and the RNFL thicknesses were 148.33±23.22, 126.36±23.45, 82.21±22.30 and 83.36±31.28 µm in the superior, inferior, temporal and nasal quadrants, respectively. In the control group, the mean GCIPL thickness was 86.98±22.37 µm, the minimum GCIPL thickness was 85.28±10.75 µm, and the RNFL thicknesses were 150.22±22.73, 154.79±60.23, 82.33±7.01 and 85.62±13.81 µm in the superior, inferior, temporal and nasal quadrants, respectively. The GCIPL and RNFL were thinner in the NMO-ON contra group than in the control group ( P deviation (MD) and corrected pattern standard deviation (PSD) in the NMO-ON group ( P <0.05). The thickness of the GCIPL and RNFL, as measured using OCT, may indicate optic nerve damage in patients with NMO.

  6. Interleukin-6 Deficiency Attenuates Retinal Ganglion Cell Axonopathy and Glaucoma-Related Vision Loss

    Directory of Open Access Journals (Sweden)

    Franklin D. Echevarria

    2017-05-01

    Full Text Available The pleotropic cytokine interleukin-6 (IL-6 is implicated in retinal ganglion cell (RGC survival and degeneration, including that associated with glaucoma. IL-6 protects RGCs from pressure-induced apoptosis in vitro. However, it is unknown how IL-6 impacts glaucomatous degeneration in vivo. To study how IL-6 influences glaucomatous RGC axonopathy, accompanying glial reactivity, and resultant deficits in visual function, we performed neural tracing, histological, and neurobehavioral assessments in wildtype (B6;129SF2/J; WT and IL-6 knock-out mice (B6;129S2-IL6tm1kopf/J; IL-6-/- after 8 weeks of unilateral or bilateral microbead-induced glaucoma (microbead occlusion model. IOP increased by 20% following microbead injection in both genotypes (p < 0.05. However, deficits in wound healing at the site of corneal injection were noted. In WT mice, elevated IOP produced degenerating axon profiles and decreased axon density in the optic nerve by 15% (p < 0.01. In IL-6-/- mice, axon density in the optic nerve did not differ between microbead- and saline-injected mice (p > 0.05 and degenerating axon profiles were minimal. Preservation of RGC axons was reflected in visual function, where visual acuity decreased significantly in a time-dependent manner with microbead-induced IOP elevation in WT (p < 0.001, but not IL-6-/- mice (p > 0.05. Despite this preservation of RGC axons and visual acuity, both microbead-injected WT and IL-6-/- mice exhibited a 50% decrease in anterograde CTB transport to the superior colliculus, as compared to saline-injected controls (p < 0.01. Assessment of glial reactivity revealed no genotype- or IOP-dependent changes in retinal astrocytes. IOP elevation decreased microglia density and percent retinal area covered in WT mice (p < 0.05, while IL-6-/- mice exhibited only a decrease in density (p < 0.05. Together, our findings indicate that two defining features of RGC axonopathy—axon transport deficits and structural degeneration of

  7. Neuroprotection of rat retinal ganglion cells mediated through alpha7 nicotinic acetylcholine receptors.

    Science.gov (United States)

    Iwamoto, K; Mata, D; Linn, D M; Linn, C L

    2013-05-01

    Glutamate-induced excitotoxicity is thought to play an important role in several neurodegenerative diseases in the central nervous system (CNS). In this study, neuroprotection against glutamate-induced excitotoxicity was analyzed using acetylcholine (ACh), nicotine and the α7 specific nicotinic acetylcholine receptor (α7 nAChR) agonist, N-[(3R)-1-azabicyclo[2.2.2]oct-3-yl]-4-chlorobenzamide hydrochloride (PNU-282987), in cultured adult rat retinal neurons. Adult Long Evans rat retinas were dissociated and retinal ganglion cells (RGCs) were isolated from all other retinal tissue using a two-step panning technique. Once isolated, RGCs were cultured under various pharmacological conditions to demonstrate excitotoxicity and neuroprotection against excitotoxicity. After 3 days, RGCs were immunostained with antibodies against the glycoprotein, Thy 1.1, counted and cell survival was assessed relative to control untreated conditions. 500 μM glutamate induced excitotoxicity in large and small RGCs in an adult rat dissociated culture. After 3 days in culture with glutamate, the cell survival of large RGCs decreased by an average of 48.16% while the cell survival of small RGCs decreased by an average of 42.03%. Using specific glutamate receptor agonists and antagonists, we provide evidence that the excitotoxic response was mediated through α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA)/kainic acid (KA) and N-methyl-d-aspartate (NMDA) glutamate receptors through an apoptotic mechanism. However, the excitotoxic effect of glutamate on all RGCs was eliminated if cells were cultured for an hour with 10 μM ACh, 100 μM nicotine or 100 nM of the α7 nAChR agonist, PNU-282987, before the glutamate insult. Inhibition studies using 10nM methyllycaconitine (MLA) or α-bungarotoxin (α-Bgt) supported the hypothesis that neuroprotection against glutamate-induced excitotoxicity on rat RGCs was mediated through α7 nAChRs. In immunocytochemical studies, double

  8. Co-expression of two subtypes of melatonin receptor on rat M1-type intrinsically photosensitive retinal ganglion cells.

    Directory of Open Access Journals (Sweden)

    Wen-Long Sheng

    Full Text Available Intrinsically photosensitive retinal ganglion cells (ipRGCs are involved in circadian and other non-image forming visual responses. An open question is whether the activity of these neurons may also be under the regulation mediated by the neurohormone melatonin. In the present work, by double-staining immunohistochemical technique, we studied the expression of MT1 and MT2, two known subtypes of mammalian melatonin receptors, in rat ipRGCs. A single subset of retinal ganglion cells labeled by the specific antibody against melanopsin exhibited the morphology typical of M1-type ipRGCs. Immunoreactivity for both MT1 and MT2 receptors was clearly seen in the cytoplasm of all labeled ipRGCs, indicating that these two receptors were co-expressed in each of these neurons. Furthermore, labeling for both the receptors were found in neonatal M1 cells as early as the day of birth. It is therefore highly plausible that retinal melatonin may directly modulate the activity of ipRGCs, thus regulating non-image forming visual functions.

  9. Pleomorphic hyalinizing angiectatic tumor of soft parts: case report with unusual ganglion-like cells and review of the literature.

    Science.gov (United States)

    Changchien, Yi-Che; Bocskai, Pál; Kovács, Ilona; Hargitai, Zoltán; Kollár, Sándor; Török, Miklós

    2014-12-01

    Pleomorphic hyalinizing angiectatic tumor (PHAT) is a recently described, non-metastasizing tumor of uncertain lineage. This tumor distributes equally between the genders and has a predilection for the subcutaneous soft tissue, particularly in lower extremity, other locations are rare. Based on the recent literature, PHAT is suspected to encompass the morphological spectrum with other tumors such as myxoinflammatory fibroblastic sarcoma (MIFS) and hemosiderotic fibrolipomatous tumor (HFLT), although cytogenetic data remain inconsistent. We report a case of PHAT that arose in the upper arm with unusual morphology which showed ganglion-like cells similar to Reed-Sternberg-like cells found in MIFS. The tumor had strong immunohistochemical expression of CD34, CD99, and was negative for S-100. The ganglion-like cells were positive for both CD34 and CD68 but negative for CD30. The translocation between chromosome 1 and 10, a frequent finding of MIFS and HFLT, was not identified by FISH excluding the possibility of hybrid PHAT and MIFS. We conclude FISH can be a potential useful tool to separate PHAT with atypical morphology from hybrid tumor in doubted cases. Due to the rarity of PHAT and lack of consistent pathogenetic signatures, more cases and further studies will be needed to elucidate the pathogenesis and nature of this tumor. Copyright © 2014 Elsevier GmbH. All rights reserved.

  10. Medial ganglionic eminence–like cells derived from human embryonic stem cells correct learning and memory deficits

    Science.gov (United States)

    Liu, Yan; Weick, Jason P; Liu, Huisheng; Krencik, Robert; Zhang, Xiaoqing; Ma, Lixiang; Zhou, Guo-min; Ayala, Melvin; Zhang, Su-Chun

    2013-01-01

    Dysfunction of basal forebrain cholinergic neurons (BFCNs) and γ-aminobutyric acid (GABA) interneurons, derived from medial ganglionic eminence (MGE), is implicated in disorders of learning and memory. Here we present a method for differentiating human embryonic stem cells (hESCs) to a nearly uniform population of NKX2.1+ MGE-like progenitor cells. After transplantation into the hippocampus of mice in which BFCNs and some GABA neurons in the medial septum had been destroyed by mu P75-saporin, human MGE-like progenitors, but not ventral spinal progenitors, produced BFCNs that synaptically connected with endogenous neurons, whereas both progenitors generated similar populations of GABA neurons. Mice transplanted with MGE-like but not spinal progenitors showed improvements in learning and memory deficits. These results suggest that progeny of the MGE-like progenitors, particularly BFCNs, contributed to learning and memory. Our findings support the prospect of using human stem cell–derived MGE-like progenitors in developing therapies for neurological disorders of learning and memory. PMID:23604284

  11. The influence of venous blood flow on the retinal ganglion cell complex in patients with primary open angle glaucoma

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    N. I. Kurysheva

    2014-07-01

    Full Text Available Purpose: To study the influence of venous blood flow on the ganglion cell complex (GCC in patients with preperimetric and perimetric open angle glaucoma.Methods: 74 patients were included in the research. 59 eyes and 62 eyes were diagnosed with preperimetric and perimetric open angle glaucoma respectively. The mean age was 56.5±10.5 years. 22 (12 female and 10 male healthy individuals constituted the control group. The ganglion cell complex and retinal nerve fibre layer were evaluated with the help of optical coherence tomography (RTVue-100 OCT, Optovue, Inc., Fremont, CA. Ocular blood flow was measured by Color Doppler Imaging (multifunctional VOLUSON 730 ProSystem. The statistical analysis included correlation between GCC and RNFL thickness in both glaucoma groups.Results: The results showed a statistically significant reduction of venous blood flow velocity in both glaucoma groups compared to the control group. No difference in venous blood flow parameters between two glaucoma groups was found, except resistance index, which was higher in perimetric group in comparison to preperimetric group. A correlation was also obtained between venous blood flow parameters and GCC and RNFL thickness in both glaucoma groups.Conclusion: Early GCC damage in glaucoma might occur due to venous blood flow reduction. This fact may be of great value in understanding glaucoma pathogenesis and search for novel treatment options.

  12. Effect of SIRT1 regulating cholesterol synthesis in repairing retinal ganglion cells after optic nerve injury in rats

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    Yan Zhang

    2014-10-01

    Full Text Available AIM: To investigate the repair mechanism associated with cholesterol synthesis regulated by silent information regulator 1(SIRT1in rat model of optic nerve damage. METHODS: Preparation of optic nerve damage in 70 rats was randomly divided into normal group(10 rats, resveratrol treatment group(experimental group 30 ratsand PBS buffer control group(30 rats. The experimental group and control group was further divided into 3 subgroups(each group 10 rats, respectively. After 7, 14, 21d injected resveratrol or PBS, optic nerve injury were observed, then the rats were sacrificed. Retina was segregated; the surviving retinal ganglion cell(RGCswas counted. Dissection of optic nerve, cholesterol content of them were tested; RT-PCR was used to detect mRNA expression of SIRT1, SREBP2 and HMGCR; Western blot assay was used to test the protein expression levels of SIRT1, cholesterol regulatory element binding protein 2(SREBP2and HMGCR. RESULTS: The numbers of RGCs and cholesterol levels of rat model with optic nerve injury decreased significantly(PPPPCONCLUSION: Up-regulating the expression of SIRT1, SREBP2 and down-regulating HMGCR by resveratrol could repair the injury of optic nerve through promoting the synthesis of cholesterol in neurons and retinal ganglion cells in the repair process. SIRT1 may be as a promising new target for treatment on optic nerve damage.

  13. Melanopsin-expressing retinal ganglion cells are resistant to cell injury, but not always

    DEFF Research Database (Denmark)

    Georg, Birgitte; Ghelli, Anna; Giordano, Carla

    2017-01-01

    Atrophy). However, these cells are affected in other neurodegenerative conditions, such as glaucoma and Alzheimer's disease. We here review the current evidences that may underlie this dichotomy. We also present our unpublished data on cell experiments demonstrating that melanopsin itself does not explain...

  14. The intricacies of neurotrophic factor therapy for retinal ganglion cell rescue in glaucoma: a case for gene therapy

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    Marianna Foldvari

    2016-01-01

    Full Text Available Regeneration of damaged retinal ganglion cells (RGC and their axons is an important aspect of reversing vision loss in glaucoma patients. While current therapies can effectively lower intraocular pressure, they do not provide extrinsic support to RGCs to actively aid in their protection and regeneration. The unmet need could be addressed by neurotrophic factor gene therapy, where plasmid DNA, encoding neurotrophic factors, is delivered to retinal cells to maintain sufficient levels of neurotrophins in the retina. In this review, we aim to describe the intricacies in the design of the therapy including: the choice of neurotrophic factor, the site and route of administration and target cell populations for gene delivery. Furthermore, we also discuss the challenges currently being faced in RGC-related therapy development with special considerations to the existence of multiple RGC subtypes and the lack of efficient and representative in vitro models for rapid and reliable screening in the drug development process.

  15. Asymmetry between ON and OFF α ganglion cells of mouse retina: integration of signal and noise from synaptic inputs.

    Science.gov (United States)

    Freed, Michael A

    2017-11-15

    Bipolar and amacrine cells presynaptic to the ON sustained α cell of mouse retina provide currents with a higher signal-to-noise power ratio (SNR) than those presynaptic to the OFF sustained α cell. Yet the ON cell loses proportionately more SNR from synaptic inputs to spike output than the OFF cell does. The higher SNR of ON bipolar cells at the beginning of the ON pathway compensates for losses incurred by the ON ganglion cell, and improves the processing of positive contrasts. ON and OFF pathways in the retina include functional pairs of neurons that, at first glance, appear to have symmetrically similar responses to brightening and darkening, respectively. Upon careful examination, however, functional pairs exhibit asymmetries in receptive field size and response kinetics. Until now, descriptions of how light-adapted retinal circuitry maintains a preponderance of signal over the noise have not distinguished between ON and OFF pathways. Here I present evidence of marked asymmetries between members of a functional pair of sustained α ganglion cells in the mouse retina. The ON cell exhibited a proportionately greater loss of signal-to-noise power ratio (SNR) from its presynaptic arrays to its postsynaptic currents. Thus the ON cell combines signal and noise from its presynaptic arrays of bipolar and amacrine cells less efficiently than the OFF cell does. Yet the inefficiency of the ON cell is compensated by its presynaptic arrays providing a higher SNR than the arrays presynaptic to the OFF cell, apparently to improve visual processing of positive contrasts. Dynamic clamp experiments were performed that introduced synaptic conductances into ON and OFF cells. When the amacrine-modulated conductance was removed, the ON cell's spike train exhibited an increase in SNR. The OFF cell, however, showed the opposite effect of removing amacrine input, which was a decrease in SNR. Thus ON and OFF cells have different modes of synaptic integration with direct effects on

  16. Chlorpyrifos- and chlorpyrifos oxon-induced neurite retraction in pre-differentiated N2a cells is associated with transient hyperphosphorylation of neurofilament heavy chain and ERK 1/2

    International Nuclear Information System (INIS)

    Sindi, Ramya A.; Harris, Wayne; Arnott, Gordon; Flaskos, John; Lloyd Mills, Chris; Hargreaves, Alan J.

    2016-01-01

    Chlorpyrifos (CPF) and CPF-oxon (CPO) are known to inhibit neurite outgrowth but little is known about their ability to induce neurite retraction in differentiating neuronal cells. The aims of this study were to determine the ability of these compounds to destabilize neurites and to identify the key molecular events involved. N2a cells were induced to differentiate for 20 h before exposure to CPF or CPO for 2–8 h. Fixed cell monolayers labeled with carboxyfluorescein succinimidyl ester or immunofluorescently stained with antibodies to tubulin (B512) or phosphorylated neurofilament heavy chain (Ta51) showed time- and concentration-dependent reductions in numbers and length of axon-like processes compared to the control, respectively, retraction of neurites being observed within 2 h of exposure by live cell imaging. Neurofilament disruption was also observed in treated cells stained by indirect immunofluorescence with anti-phosphorylated neurofilament heavy chain (NFH) monoclonal antibody SMI34, while the microtubule network was unaffected. Western blotting analysis revealed transiently increased levels of reactivity of Ta51 after 2 h exposure and reduced levels of reactivity of the same antibody following 8 h treatment with both compounds, whereas reactivity with antibodies to anti-total NFH or anti-tubulin was not affected. The alteration in NFH phosphorylation at 2 h exposure was associated with increased activation of extracellular signal-regulated protein kinase ERK 1/2. However, increased levels of phosphatase activity were observed following 8 h exposure. These findings suggest for the first time that organophosphorothionate pesticide-induced neurite retraction in N2a cells is associated with transient increases in NFH phosphorylation and ERK1/2 activation. - Highlights: • Chlorpyrifos and chlorpyrifos oxon induced rapid neurite retraction in N2a cells. • This occurred following transient hyperphosphorylation of ERK 1/2. • It was concomitant with

  17. Retinofugal Projections from Melanopsin-Expressing Retinal Ganglion Cells Revealed by Intraocular Injections of Cre-Dependent Virus.

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    Anton Delwig

    Full Text Available To understand visual functions mediated by intrinsically photosensitive melanopsin-expressing retinal ganglion cells (mRGCs, it is important to elucidate axonal projections from these cells into the brain. Initial studies reported that melanopsin is expressed only in retinal ganglion cells within the eye. However, recent studies in Opn4-Cre mice revealed Cre-mediated marker expression in multiple brain areas. These discoveries complicate the use of melanopsin-driven genetic labeling techniques to identify retinofugal projections specifically from mRGCs. To restrict labeling to mRGCs, we developed a recombinant adeno-associated virus (AAV carrying a Cre-dependent reporter (human placental alkaline phosphatase that was injected into the vitreous of Opn4-Cre mouse eyes. The labeling observed in the brain of these mice was necessarily restricted specifically to retinofugal projections from mRGCs in the injected eye. We found that mRGCs innervate multiple nuclei in the basal forebrain, hypothalamus, amygdala, thalamus and midbrain. Midline structures tended to be bilaterally innervated, whereas the lateral structures received mostly contralateral innervation. As validation of our approach, we found projection patterns largely corresponded with previously published results; however, we have also identified a few novel targets. Our discovery of projections to the central amygdala suggests a possible direct neural pathway for aversive responses to light in neonates. In addition, projections to the accessory optic system suggest that mRGCs play a direct role in visual tracking, responses that were previously attributed to other classes of retinal ganglion cells. Moreover, projections to the zona incerta raise the possibility that mRGCs could regulate visceral and sensory functions. However, additional studies are needed to investigate the actual photosensitivity of mRGCs that project to the different brain areas. Also, there is a concern of "overlabeling

  18. Retinal ganglion cell-inner plexiform and nerve fiber layers in neuromyelitis optica

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    Sai-Jing Hu

    2018-01-01

    Full Text Available AIM: To determine the thickness of the retinal ganglion cell-inner plexiform layer (GCIPL and the retinal nerve fiber layer (RNFL in patients with neuromyelitis optica (NMO. METHODS: We conducted a cross-sectional study that included 30 NMO patients with a total of 60 eyes. Based on the presence or absence of optic neuritis (ON, subjects were divided into either the NMO-ON group (30 eyes or the NMO-ON contra group (10 eyes. A detailed ophthalmologic examination was performed for each group; subsequently, the GCIPL and the RNFL were measured using high-definition optical coherence tomography (OCT. RESULTS: In the NMO-ON group, the mean GCIPL thickness was 69.28±21.12 μm, the minimum GCIPL thickness was 66.02±10.02 μm, and the RNFL thickness were 109.33±11.23, 110.47±3.10, 64.92±12.71 and 71.21±50.22 μm in the superior, inferior, temporal and nasal quadrants, respectively. In the NMO-ON contra group, the mean GCIPL thickness was 85.12±17.09 μm, the minimum GCIPL thickness was 25.39±25.1 μm, and the RNFL thicknesses were 148.33±23.22, 126.36±23.45, 82.21±22.30 and 83.36±31.28 μm in the superior, inferior, temporal and nasal quadrants, respectively. In the control group, the mean GCIPL thickness was 86.98±22.37 μm, the minimum GCIPL thickness was 85.28±10.75 μm, and the RNFL thicknesses were 150.22±22.73, 154.79±60.23, 82.33±7.01 and 85.62±13.81 μm in the superior, inferior, temporal and nasal quadrants, respectively. The GCIPL and RNFL were thinner in the NMO-ON contra group than in the control group (P<0.05; additionally, the RNFL was thinner in the inferior quadrant in the NMO-ON group than in the control group (P<0.05. Significant correlations were observed between the GCIPL and RNFL thickness measurements as well as between thickness measurements and the two visual field parameters of mean deviation (MD and corrected pattern standard deviation (PSD in the NMO-ON group (P<0.05. CONCLUSION: The thickness of the GCIPL

  19. Proteomic analysis of differentiating neuroblastoma cells treated with sub-lethal neurite inhibitory concentrations of diazinon: Identification of novel biomarkers of effect

    International Nuclear Information System (INIS)

    Harris, W.; Sachana, M.; Flaskos, J.; Hargreaves, A.J.

    2009-01-01

    In previous work we showed that sub-lethal levels of diazinon inhibited neurite outgrowth in differentiating N2a neuroblastoma cells. Western blotting analysis targeted at proteins involved in axon growth and stress responses, revealed that such exposure led to a reduction in the levels of neurofilament heavy chain, microtubule associated protein 1 B (MAP 1B) and HSP-70. The aim of this study was to apply the approach of 2 dimensional polyacrylamide gel electrophoresis and mass spectrometry to identify novel biomarkers of effect. A number of proteins were found to be up-regulated compared to the control on silver-stained gels. These were classified in to 3 main groups of proteins: cytosolic factors, chaperones and the actin-binding protein cofilin, all of which are involved in cell differentiation, survival or metabolism. The changes observed for cofilin were further confirmed by quantitative Western blotting analysis with anti-actin and anti-cofilin antibodies. Indirect immunofluorescence staining with the same antibodies indicated that the microfilament network was disrupted in diazinon-treated cells. Our data suggest that microfilament organisation is disrupted by diazinon exposure, which may be related to increased cofilin expression.

  20. Microelectrode array-induced neuronal alignment directs neurite outgrowth: analysis using a fast Fourier transform (FFT).

    Science.gov (United States)

    Radotić, Viktorija; Braeken, Dries; Kovačić, Damir

    2017-12-01

    Many studies have shown that the topography of the substrate on which neurons are cultured can promote neuronal adhesion and guide neurite outgrowth in the same direction as the underlying topography. To investigate this effect, isotropic substrate-complementary metal-oxide-semiconductor (CMOS) chips were used as one example of microelectrode arrays (MEAs) for directing neurite growth of spiral ganglion neurons. Neurons were isolated from 5 to 7-day-old rat pups, cultured 1 day in vitro (DIV) and 4 DIV, and then fixed with 4% paraformaldehyde. For analysis of neurite alignment and orientation, fast Fourier transformation (FFT) was used. Results revealed that on the micro-patterned surface of a CMOS chip, neurons orient their neurites along three directional axes at 30, 90, and 150° and that neurites aligned in straight lines between adjacent pillars and mostly followed a single direction while occasionally branching perpendicularly. We conclude that the CMOS substrate guides neurites towards electrodes by means of their structured pillar organization and can produce electrical stimulation of aligned neurons as well as monitoring their neural activities once neurites are in the vicinity of electrodes. These findings are of particular interest for neural tissue engineering with the ultimate goal of developing a new generation of MEA essential for improved electrical stimulation of auditory neurons.

  1. Protective Effect of ALA in Crushed Optic Nerve Cat Retinal Ganglion Cells Using a New Marker RBPMS.

    Science.gov (United States)

    Wang, Yanling; Wang, Wenyao; Liu, Jessica; Huang, Xin; Liu, Ruixing; Xia, Huika; Brecha, Nicholas C; Pu, Mingliang; Gao, Jie

    2016-01-01

    In this study we first sought to determine whether RNA-binding protein with multiple splicing (RBPMS) can serve as a specific marker for cat retina ganglion cells (RGCs) using retrograde labeling and immunohistochemistry staining. RBPM was then used as an RGC marker to study RGC survival after optic nerve crush (ONC) and alpha-lipoic acid (ALA) treatment in cats. ALA treatment yielded a peak density of RBPMS-alpha cells within the peak isodensity zone (>60/mm2) which did not differ from ONC retinas. The area within the zone was significantly enlarged (control: 2.3%, ONC: 0.06%, ONC+ALA: 0.1%). As for the 10-21/mm2 zone, ALA treatment resulted in a significant increase in area (control: 34.5%, ONC: 12.1%, ONC+ALA: 35.9%). ALA can alleviate crush-induced RGC injury.

  2. Matrix metalloproteinase 2 and membrane type 1 matrix metalloproteinase co-regulate axonal outgrowth of mouse retinal ganglion cells

    DEFF Research Database (Denmark)

    Gaublomme, Djoere; Buyens, Tom; De Groef, Lies

    2014-01-01

    , we were able to show that broad-spectrum MMP inhibition reduces axon outgrowth of mouse retinal ganglion cells (RGCs), implicating MMPs as beneficial factors in axonal regeneration. Additional studies, using more specific MMP inhibitors and MMP-deficient mice, disclosed that both MMP-2 and MT1-MMP......, but not MMP-9, are involved in this process. Furthermore, administration of a novel antibody to MT1-MMP that selectively blocks pro-MMP-2 activation revealed a functional co-involvement of these proteinases in determining RGC axon outgrowth. Subsequent immunostainings showed expression of both MMP-2 and MT1......-MMP in RGC axons and glial cells. Finally, results from combined inhibition of MMP-2 and β1-integrin were suggestive for a functional interaction between these molecules. Overall, our data indicate MMP-2 and MT1-MMP as promising axonal outgrowth-promoting molecules. Axonal regeneration in the central...

  3. Activated Müller Cells Involved in ATP-Induced Upregulation of P2X7 Receptor Expression and Retinal Ganglion Cell Death

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    Ying Xue

    2016-01-01

    Full Text Available P2X7 receptor (P2X7R, an ATP-gated ion channel, plays an important role in glaucomatous retinal ganglion cell (RGC apoptotic death, in which activated retinal Müller glial cells may be involved by releasing ATP. In the present study, we investigated whether and how activated Müller cells may induce changes in P2X7R expression in RGCs by using immunohistochemistry and Western blot techniques. Intravitreal injection of DHPG, a group I metabotropic glutamate receptor (mGluR I agonist, induced upregulation of GFAP expression, suggestive of Müller cell activation (gliosis, as we previously reported. Accompanying Müller cell activation, P2X7R protein expression was upregulated, especially in the cells of ganglion cell layer (GCL, which was reversed by coinjection of brilliant blue G (BBG, a P2X7R blocker. In addition, intravitreal injection of ATP also induced upregulation of P2X7R protein expression. Similar results were observed in cultured retinal neurons by ATP treatment. Moreover, both DHPG and ATP intravitreal injection induced a reduction in the number of fluorogold retrogradely labeled RGCs, and the DHPG effect was partially rescued by coinjection of BBG. All these results suggest that activated Müller cells may release ATP and, in turn, induce upregulation of P2X7R expression in the cells of GCL, thus contributing to RGC death.

  4. Optimal voltage stimulation parameters for network-mediated responses in wild type and rd10 mouse retinal ganglion cells

    Science.gov (United States)

    Jalligampala, Archana; Sekhar, Sudarshan; Zrenner, Eberhart; Rathbun, Daniel L.

    2017-04-01

    To further improve the quality of visual percepts elicited by microelectronic retinal prosthetics, substantial efforts have been made to understand how retinal neurons respond to electrical stimulation. It is generally assumed that a sufficiently strong stimulus will recruit most retinal neurons. However, recent evidence has shown that the responses of some retinal neurons decrease with excessively strong stimuli (a non-monotonic response function). Therefore, it is necessary to identify stimuli that can be used to activate the majority of retinal neurons even when such non-monotonic cells are part of the neuronal population. Taking these non-monotonic responses into consideration, we establish the optimal voltage stimulation parameters (amplitude, duration, and polarity) for epiretinal stimulation of network-mediated (indirect) ganglion cell responses. We recorded responses from 3958 mouse retinal ganglion cells (RGCs) in both healthy (wild type, WT) and a degenerating (rd10) mouse model of retinitis pigmentosa—using flat-mounted retina on a microelectrode array. Rectangular monophasic voltage-controlled pulses were presented with varying voltage, duration, and polarity. We found that in 4-5 weeks old rd10 mice the RGC thresholds were comparable to those of WT. There was a marked response variability among mouse RGCs. To account for this variability, we interpolated the percentage of RGCs activated at each point in the voltage-polarity-duration stimulus space, thus identifying the optimal voltage-controlled pulse (-2.4 V, 0.88 ms). The identified optimal voltage pulse can activate at least 65% of potentially responsive RGCs in both mouse strains. Furthermore, this pulse is well within the range of stimuli demonstrated to be safe and effective for retinal implant patients. Such optimized stimuli and the underlying method used to identify them support a high yield of responsive RGCs and will serve as an effective guideline for future in vitro investigations of

  5. Negative impact of rAAV2 mediated expression of SOCS3 on the regeneration of adult retinal ganglion cell axons

    NARCIS (Netherlands)

    Hellstrom, M.; Muhling, J.; Ehlert, E.M.; Verhaagen, J.; Pollett, M.A.; Hu, Y.; Harvey, A.R.

    2011-01-01

    Intravitreal injections of recombinant ciliary neurotrophic factor (rCNTF) protect adult rat retinal ganglion cells (RGCs) after injury and stimulate regeneration, an effect enhanced by co-injection with a cAMP analogue (CPT-cAMP). This effect is partly mediated by PKA and associated signaling

  6. Detection of macular ganglion cell loss in preperimetric glaucoma patients with localized retinal nerve fibre defects by spectral-domain optical coherence tomography.

    Science.gov (United States)

    Na, Jung Hwa; Lee, Kyoungsub; Lee, Jong Rak; Baek, Seunghee; Yoo, Sung Jun; Kook, Michael S

    2013-12-01

    To evaluate and compare the utility of ganglion cell complex with peripapillary retinal nerve fibre layer and optic nerve head measurements for detection of localized defects in patients with preperimetric glaucoma using spectral-domain optical coherence tomography. Prospective study. Preperimetric glaucoma patients. A total of 105 eyes with preperimetric glaucoma and 68 age- and refractive error-matched control eyes were enrolled. The ability to detect localized retinal nerve fibre layer defects by RTVue-100 spectral-domain optical coherence tomography (Optovue, Inc., Fremont, CA, USA) was assessed calculating the areas under receiver operating characteristic curves. The ability to detect localized retinal nerve fibre layer defects by spectral-domain optical coherence tomography. Global volume loss and superior ganglion cell complex thickness showed the largest area under receiver operating characteristic curve values (both areas under receiver operating characteristic curves 0.84, P fibre layer thickness afforded the best diagnostic capability (area under receiver operating characteristic curve 0.89, P optic nerve head parameters, the horizontal cup:disc ratio yielded the highest area under receiver operating characteristic curve (0.85, P fibre layer, and optic nerve head) (P > 0.02). Ganglion cell complex thickness was significantly reduced in eyes with preperimetric glaucoma. Ganglion cell complex imaging using spectral-domain optical coherence tomography may be a useful ancillary modality for detection of early macular changes in glaucomatous eyes with localized retinal nerve fibre layer defects. © 2013 Royal Australian and New Zealand College of Ophthalmologists.

  7. Effect of myopia on ganglion cell complex and peripapillary retinal nerve fibre layer measurements: a Fourier-domain optical coherence tomography study of young Chinese persons.

    Science.gov (United States)

    Zhao, Zhennan; Jiang, Chunhui

    2013-08-01

    To investigate the change of the ganglion cell complex and peripapillary retinal nerve fibre layer under different refractive conditions, as measured by optical coherence tomography. Cross-sectional observational study. A total of 107 eyes from 107 subjects were studied. Ganglion cell complex and retinal nerve fibre layer were studied by a spectral-domain system. Their relationship against spherical equivalents and axial length was studied. The thickness of ganglion cell complex and retinal nerve fibre layer at different area and under different refractive conditions as measurements by optical coherence tomography. The average, superior and inferior macular ganglion cell complex thickness was significantly associated with both spherical equivalents (all P fibre layer thicknesses from the superior, inferior and temporal quadrants were associated with both spherical equivalents (all P fibre layer thickness was studied in sections, then some parts of the upper and lower temporal part showed no correlation with either spherical equivalents or axial length (all P fibre layer were correlated with refraction and axial length. Therefore, when using these for clinical purposes, attention must be paid to the refractive state of the patient. © 2012 The Authors. Clinical and Experimental Ophthalmology © 2012 Royal Australian and New Zealand College of Ophthalmologists.

  8. Patterns of Retinal Ganglion Cell Damage in Neurodegenerative Disorders: Parvocellular vs Magnocellular Degeneration in Optical Coherence Tomography Studies

    Directory of Open Access Journals (Sweden)

    Chiara La Morgia

    2017-12-01

    Full Text Available Many neurodegenerative disorders, such as Parkinson’s disease (PD and Alzheimer’s disease (AD, are characterized by loss of retinal ganglion cells (RGCs as part of the neurodegenerative process. Optical coherence tomography (OCT studies demonstrated variable degree of optic atrophy in these diseases. However, the pattern of degenerative changes affecting the optic nerve (ON can be different. In particular, neurodegeneration is more evident for magnocellular RGCs in AD and multiple system atrophy with a pattern resembling glaucoma. Conversely, in PD and Huntington’s disease, the parvocellular RGCs are more vulnerable. This latter pattern closely resembles that of mitochondrial optic neuropathies, possibly pointing to similar pathogenic mechanisms. In this review, the currently available evidences on OCT findings in these neurodegenerative disorders are summarized with particular emphasis on the different pattern of RGC loss. The ON degeneration could become a validated biomarker of the disease, which may turn useful to follow natural history and possibly assess therapeutic efficacy.

  9. Neurite, a finite difference large scale parallel program for the simulation of electrical signal propagation in neurites under mechanical loading.

    Directory of Open Access Journals (Sweden)

    Julián A García-Grajales

    Full Text Available With the growing body of research on traumatic brain injury and spinal cord injury, computational neuroscience has recently focused its modeling efforts on neuronal functional deficits following mechanical loading. However, in most of these efforts, cell damage is generally only characterized by purely mechanistic criteria, functions of quantities such as stress, strain or their corresponding rates. The modeling of functional deficits in neurites as a consequence of macroscopic mechanical insults has been rarely explored. In particular, a quantitative mechanically based model of electrophysiological impairment in neuronal cells, Neurite, has only very recently been proposed. In this paper, we present the implementation details of this model: a finite difference parallel program for simulating electrical signal propagation along neurites under mechanical loading. Following the application of a macroscopic strain at a given strain rate produced by a mechanical insult, Neurite is able to simulate the resulting neuronal electrical signal propagation, and thus the corresponding functional deficits. The simulation of the coupled mechanical and electrophysiological behaviors requires computational expensive calculations that increase in complexity as the network of the simulated cells grows. The solvers implemented in Neurite--explicit and implicit--were therefore parallelized using graphics processing units in order to reduce the burden of the simulation costs of large scale scenarios. Cable Theory and Hodgkin-Huxley models were implemented to account for the electrophysiological passive and active regions of a neurite, respectively, whereas a coupled mechanical model accounting for the neurite mechanical behavior within its surrounding medium was adopted as a link between electrophysiology and mechanics. This paper provides the details of the parallel implementation of Neurite, along with three different application examples: a long myelinated axon

  10. 3D-Printed pHEMA Materials for Topographical and Biochemical Modulation of Dorsal Root Ganglion Cell Response.

    Science.gov (United States)

    Badea, Adina; McCracken, Joselle M; Tillmaand, Emily G; Kandel, Mikhail E; Oraham, Aaron W; Mevis, Molly B; Rubakhin, Stanislav S; Popescu, Gabriel; Sweedler, Jonathan V; Nuzzo, Ralph G

    2017-09-13

    Understanding and controlling the interactions occurring between cells and engineered materials are central challenges toward progress in the development of biomedical devices. In this work, we describe materials for direct ink writing (DIW), an extrusion-based type of 3D printing, that embed a custom synthetic protein (RGD-PDL) within the microfilaments of 3D-hydrogel scaffolds to modify these interactions and differentially direct tissue-level organization of complex cell populations in vitro. The RGD-PDL is synthesized by modifying poly-d-lysine (PDL) to varying extents with peptides containing the integrin-binding motif Arg-Gly-Asp (RGD). Compositional gradients of the RGD-PDL presented by both patterned and thin-film poly(2-hydroxyethyl) methacrylate (pHEMA) substrates allow the patterning of cell-growth compliance in a grayscale form. The surface chemistry-dependent guidance of cell growth on the RGD-PDL-modified pHEMA materials is demonstrated using a model NIH-3T3 fibroblast cell line. The formation of a more complex cellular system-organotypic primary murine dorsal root ganglion (DRG)-in culture is also achieved on these scaffolds, where distinctive forms of cell growth and migration guidance are seen depending on their RGD-PDL content and topography. This experimental platform for the study of physicochemical factors on the formation and the reorganization of organotypic cultures offers useful capabilities for studies in tissue engineering, regenerative medicine, and diagnostics.

  11. INFLUENCE OF DIFFERENT INCUBATOR MODELS ON MAGNETIC FIELD-INDUCED CHANGES IN NEURITE OUTGROWTH IN PC-12 CELLS

    Science.gov (United States)

    OBJECTIVE: Devise a method to standardize responses of cells to MF-exposure in different incubator environments. METHODS: We compared the cell responses to generated MF in a standard cell-culture incubator (Forma, model #3158) with cell responses to the same exposure when a mu-m...

  12. RabGEF1/Rabex-5 Regulates TrkA-Mediated Neurite Outgrowth and NMDA-Induced Signaling Activation in NGF-Differentiated PC12 Cells

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    Tam, See-Ying; Lilla, Jennifer N.; Chen, Ching-Cheng; Kalesnikoff, Janet; Tsai, Mindy

    2015-01-01

    Nerve growth factor (NGF) binds to its cognate receptor TrkA and induces neuronal differentiation by activating distinct downstream signal transduction events. RabGEF1 (also known as Rabex-5) is a guanine nucleotide exchange factor for Rab5, which regulates early endosome fusion and vesicular trafficking in endocytic pathways. Here, we used the antisense (AS) expression approach to induce an NGF-dependent sustained knockdown of RabGEF1 protein expression in stable PC12 transfectants. We show that RabGEF1 is a negative regulator of NGF-induced neurite outgrowth and modulates other cellular and signaling processes that are activated by the interaction of NGF with TrkA receptors, such as cell cycle progression, cessation of proliferation, and activation of NGF-mediated downstream signaling responses. Moreover, RabGEF1 can bind to Rac1, and the activation of Rac1 upon NGF treatment is significantly enhanced in AS transfectants, suggesting that RabGEF1 is a negative regulator of NGF-induced Rac1 activation in PC12 cells. Furthermore, we show that RabGEF1 can also interact with NMDA receptors by binding to the NR2B subunit and its associated binding partner SynGAP, and negatively regulates activation of nitric oxide synthase activity induced by NMDA receptor stimulation in NGF-differentiated PC12 cells. Our data suggest that RabGEF1 is a negative regulator of TrkA-dependent neuronal differentiation and of NMDA receptor-mediated signaling activation in NGF-differentiated PC12 cells. PMID:26588713

  13. Interferon-gamma (IFN-γ-mediated retinal ganglion cell death in human tyrosinase T cell receptor transgenic mouse.

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    Shahid Husain

    Full Text Available We have recently demonstrated the characterization of human tyrosinase TCR bearing h3T-A2 transgenic mouse model, which exhibits spontaneous autoimmune vitiligo and retinal dysfunction. The purpose of current study was to determine the role of T cells and IFN-γ in retina dysfunction and retinal ganglion cell (RGC death using this model. RGC function was measured by pattern electroretinograms (ERGs in response to contrast reversal of patterned visual stimuli. RGCs were visualized by fluorogold retrograde-labeling. Expression of CD3, IFN-γ, GFAP, and caspases was measured by immunohistochemistry and Western blotting. All functional and structural changes were measured in 12-month-old h3T-A2 mice and compared with age-matched HLA-A2 wild-type mice. Both pattern-ERGs (42%, p = 0.03 and RGC numbers (37%, p = 0.0001 were reduced in h3T-A2 mice when compared with wild-type mice. The level of CD3 expression was increased in h3T-A2 mice (h3T-A2: 174 ± 27% vs. HLA-A2: 100%; p = 0.04. The levels of effector cytokine IFN-γ were also increased significantly in h3T-A2 mice (h3T-A2: 189 ± 11% vs. HLA-A2: 100%; p = 0.023. Both CD3 and IFN-γ immunostaining were increased in nerve fiber (NF and RGC layers of h3T-A2 mice. In addition, we have seen a robust increase in GFAP staining in h3T-A2 mice (mainly localized to NF layer, which was substantially reduced in IFN-γ ((-/- knockout h3T-A2 mice. We also have seen an up-regulation of caspase-3 and -9 in h3T-A2 mice. Based on our data we conclude that h3T-A2 transgenic mice exhibit visual defects that are mostly associated with the inner retinal layers and RGC function. This novel h3T-A2 transgenic mouse model provides opportunity to understand RGC pathology and test neuroprotective strategies to rescue RGCs.

  14. Neuroprotection of a novel cyclopeptide C*HSDGIC* from the cyclization of PACAP (1-5 in cellular and rodent models of retinal ganglion cell apoptosis.

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    Huanhuan Cheng

    Full Text Available To investigate the protective effects of a novel cyclopeptide C*HSDGIC* (CHC from the cyclization of Pituitary adenylate cyclase-activating polypeptide (PACAP (1-5 in cellular and rodent models of retinal ganglion cell apoptosis.Double-labeling immunohistochemistry was used to detect the expression of Thy-1 and PACAP receptor type 1 in a retinal ganglion cell line RGC-5. The apoptosis of RGC-5 cells was induced by 0.02 J/cm(2 Ultraviolet B irradiation. MTT assay, flow cytometry, fluorescence microscopy were used to investigate the viability, the level of reactive oxygen species (ROS and apoptosis of RGC-5 cells respectively. CHC attenuated apoptotic cell death induced by Ultraviolet B irradiation and inhibited the excessive generation of ROS. Moreover, CHC treatment resulted in decreased expression of Bax and concomitant increase of Bcl-2, as was revealed by western-blot analysis. The in vivo apoptosis of retinal ganglion cells was induced by injecting 50 mM N-methyl-D-aspartate (NMDA (100 nmol in a 2 µL saline solution intravitreally, and different dosages of CHC were administered. At day 7, rats in CHC+ NMDA-treated groups showed obvious aversion to light when compared to NMDA rats. Electroretinogram recordings revealed a marked decrease in the amplitudes of a-wave, b-wave, and photopic negative response due to NMDA damage. In retina receiving intravitreal NMDA and CHC co-treatment, these values were significantly increased. CHC treatment also resulted in less NMDA-induced cell loss and a decrease in the proportion of dUTP end-labeling-positive cells in ganglion cell line.C*HSDGIC*, a novel cyclopeptide from PACAP (1-5 attenuates apoptosis in RGC-5 cells and inhibits NMDA-induced retinal neuronal death. The beneficial effects may occur via the mitochondria pathway. PACAP derivatives like CHC may serve as a promising candidate for neuroprotection in glaucoma.

  15. Network oscillations drive correlated spiking of ON and OFF ganglion cells in the rd1 mouse model of retinal degeneration.

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    David J Margolis

    Full Text Available Following photoreceptor degeneration, ON and OFF retinal ganglion cells (RGCs in the rd-1/rd-1 mouse receive rhythmic synaptic input that elicits bursts of action potentials at ∼ 10 Hz. To characterize the properties of this activity, RGCs were targeted for paired recording and morphological classification as either ON alpha, OFF alpha or non-alpha RGCs using two-photon imaging. Identified cell types exhibited rhythmic spike activity. Cross-correlation of spike trains recorded simultaneously from pairs of RGCs revealed that activity was correlated more strongly between alpha RGCs than between alpha and non-alpha cell pairs. Bursts of action potentials in alpha RGC pairs of the same type, i.e. two ON or two OFF cells, were in phase, while bursts in dissimilar alpha cell types, i.e. an ON and an OFF RGC, were 180 degrees out of phase. This result is consistent with RGC activity being driven by an input that provides correlated excitation to ON cells and inhibition to OFF cells. A2 amacrine cells were investigated as a candidate cellular mechanism and found to display 10 Hz oscillations in membrane voltage and current that persisted in the presence of antagonists of fast synaptic transmission and were eliminated by tetrodotoxin. Results support the conclusion that the rhythmic RGC activity originates in a presynaptic network of electrically coupled cells including A2s via a Na(+-channel dependent mechanism. Network activity drives out of phase oscillations in ON and OFF cone bipolar cells, entraining similar frequency fluctuations in RGC spike activity over an area of retina that migrates with changes in the spatial locus of the cellular oscillator.

  16. Network oscillations drive correlated spiking of ON and OFF ganglion cells in the rd1 mouse model of retinal degeneration.

    Science.gov (United States)

    Margolis, David J; Gartland, Andrew J; Singer, Joshua H; Detwiler, Peter B

    2014-01-01

    Following photoreceptor degeneration, ON and OFF retinal ganglion cells (RGCs) in the rd-1/rd-1 mouse receive rhythmic synaptic input that elicits bursts of action potentials at ∼ 10 Hz. To characterize the properties of this activity, RGCs were targeted for paired recording and morphological classification as either ON alpha, OFF alpha or non-alpha RGCs using two-photon imaging. Identified cell types exhibited rhythmic spike activity. Cross-correlation of spike trains recorded simultaneously from pairs of RGCs revealed that activity was correlated more strongly between alpha RGCs than between alpha and non-alpha cell pairs. Bursts of action potentials in alpha RGC pairs of the same type, i.e. two ON or two OFF cells, were in phase, while bursts in dissimilar alpha cell types, i.e. an ON and an OFF RGC, were 180 degrees out of phase. This result is consistent with RGC activity being driven by an input that provides correlated excitation to ON cells and inhibition to OFF cells. A2 amacrine cells were investigated as a candidate cellular mechanism and found to display 10 Hz oscillations in membrane voltage and current that persisted in the presence of antagonists of fast synaptic transmission and were eliminated by tetrodotoxin. Results support the conclusion that the rhythmic RGC activity originates in a presynaptic network of electrically coupled cells including A2s via a Na(+)-channel dependent mechanism. Network activity drives out of phase oscillations in ON and OFF cone bipolar cells, entraining similar frequency fluctuations in RGC spike activity over an area of retina that migrates with changes in the spatial locus of the cellular oscillator.

  17. Electric field-induced astrocyte alignment directs neurite outgrowth.

    Science.gov (United States)

    Alexander, John K; Fuss, Babette; Colello, Raymond J

    2006-05-01

    The extension and directionality of neurite outgrowth are key to achieving successful target connections during both CNS development and during the re-establishment of connections lost after neural trauma. The degree of axonal elongation depends, in large part, on the spatial arrangement of astrocytic processes rich in growth-promoting proteins. Because astrocytes in culture align their processes on exposure to an electrical field of physiological strength, we sought to determine the extent to which aligned astrocytes affect neurite outgrowth. To this end, dorsal root ganglia cells were seeded onto cultured rat astrocytes that were pre-aligned by exposure to an electric field of physiological strength (500 mV mm(-1)). Using confocal microscopy and digital image analysis, we found that neurite outgrowth at 24 hours and at 48 hours is enhanced significantly and directed consistently along the aligned astrocyte processes. Moreover, this directed neurite outgrowth is maintained when grown on fixed, aligned astrocytes. Collectively, these results indicate that endogenous electric fields present within the developing CNS might act to align astrocyte processes, which can promote and direct neurite growth. Furthermore, these results demonstrate a simple method to produce an aligned cellular substrate, which might be used to direct regenerating neurites.

  18. Isolation and Molecular Profiling of Primary Mouse Retinal Ganglion Cells: Comparison of Phenotypes from Healthy and Glaucomatous Retinas.

    Science.gov (United States)

    Chintalapudi, Sumana R; Djenderedjian, Levon; Stiemke, Andrew B; Steinle, Jena J; Jablonski, Monica M; Morales-Tirado, Vanessa M

    2016-01-01

    Loss of functional retinal ganglion cells (RGC) is an element of retinal degeneration that is poorly understood. This is in part due to the lack of a reliable and validated protocol for the isolation of primary RGCs. Here we optimize a feasible, reproducible, standardized flow cytometry-based protocol for the isolation and enrichment of homogeneous RGC with the Thy1.2(hi)CD48(neg)CD15(neg)CD57(neg) surface phenotype. A three-step validation process was performed by: (1) genomic profiling of 25-genes associated with retinal cells; (2) intracellular labeling of homogeneous sorted cells for the intracellular RGC-markers SNCG, brain-specific homeobox/POU domain protein 3A (BRN3A), TUJ1, and RNA-binding protein with multiple splicing (RBPMS); and (3) by applying the methodology on RGC from a mouse model with elevated intraocular pressure (IOP) and optic nerve damage. Use of primary RGC cultures will allow for future careful assessment of important cell specific pathways in RGC to provide mechanistic insights into the declining of visual acuity in aged populations and those suffering from retinal neurodegenerative diseases.

  19. Isolation and Molecular Profiling of Primary Mouse Retinal Ganglion Cells: Comparison of Phenotypes from Healthy and Glaucomatous Retinas

    Science.gov (United States)

    Chintalapudi, Sumana R.; Djenderedjian, Levon; Stiemke, Andrew B.; Steinle, Jena J.; Jablonski, Monica M.; Morales-Tirado, Vanessa M.

    2016-01-01

    Loss of functional retinal ganglion cells (RGC) is an element of retinal degeneration that is poorly understood. This is in part due to the lack of a reliable and validated protocol for the isolation of primary RGCs. Here we optimize a feasible, reproducible, standardized flow cytometry-based protocol for the isolation and enrichment of homogeneous RGC with the Thy1.2hiCD48negCD15negCD57neg surface phenotype. A three-step validation process was performed by: (1) genomic profiling of 25-genes associated with retinal cells; (2) intracellular labeling of homogeneous sorted cells for the intracellular RGC-markers SNCG, brain-specific homeobox/POU domain protein 3A (BRN3A), TUJ1, and RNA-binding protein with multiple splicing (RBPMS); and (3) by applying the methodology on RGC from a mouse model with elevated intraocular pressure (IOP) and optic nerve damage. Use of primary RGC cultures will allow for future careful assessment of important cell specific pathways in RGC to provide mechanistic insights into the declining of visual acuity in aged populations and those suffering from retinal neurodegenerative diseases. PMID:27242509

  20. Quinuclidine compounds differently act as agonists of Kenyon cell nicotinic acetylcholine receptors and induced distinct effect on insect ganglionic depolarizations.

    Science.gov (United States)

    Mathé-Allainmat, Monique; Swale, Daniel; Leray, Xavier; Benzidane, Yassine; Lebreton, Jacques; Bloomquist, Jeffrey R; Thany, Steeve H

    2013-12-01

    We have recently demonstrated that a new quinuclidine benzamide compound named LMA10203 acted as an agonist of insect nicotinic acetylcholine receptors. Its specific pharmacological profile on cockroach dorsal unpaired median neurons (DUM) helped to identify alpha-bungarotoxin-insensitive nAChR2 receptors. In the present study, we tested its effect on cockroach Kenyon cells. We found that it induced an inward current demonstrating that it bounds to nicotinic acetylcholine receptors expressed on Kenyon cells. Interestingly, LMA10203-induced currents were completely blocked by the nicotinic antagonist α-bungarotoxin. We suggested that LMA10203 effect occurred through the activation of α-bungarotoxin-sensitive receptors and did not involve α-bungarotoxin-insensitive nAChR2, previously identified in DUM neurons. In addition, we have synthesized two new compounds, LMA10210 and LMA10211, and compared their effects on Kenyon cells. These compounds were members of the 3-quinuclidinyl benzamide or benzoate families. Interestingly, 1 mM LMA10210 was not able to induce an inward current on Kenyon cells compared to LMA10211. Similarly, we did not find any significant effect of LMA10210 on cockroach ganglionic depolarization, whereas these three compounds were able to induce an effect on the central nervous system of the third instar M. domestica larvae. Our data suggested that these three compounds could bind to distinct cockroach nicotinic acetylcholine receptors.

  1. Selective deletion of cochlear hair cells causes rapid age-dependent changes in spiral ganglion and cochlear nucleus neurons.

    Science.gov (United States)

    Tong, Ling; Strong, Melissa K; Kaur, Tejbeer; Juiz, Jose M; Oesterle, Elizabeth C; Hume, Clifford; Warchol, Mark E; Palmiter, Richard D; Rubel, Edwin W

    2015-05-20

    During nervous system development, critical periods are usually defined as early periods during which manipulations dramatically change neuronal structure or function, whereas the same manipulations in mature animals have little or no effect on the same property. Neurons in the ventral cochlear nucleus (CN) are dependent on excitatory afferent input for survival during a critical period of development. Cochlear removal in young mammals and birds results in rapid death of target neurons in the CN. Cochlear removal in older animals results in little or no neuron death. However, the extent to which hair-cell-specific afferent activity prevents neuronal death in the neonatal brain is unknown. We further explore this phenomenon using a new mouse model that allows temporal control of cochlear hair cell deletion. Hair cells express the human diphtheria toxin (DT) receptor behind the Pou4f3 promoter. Injections of DT resulted in nearly complete loss of organ of Corti hair cells within 1 week of injection regardless of the age of injection. Injection of DT did not influence surrounding supporting cells directly in the sensory epithelium or spiral ganglion neurons (SGNs). Loss of hair cells in neonates resulted in rapid and profound neuronal loss in the ventral CN, but not when hair cells were eliminated at a more mature age. In addition, normal survival of SGNs was dependent on hair cell integrity early in development and less so in mature animals. This defines a previously undocumented critical period for SGN survival. Copyright © 2015 the authors 0270-6474/15/357878-14$15.00/0.

  2. Action potentials in retinal ganglion cells are initiated at the site of maximal curvature of the extracellular potential.

    Science.gov (United States)

    Eickenscheidt, Max; Zeck, Günther

    2014-06-01

    The initiation of an action potential by extracellular stimulation occurs after local depolarization of the neuronal membrane above threshold. Although the technique shows remarkable clinical success, the site of action and the relevant stimulation parameters are not completely understood. Here we identify the site of action potential initiation in rabbit retinal ganglion cells (RGCs) interfaced to an array of extracellular capacitive stimulation electrodes. We determine which feature of the extracellular potential governs action potential initiation by simultaneous stimulation and recording RGCs interfaced in epiretinal configuration. Stimulation electrodes were combined to areas of different size and were presented at different positions with respect to the RGC. Based on stimulation by electrodes beneath the RGC soma and simultaneous sub-millisecond latency measurement we infer axonal initiation at the site of maximal curvature of the extracellular potential. Stimulation by electrodes at different positions along the axon reveals a nearly constant threshold current density except for a narrow region close to the cell soma. These findings are explained by the concept of the activating function modified to consider a region of lower excitability close to the cell soma. We present a framework how to estimate the site of action potential initiation and the stimulus required to cross threshold in neurons tightly interfaced to capacitive stimulation electrodes. Our results underscore the necessity of rigorous electrical characterization of the stimulation electrodes and of the interfaced neural tissue.

  3. Direct Reprogramming of Spiral Ganglion Non-neuronal Cells into Neurons: Toward Ameliorating Sensorineural Hearing Loss by Gene Therapy

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    Teppei Noda

    2018-02-01

    Full Text Available Primary auditory neurons (PANs play a critical role in hearing by transmitting sound information from the inner ear to the brain. Their progressive degeneration is associated with excessive noise, disease and aging. The loss of PANs leads to permanent hearing impairment since they are incapable of regenerating. Spiral ganglion non-neuronal cells (SGNNCs, comprised mainly of glia, are resident within the modiolus and continue to survive after PAN loss. These attributes make SGNNCs an excellent target for replacing damaged PANs through cellular reprogramming. We used the neurogenic pioneer transcription factor Ascl1 and the auditory neuron differentiation factor NeuroD1 to reprogram SGNNCs into induced neurons (iNs. The overexpression of both Ascl1 and NeuroD1 in vitro generated iNs at high efficiency. Transcriptome analyses revealed that iNs displayed a transcriptome profile resembling that of endogenous PANs, including expression of several key markers of neuronal identity: Tubb3, Map2, Prph, Snap25, and Prox1. Pathway analyses indicated that essential pathways in neuronal growth and maturation were activated in cells upon neuronal induction. Furthermore, iNs extended projections toward cochlear hair cells and cochlear nucleus neurons when cultured with each respective tissue. Taken together, our study demonstrates that PAN-like neurons can be generated from endogenous SGNNCs. This work suggests that gene therapy can be a viable strategy to treat sensorineural hearing loss caused by degeneration of PANs.

  4. Action potentials in retinal ganglion cells are initiated at the site of maximal curvature of the extracellular potential

    Science.gov (United States)

    Eickenscheidt, Max; Zeck, Günther

    2014-06-01

    Objective. The initiation of an action potential by extracellular stimulation occurs after local depolarization of the neuronal membrane above threshold. Although the technique shows remarkable clinical success, the site of action and the relevant stimulation parameters are not completely understood. Approach. Here we identify the site of action potential initiation in rabbit retinal ganglion cells (RGCs) interfaced to an array of extracellular capacitive stimulation electrodes. We determine which feature of the extracellular potential governs action potential initiation by simultaneous stimulation and recording RGCs interfaced in epiretinal configuration. Stimulation electrodes were combined to areas of different size and were presented at different positions with respect to the RGC. Main results. Based on stimulation by electrodes beneath the RGC soma and simultaneous sub-millisecond latency measurement we infer axonal initiation at the site of maximal curvature of the extracellular potential. Stimulation by electrodes at different positions along the axon reveals a nearly constant threshold current density except for a narrow region close to the cell soma. These findings are explained by the concept of the activating function modified to consider a region of lower excitability close to the cell soma. Significance. We present a framework how to estimate the site of action potential initiation and the stimulus required to cross threshold in neurons tightly interfaced to capacitive stimulation electrodes. Our results underscore the necessity of rigorous electrical characterization of the stimulation electrodes and of the interfaced neural tissue.

  5. N-Acetylcysteine protects inner ear hair cells and spiral ganglion neurons from manganese exposure by regulating ROS levels.

    Science.gov (United States)

    Wang, Weilong; Li, Dan; Ding, Xuerui; Zhao, Qianqian; Chen, Jiawei; Tian, Keyong; Qiu, Yang; Lu, Lianjun

    2017-09-05

    Manganese (Mn) is an indispensable cofactor for many enzymes and a basic factor for many reproductive and metabolic pathways. However, exposure to high concentrations of Mn can result in deleterious effects on the central nervous system and peripheral nerves, including nerves associated with the auditory system. Based on our studies of cochlear organotypic cultures, Mn exposure induces a significant loss of hair cells (HCs), auditory nerve fibers (ANFs) and spiral ganglion neurons (SGNs) in a concentration-dependent manner. Additionally, N-acetylcysteine (NAC), a glutathione (GSH) provider and a direct scavenger of reactive oxygen species (ROS), clearly decreases Mn-induced ROS accumulation, caspase-3 activation and TUNEL staining, which indicate increased cell survival. Based on these results, Mn exposure exerts ototoxic and neurotoxic effects on the auditory system. Furthermore, 20mM NAC may prevent 1mM Mn-induced hair cell loss and axonal degeneration, indicating that NAC could be a promising drug for clinical applications. Copyright © 2017 Elsevier B.V. All rights reserved.

  6. Direct Reprogramming of Spiral Ganglion Non-neuronal Cells into Neurons: Toward Ameliorating Sensorineural Hearing Loss by Gene Therapy.

    Science.gov (United States)

    Noda, Teppei; Meas, Steven J; Nogami, Jumpei; Amemiya, Yutaka; Uchi, Ryutaro; Ohkawa, Yasuyuki; Nishimura, Koji; Dabdoub, Alain

    2018-01-01

    Primary auditory neurons (PANs) play a critical role in hearing by transmitting sound information from the inner ear to the brain. Their progressive degeneration is associated with excessive noise, disease and aging. The loss of PANs leads to permanent hearing impairment since they are incapable of regenerating. Spiral ganglion non-neuronal cells (SGNNCs), comprised mainly of glia, are resident within the modiolus and continue to survive after PAN loss. These attributes make SGNNCs an excellent target for replacing damaged PANs through cellular reprogramming. We used the neurogenic pioneer transcription factor Ascl1 and the auditory neuron differentiation factor NeuroD1 to reprogram SGNNCs into induced neurons (iNs). The overexpression of both Ascl1 and NeuroD1 in vitro generated iNs at high efficiency. Transcriptome analyses revealed that iNs displayed a transcriptome profile resembling that of endogenous PANs, including expression of several key markers of neuronal identity: Tubb3, Map2, Prph, Snap25, and Prox1. Pathway analyses indicated that essential pathways in neuronal growth and maturation were activated in cells upon neuronal induction. Furthermore, iNs extended projections toward cochlear hair cells and cochlear nucleus neurons when cultured with each respective tissue. Taken together, our study demonstrates that PAN-like neurons can be generated from endogenous SGNNCs. This work suggests that gene therapy can be a viable strategy to treat sensorineural hearing loss caused by degeneration of PANs.

  7. Two transcription factors, Pou4f2 and Isl1, are sufficient to specify the retinal ganglion cell fate.

    Science.gov (United States)

    Wu, Fuguo; Kaczynski, Tadeusz J; Sethuramanujam, Santhosh; Li, Renzhong; Jain, Varsha; Slaughter, Malcolm; Mu, Xiuqian

    2015-03-31

    As with other retinal cell types, retinal ganglion cells (RGCs) arise from multipotent retinal progenitor cells (RPCs), and their formation is regulated by a hierarchical gene-regulatory network (GRN). Within this GRN, three transcription factors--atonal homolog 7 (Atoh7), POU domain, class 4, transcription factor 2 (Pou4f2), and insulin gene enhancer protein 1 (Isl1)--occupy key node positions at two different stages of RGC development. Atoh7 is upstream and is required for RPCs to gain competence for an RGC fate, whereas Pou4f2 and Isl1 are downstream and regulate RGC differentiation. However, the genetic and molecular basis for the specification of the RGC fate, a key step in RGC development, remains unclear. Here we report that ectopic expression of Pou4f2 and Isl1 in the Atoh7-null retina using a binary knockin-transgenic system is sufficient for the specification of the RGC fate. The RGCs thus formed are largely normal in gene expression, survive to postnatal stages, and are physiologically functional. Our results indicate that Pou4f2 and Isl1 compose a minimally sufficient regulatory core for the RGC fate. We further conclude that during development a core group of limited transcription factors, including Pou4f2 and Isl1, function downstream of Atoh7 to determine the RGC fate and initiate RGC differentiation.

  8. High-resolution X-ray tomography of the human inner ear: synchrotron radiation-based study of nerve fibre bundles, membranes and ganglion cells.

    Science.gov (United States)

    Lareida, A; Beckmann, F; Schrott-Fischer, A; Glueckert, R; Freysinger, W; Müller, B

    2009-04-01

    The combination of osmium tetroxide staining and high-resolution tomographic imaging using monochromatic X rays allows visualizing cellular structures of the human inner ear, that is, the organ of Corti, the stria vascularis and further soft tissues of the membranous labyrinth, in three-dimensional space with isotropic micrometre resolution. This approach permits to follow the course of nerve fibre bundles in a major part of the specimen and reveals the detailed three-dimensional arrangement of individual ganglion cells with distinct nuclei by means of X-ray tomography for the first time. The non-destructive neuron cell counting in a selected volume of 125 microm x 800 microm x 600 microm = 0.06 mm(3) gives rise to the estimate that 2000 ganglion cells are present along 1 mm organ of Corti.

  9. Investigating the expression of metabotropic glutamate receptors in trigeminal ganglion neurons and satellite glial cells: implications for craniofacial pain.

    Science.gov (United States)

    Boye Larsen, Dennis; Ingemann Kristensen, Gunda; Panchalingam, Vinodenee; Laursen, Jens Christian; Nørgaard Poulsen, Jeppe; Skallerup Andersen, Maria; Kandiah, Aginsha; Gazerani, Parisa

    2014-08-01

    Previous studies have demonstrated that various subtypes of the metabotropic glutamate receptors (mGluRs) are expressed in the dorsal root ganglion (DRG) of the peripheral nervous system (PNS), implicating that glutamate potentially contributes to sensory transmission through these receptors. While mGluR expression has been investigated largely in the DRG, the present study focused on mGluR expression on neurons and satellite glial cells (SGCs) of the trigeminal ganglion (TG). To address the presence of mGluRs in rat TG neurons and their corresponding SGCs, the trigeminal ganglia from six adult male Wistar rats were isolated and immunohistochemistry and immunocytochemistry were performed. The expression of mGluR1α-, mGluR2/3- and mGluR8 on TG neurons and SGCs was investigated in tissue slices and isolated cells. 35.1 ± 6.0% of the TG neurons were positive for mGluR1α, whereas 39.9 ± 7.7% and 55.5 ± 6.3% were positive for mGluR2/3 and mGluR8, respectively. Immunoreactive neurons expressing mGluRs were mainly medium- to large sized, with a smaller population of small-sized neurons showing immunoreactivity. The SGCs showed immunoreactivity toward mGluR1α and mGluR8, but not mGluR2/3, both in the tissue and in isolated cells. Findings from the present study showed that trigeminal neurons express mGluR1α, mGluR2/3 and mGluR8, while SGCs only express mGluR1α and mGluR8. This novel evidence may advance investigations on a possible role of mGluRs in relation to trigeminal pain transmission within the craniofacial region.

  10. Activation of neurokinin-1 receptors increases the excitability of guinea pig dorsal root ganglion cells.

    Science.gov (United States)

    Zhang, Xiulin; Pietra, Claudio; Lovati, Emanuela; de Groat, William C

    2012-10-01

    The suppression of overactive bladder symptoms in patients and overactive bladder reflexes in animal models by neurokinin (NK)-1 receptor antagonists raises the possibility that these drugs target sensory neurons. This mechanism was evaluated by examining the interactions between a specific NK-1 agonist, [Sar(9),Met(O(2))(11)]-substance P (Sar-Met-SP), and a potent NK-1 antagonist, netupitant (NTP), on small size (20-30 μm) dissociated L6 and S1 dorsal root ganglion (DRG) neurons from female guinea pigs. Current-clamp recording revealed that Sar-Met-SP (1 μM) elicited membrane depolarization (average 8.05 ± 1.38 mV) in 27% (18 of 65) of DRG neurons. In 74% of the remaining neurons (35 of 47) Sar-Met-SP decreased the rheobase for action potential (AP) generation and increased the response to a suprathreshold stimulus (3 times rheobase) without changing the membrane potential. Sar-Met-SP also induced changes in the action potential (AP) wave form, including 1) an increase in overshoot (average 5 mV, n = 35 neurons), 2) a prolongation of AP duration (from 4.64 to 5.29 ms, n = 34), and 3) a reduction in the maximal rate of AP repolarization. NTP (200 nM) reversed the Sar-Met-SP-induced changes. Ca(2+) imaging showed that application of Sar-Met-SP (1 μM) decreased the tachyphylaxis induced by repeated application of capsaicin (0.5 μM), an effect blocked by pretreatment with NTP (200 nM). These results raise the possibility that activation of NK-1 receptors in primary sensory neurons plays a role in the generation of overactive bladder and that block of NK-1 receptors in these neurons may contribute to efficacy of NK-1 antagonists in the treatment of overactive bladder symptoms.

  11. Characterization of spiral ganglion neurons cultured on silicon micro-pillar substrates for new auditory neuro-electronic interfaces

    Science.gov (United States)

    Mattotti, M.; Micholt, L.; Braeken, D.; Kovačić, D.

    2015-04-01

    Objective. One of the strategies to improve cochlear implant technology is to increase the number of electrodes in the neuro-electronic interface. The objective was to characterize in vitro cultures of spiral ganglion neurons (SGN) cultured on surfaces of novel silicon micro-pillar substrates (MPS). Approach. SGN from P5 rat pups were cultured on MPS with different micro-pillar widths (1-5.6 μm) and spacings (0.6-15 μm) and were compared with control SGN cultures on glass coverslips by immunocytochemistry and scanning electron microscopy (SEM). Main results. Overall, MPS support SGN growth equally well as the control glass surfaces. Micro-pillars of a particular size-range (1.2-2.4 μm) were optimal in promoting SGN presence, neurite growth and alignment. On this specific micro-pillar size, more SGN were present, and neurites were longer and more aligned. SEM pictures highlight how cells on micro-pillars with smaller spacings grow directly on top of pillars, while at wider spacings (from 3.2 to 15 μm) they grow on the bottom of the surface, losing contact guidance. Further, we found that MPS encourage more monopolar and bipolar SGN morphologies compared to the control condition. Finally, MPS induce longest neurite growth with minimal interaction of S100+ glial cells. Significance. These results indicate that silicon micro-pillar substrates create a permissive environment for the growth of primary auditory neurons promoting neurite sprouting and are a promising technology for future high-density three-dimensional CMOS-based auditory neuro-electronic interfaces.

  12. Quercetin Declines Apoptosis, Ameliorates Mitochondrial Function and Improves Retinal Ganglion Cell Survival and Function in In Vivo Model of Glaucoma in Rat and Retinal Ganglion Cell Culture In Vitro

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    Feng-Juan Gao

    2017-09-01

    Full Text Available Glaucoma is a progressive neuropathy characterized by the loss of retinal ganglion cells (RGCs. Strategies that delay or halt RGC loss have been recognized as potentially beneficial for rescuing vision in glaucoma patients. Quercetin (Qcn is a natural and important dietary flavonoid compound, widely distributed in fruits and vegetables. Mounting evidence suggests that Qcn has numerous neuroprotective effects. However, whether Qcn exerts neuroprotective effects on RGC in glaucoma is poorly understood. In this study, we investigated the protective effect of Qcn against RGC damage in a rat chronic ocular hypertension (COHT model invivo and hypoxia-induced primary cultured RGC damage in vitro, and we further explored the underlying neuroprotective mechanisms. We found that Qcn not only improved RGC survival and function from a very early stage of COHT invivo, it promoted the survival of hypoxia-treated primary cultured RGCs invitro via ameliorating mitochondrial function and preventing mitochondria-mediated apoptosis. Our findings suggest that Qcn has direct protective effects on RGCs that are independent of lowering the intraocular pressure (IOP. Qcn may be a promising therapeutic agent for improving RGC survival and function in glaucomatous neurodegeneration.

  13. Cross-talk between androgen receptor/filamin A and TrkA regulates neurite outgrowth in PC12 cells

    Science.gov (United States)

    Di Donato, Marzia; Bilancio, Antonio; D'Amato, Loredana; Claudiani, Pamela; Oliviero, Maria Antonietta; Barone, Maria Vittoria; Auricchio, Alberto; Appella, Ettore; Migliaccio, Antimo; Auricchio, Ferdinando; Castoria, Gabriella

    2015-01-01

    Steroids and growth factors control neuronal development through their receptors under physiological and pathological conditions. We show that PC12 cells harbor endogenous androgen receptor (AR), whose inhibition or silencing strongly interferes with neuritogenesis stimulated by the nonaromatizable synthetic androgen R1881 or NGF. This implies a role for AR not only in androgen signaling, but also in NGF signaling. In turn, a pharmacological TrkA inhibitor interferes with NGF- or androgen-induced neuritogenesis. In addition, androgen or NGF triggers AR association with TrkA, TrkA interaction with PI3-K δ, and downstream activation of PI3-K δ and Rac in PC12 cells. Once associated with AR, filamin A (FlnA) contributes to androgen or NGF neuritogenesis, likely through its interaction with signaling effectors, such as Rac. This study thus identifies a previously unrecognized reciprocal cross-talk between AR and TrkA, which is controlled by β1 integrin. The contribution of FlnA/AR complex and PI3-K δ to neuronal differentiation by androgens and NGF is also novel. This is the first description of AR function in PC12 cells. PMID:26063730

  14. Electrospun silk fibroin scaffolds coated with reduced graphene promote neurite outgrowth of PC-12 cells under electrical stimulation.

    Science.gov (United States)

    Aznar-Cervantes, Salvador; Pagán, Ana; Martínez, Jose G; Bernabeu-Esclapez, Antonia; Otero, Toribio F; Meseguer-Olmo, Luis; Paredes, Juan I; Cenis, Jose L

    2017-10-01

    Novel approaches to neural research require biocompatible materials capable to act as electrode structures or scaffolds for tissue engineering in order to stimulate or restore the functionality of damaged tissues. This work offers promising results that indicate the potential use of electrospun silk fibroin (SF) scaffolds coated with reduced graphene oxide (rGO) in this sense. The coated material becomes conductor and electroactive. A complete characterisation of SF/rGO scaffolds is provided in terms of electrochemistry, mechanical behaviour and chemical conformation of fibroin. The excellent biocompatibility of this novel material is proved with cultures of PC-12 cells. The coating with rGO improved the adhesion of cells in comparison with cells growing onto the surface of pure SF scaffolds. Also, the use of SF/rGO scaffolds combined with electrical stimulation promoted the differentiation into neural phenotypes reaching comparable or even superior levels to those obtained by means of the traditional treatment with neural growth factor (NGF). Copyright © 2017 Elsevier B.V. All rights reserved.

  15. Spatially restricted electrical activation of retinal ganglion cells in the rabbit retina by hexapolar electrode return configuration

    Science.gov (United States)

    Habib, Amgad G.; Cameron, Morven A.; Suaning, Gregg J.; Lovell, Nigel H.; Morley, John W.

    2013-06-01

    Objective. Visual prostheses currently in development aim to restore some form of vision to patients suffering from diseases such as age-related macular degeneration and retinitis pigmentosa. Most rely on electrically stimulating inner retinal cells via electrodes implanted on or near the retina, resulting in percepts of light termed ‘phosphenes’. Activation of spatially distinct populations of cells in the retina is key for pattern vision to be produced. To achieve this, the electrical stimulation must be localized, activating cells only in the direct vicinity of the stimulating electrode(s). With this goal in mind, a hexagonal return (hexapolar) configuration has been proposed as an alternative to the traditional monopolar or bipolar return configurations for electrically stimulating the retina. This study investigated the efficacy of the hexapolar configuration in localizing the activation of retinal ganglion cells (RGCs), compared to a monopolar configuration. Approach. Patch-clamp electrophysiology was used to measure the activation thresholds of RGCs in whole-mount rabbit retina to monopolar and hexapolar electrical stimulation, applied subretinally. Main results. Hexapolar activation thresholds for RGCs located outside the hex guard were found to be significantly (>2 fold) higher than those located inside the area of tissue bounded by the hex guard. The hexapolar configuration localized the activation of RGCs more effectively than its monopolar counterpart. Furthermore, no difference in hexapolar thresholds or localization was observed when using cathodic-first versus anodic-first stimulation. Significance. The hexapolar configuration may provide an improved method for electrically stimulating spatially distinct populations of cells in retinal tissue.

  16. Development of a cell-based treatment for long-term neurotrophin expression and spiral ganglion neuron survival.

    Science.gov (United States)

    Zanin, M P; Hellström, M; Shepherd, R K; Harvey, A R; Gillespie, L N

    2014-09-26

    Spiral ganglion neurons (SGNs), the target cells of the cochlear implant, undergo gradual degeneration following loss of the sensory epithelium in deafness. The preservation of a viable population of SGNs in deafness can be achieved in animal models with exogenous application of neurotrophins such as brain-derived neurotrophic factor (BDNF) and neurotrophin-3. For translation into clinical application, a suitable delivery strategy that provides ongoing neurotrophic support and promotes long-term SGN survival is required. Cell-based neurotrophin treatment has the potential to meet the specific requirements for clinical application, and we have previously reported that Schwann cells genetically modified to express BDNF can support SGN survival in deafness for 4 weeks. This study aimed to investigate various parameters important for the development of a long-term cell-based neurotrophin treatment to support SGN survival. Specifically, we investigated different (i) cell types, (ii) gene transfer methods and (iii) neurotrophins, in order to determine which variables may provide long-term neurotrophin expression and which, therefore, may be the most effective for supporting long-term SGN survival in vivo. We found that fibroblasts that were nucleofected to express BDNF provided the most sustained neurotrophin expression, with ongoing BDNF expression for at least 30 weeks. In addition, the secreted neurotrophin was biologically active and elicited survival effects on SGNs in vitro. Nucleofected fibroblasts may therefore represent a method for safe, long-term delivery of neurotrophins to the deafened cochlea to support SGN survival in deafness. Copyright © 2014 IBRO. Published by Elsevier Ltd. All rights reserved.

  17. Quantitative and Topographical Analysis of the Losses of Cone Photoreceptors and Retinal Ganglion Cells Under Taurine Depletion.

    Science.gov (United States)

    Hadj-Saïd, Wahiba; Froger, Nicolas; Ivkovic, Ivana; Jiménez-López, Manuel; Dubus, Élisabeth; Dégardin-Chicaud, Julie; Simonutti, Manuel; Quénol, César; Neveux, Nathalie; Villegas-Pérez, María Paz; Agudo-Barriuso, Marta; Vidal-Sanz, Manuel; Sahel, Jose-Alain; Picaud, Serge; García-Ayuso, Diego

    2016-09-01

    Taurine depletion is known to induce photoreceptor degeneration and was recently found to also trigger retinal ganglion cell (RGC) loss similar to the retinal toxicity of vigabatrin. Our objective was to study the topographical loss of RGCs and cone photoreceptors, with a distinction between the two cone types (S- and L- cones) in an animal model of induced taurine depletion. We used the taurine transporter (Tau-T) inhibitor, guanidoethane sulfonate (GES), to induce taurine depletion at a concentration of 1% in the drinking water. Spectral-domain optical coherence tomography (SD-OCT) and electroretinograms (ERG) were performed on animals after 2 months of GES treatment administered through the drinking water. Retinas were dissected as wholemounts and immunodetection of Brn3a (RGC), S-opsin (S-cones), and L-opsin (L-cones) was performed. The number of Brn3a+ RGCs, and L- and S-opsin+ cones was automatically quantified and their retinal distribution studied using isodensity maps. The treatment resulted in a significant reduction in plasma taurine levels and a profound dysfunction of visual performance as shown by ERG recordings. Optical coherence tomography analysis revealed that the retina was thinner in the taurine-depleted group. S-opsin+cones were more affected (36%) than L-opsin+cones (27%) with greater cone cell loss in the dorsal area whereas RGC loss (12%) was uniformly distributed. This study confirms that taurine depletion causes RGC and cone loss. Electroretinograms results show that taurine depletion induces retinal dysfunction in photoreceptors and in the inner retina. It establishes a gradient of cell loss depending on the cell type from S-opsin+cones, L-opsin+cones, to RGCs. The greater cell loss in the dorsal retina and of the S-cone population may underline different cellular mechanisms of cellular degeneration and suggests that S-cones may be more sensitive to light-induced retinal toxicity enhanced by the taurine depletion.

  18. Temporal properties of network-mediated responses to repetitive stimuli are dependent upon retinal ganglion cell type

    Science.gov (United States)

    Im, Maesoon; Fried, Shelley I.

    2016-04-01

    Objective. To provide artificially-elicited vision that is temporally dynamic, retinal prosthetic devices will need to repeatedly stimulate retinal neurons. However, given the diversity of physiological types of retinal ganglion cells (RGCs) as well as the heterogeneity of their responses to electric stimulation, temporal properties of RGC responses have not been adequately investigated. Here, we explored the cell type dependence of network-mediated RGC responses to repetitive electric stimulation at various stimulation rates. Approach. We examined responses of ON and OFF types of RGCs in the rabbit retinal explant to five consecutive stimuli with varying inter-stimulus intervals (10-1000 ms). Each stimulus was a 4 ms long monophasic sinusoidal cathodal current, which was applied epiretinally via a conical electrode. Spiking activity of targeted RGCs was recorded using a cell-attached patch electrode. Main results. ON and OFF cells had distinct responses to repetitive stimuli. Consistent with earlier studies, OFF cells always generated reduced responses to subsequent stimuli compared to responses to the first stimulus. In contrast, a new stimulus to ON cells suppressed all pending/ongoing responses from previous stimuli and initiated its own response that was remarkably similar to the response from a single stimulus in isolation. This previously unreported ‘reset’ behavior was observed exclusively and consistently in ON cells. These contrasts between ON and OFF cells created a range of stimulation rates (4-7 Hz) that maximized the ratio of the responses arising in ON versus OFF cells. Significance. Previous clinical testing reported that subjects perceive bright phosphenes (ON responses) and also prefer stimulation rates of 5-7 Hz. Our results suggest that responses of ON cells are weak at high rates of stimulation (> ˜7 Hz) due to the reset while responses of OFF cells are strong at low rates (cells more closely match physiological patterns (Im and Fried 2015

  19. One cell model establishment to inhibit CaMKIIγmRNA expression in the dorsal root ganglion neuron by RNA interfere.

    Science.gov (United States)

    Wen, Xianjie; Li, Xiaohong; Liang, Hua; Yang, Chenxiang; Zhong, Jiying; Wang, Hanbing; Liu, Hongzhen

    2017-09-01

    CaMKII γ in dorsal root ganglion neurons is closely related to the neuropathic pain, neuron injury induced by local anesthetics. To get great insight into the function of CaMKII γ in dorsal root ganglion neurons, we need one cell model to specially inhibit the CaMKII γ mRNA expression. The present study was aimed to establish one cell model to specially inhibit the CaMKII γ mRNA expression. We designed the CaMKII γ shRNA sequence and connected with pYr-1.1 plasmid. The ligation product of the CaMKII γ shRNA and pYr-1.1 plasmid was recombined with pAd/PL-DEST vector into pAD-CaMKIIγ-shRNA. adenovirus vector. pAD-CaMKIIγ-shRNA. adenovirus vector infected the dorsal root ganglion neuron to inhibit the CaMKII γ mRNA expression in vitro. The pAD-CaMKIIγ-shRNA adenovirus vector was verified to be correct by the digestion, sequence. And pAD-CaMKIIγ-shRNA. adenovirus vector can infect the DRG cells to inhibit the CaMKII γ mRNA or protein expression by the real-time polymerase chain reaction (PCR) or western blotting. Those results showed that we successfully constructed one adenovirus vector that can infect the dorsal root ganglion neuron to inhibit the CaMKII γ mRNA and protein expression. That will supply with one cell model for the CaMKII γ function study.

  20. Ganglion cell-inner plexiform layer and retinal nerve fibre layer changes within the macula in retinitis pigmentosa: a spectral domain optical coherence tomography study.

    Science.gov (United States)

    Yoon, Chang Ki; Yu, Hyeong Gon

    2018-03-01

    To investigate how macular ganglion cell-inner plexiform layer (GCIPL) and retinal nerve fibre layer (RNFL) thicknesses within the macula change with retinitis pigmentosa (RP) severity. Spectral domain optical coherence tomography (SD-OCT) was used to examine 177 patients with RP and 177 normal controls. An optical coherence tomography (OCT) line scan was used to grade RP severity. Retinitis pigmentosa (RP) was categorized as more advanced if there was no identifiable inner segment ellipsoid (ISe) band (NISE) and as less advanced if an ISe band could be identified and peripheral loss of ISe was apparent (IISE). Ganglion cell-inner plexiform layer (GCIPL) and RNFL thicknesses were manually measured on OCT images and analysed. Pearson's correlation analyses were used to examine correlations between GCIPL thickness, RNFL thickness, visual acuity (VA) and visual field extent in patients and controls. Ganglion cell-inner plexiform layer (GCIPL) was significantly thicker in IISE than in control eyes (p fibre layer (RNFL) was significantly thicker in eyes with IISE and NISE than in control eyes in both horizontal and vertical meridians (all p < 0.001). Ganglion cell-inner plexiform layer (GCIPL) thickness showed a weak positive correlation with vision, and RNFL thickness showed a weak negative correlation with vision and visual field extent. Based on these results, the inner retina, including the GCIPL and RNFL, maintains its gross integrity longer than the photoreceptor layer in RP. Additionally, thickening of the inner retina may have some functional implications in patients with RP. © 2017 Acta Ophthalmologica Scandinavica Foundation. Published by John Wiley & Sons Ltd.

  1. EFFECT OF INTRAVITREAL RANIBIZUMAB ON GANGLION CELL COMPLEX AND PERIPAPILLARY RETINAL NERVE FIBER LAYER IN NEOVASCULAR AGE-RELATED MACULAR DEGENERATION USING SPECTRAL DOMAIN OPTICAL COHERENCE TOMOGRAPHY.

    Science.gov (United States)

    Zucchiatti, Ilaria; Cicinelli, Maria V; Parodi, Maurizio Battaglia; Pierro, Luisa; Gagliardi, Marco; Accardo, Agostino; Bandello, Francesco

    2017-07-01

    To analyze the changes in ganglion cell complex and peripapillary retinal nerve fiber layer thickness, in central macular thickness and choroidal thickness on spectral domain optical coherence tomography in patients with neovascular age-related macular degeneration treated with intravitreal ranibizumab injections. All consecutive patients with untreated neovascular age-related macular degeneration received loading phase of three monthly intravitreal ranibizumab, followed by retreatments on a pro re nata protocol for 12 months. changes in ganglion cell complex and retinal nerve fiber layer at the end of follow-up. Secondary outcome: changes in best-corrected visual acuity, central macular thickness, and choroidal thickness at the end of follow-up. Choroidal thickness was measured at 500 μm, 1000 μm, and 1,500 μm intervals nasally, temporally, superiorly, and inferiorly to the fovea, respectively, on horizontal and vertical line scans centered on the fovea. Twenty-four eyes were included. Ganglion cell complex and peripapillary retinal nerve fiber layer thickness did not show statistically significant changes through 12 months (55.6 ± 18.5 and 81.9 ± 9.9 μm at baseline, 52.7 ± 19.3 and 84.6 ± 15.5 μm at month 12, P > 0.05). Central macular thickness showed progressive decrease from baseline to month 12, with maximum reduction at month 3 (P retinal nerve fiber layer and ganglion cell complex thickness in 1-year follow-up. Choroidal thickness in papillomacular area and central macular thickness was significantly reduced at the end of treatment. Further studies, with larger sample, longer follow-up, and greater number of injections, are warranted.

  2. Retinal nerve fiber layer and ganglion cell complex thickness assessment in patients with Alzheimer disease and mild cognitive impairment. Preliminary results

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    A. S. Tiganov

    2014-07-01

    Full Text Available Purpose: to investigate the retinal nerve fiber layer (RNFL and the macular ganglion cell complex (GCC in patients with Alzheimer`s disease and mild cognitive impairment.Methods: this study included 10 patients (20 eyes with Alzheimer`s disease, 10 patients with mild cognitive impairment and 10 age- and sex-matched healthy controls that had no history of dementia. All the subjects underwent psychiatric examination, including the Mini-Mental State Examination (MMSE, and complete ophthalmological examination, comprising optical coherence tomography and scanning laser polarimetry.Results: there was a significant decrease in GCC thickness in patients with Alzheimer`s disease compared to the control group, global loss volume of ganglion cells was higher than in control group. there was no significant difference among the groups in terms of RNFL thickness. Weak positive correlation of GCC thickness and MMSE results was observed.Conclusion: Our data confirm the retinal involvement in Alzheimer`s disease, as reflected by loss of ganglion cells. Further studies will clear up the role and contribution of dementia in pathogenesis of optic neuropathy.

  3. Protection by an oral disubstituted hydroxylamine derivative against loss of retinal ganglion cell differentiation following optic nerve crush.

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    James D Lindsey

    Full Text Available Thy-1 is a cell surface protein that is expressed during the differentiation of retinal ganglion cells (RGCs. Optic nerve injury induces progressive loss in the number of RGCs expressing Thy-1. The rate of this loss is fastest during the first week after optic nerve injury and slower in subsequent weeks. This study was undertaken to determine whether oral treatment with a water-soluble N-hydroxy-2,2,6,6-tetramethylpiperidine derivative (OT-440 protects against loss of Thy-1 promoter activation following optic nerve crush and whether this effect targets the earlier quick phase or the later slow phase. The retina of mice expressing cyan fluorescent protein under control of the Thy-1 promoter (Thy1-CFP mice was imaged using a blue-light confocal scanning laser ophthalmoscope (bCSLO. These mice then received oral OT-440 prepared in cream cheese or dissolved in water, or plain vehicle, for two weeks and were imaged again prior to unilateral optic nerve crush. Treatments and weekly imaging continued for four more weeks. Fluorescent neurons were counted in the same defined retinal areas imaged at each time point in a masked fashion. When the counts at each time point were directly compared, the numbers of fluorescent cells at each time point were greater in the animals that received OT-440 in cream cheese by 8%, 27%, 52% and 60% than in corresponding control animals at 1, 2, 3 and 4 weeks after optic nerve crush. Similar results were obtained when the vehicle was water. Rate analysis indicated the protective effect of OT-440 was greatest during the first two weeks and was maintained in the second two weeks after crush for both the cream cheese vehicle study and water vehicle study. Because most of the fluorescent cells detected by bCSLO are RGCs, these findings suggest that oral OT-440 can either protect against or delay early degenerative responses occurring in RGCs following optic nerve injury.

  4. Gene therapy with brain-derived neurotrophic factor as a protection: retinal ganglion cells in a rat glaucoma model.

    Science.gov (United States)

    Martin, Keith R G; Quigley, Harry A; Zack, Donald J; Levkovitch-Verbin, Hana; Kielczewski, Jennifer; Valenta, Danielle; Baumrind, Lisa; Pease, Mary Ellen; Klein, Ronald L; Hauswirth, William W

    2003-10-01

    To develop a modified adenoassociated viral (AAV) vector capable of efficient transfection of retinal ganglion cells (RGCs) and to test the hypothesis that use of this vector to express brain-derived neurotrophic factor (BDNF) could be protective in experimental glaucoma. Ninety-three rats received one unilateral, intravitreal injection of either normal saline (n = 30), AAV-BDNF-woodchuck hepatitis posttranscriptional regulatory element (WPRE; n = 30), or AAV-green fluorescent protein (GFP)-WPRE (n = 33). Two weeks later, experimental glaucoma was induced in the injected eye by laser application to the trabecular meshwork. Survival of RGCs was estimated by counting axons in optic nerve cross sections after 4 weeks of glaucoma. Transgene expression was assessed by immunohistochemistry, Western blot analysis, and direct visualization of GFP. The density of GFP-positive cells in retinal wholemounts was 1,828 +/- 299 cells/mm(2) (72,273 +/- 11,814 cells/retina). Exposure to elevated intraocular pressure was similar in all groups. Four weeks after initial laser treatment, axon loss was 52.3% +/- 27.1% in the saline-treated group (n = 25) and 52.3% +/- 24.2% in the AAV-GFP-WPRE group (n = 30), but only 32.3% +/- 23.0% in the AAV-BDNF-WPRE group (n = 27). Survival in AAV-BDNF-WPRE animals increased markedly and the difference was significant compared with those receiving either AAV-GFP-WPRE (P = 0.002, t-test) or saline (P = 0.006, t-test). Overexpression of the BDNF gene protects RGC as estimated by axon counts in a rat glaucoma model, further supporting the potential feasibility of neurotrophic therapy as a complement to the lowering of IOP in the treatment of glaucoma.

  5. Relationship between macular ganglion cell complex parameters and visual field parameters after tumor resection in chiasmal compression.

    Science.gov (United States)

    Ohkubo, Shinji; Higashide, Tomomi; Takeda, Hisashi; Murotani, Eiji; Hayashi, Yasuhiko; Sugiyama, Kazuhisa

    2012-01-01

    To evaluate the relationship between macular ganglion cell complex (GCC) parameters and visual field (VF) parameters in chiasmal compression and the potential for GCC parameters in order to predict the short-term postsurgical VF. Twenty-three eyes of 12 patients with chiasmal compression and 33 control eyes were studied. All patients underwent transsphenoidal tumor resection. Before surgery a 3D scan of the macula was taken using spectral-domain optical coherence tomography. All patients underwent Humphrey 24-2 VF testing after surgery. Spearman's rank correlation coefficients were used to evaluate the relationship between the GCC parameters and VF parameters [mean deviation (MD), pattern standard deviation]. Coefficients of determination (R2) were calculated using linear regression. Average thickness in the patients was significantly thinner than that of controls. Average thickness, global loss volume and focal loss volume (FLV) significantly correlated with the MD. We observed the greatest R2 between FLV and MD. Examining the macular GCC was useful for evaluating structural damage in patients with chiasmal compression. Preoperative GCC parameters, especially FLV, may be useful in predicting visual function following surgical decompression of chiasmal compression.

  6. Selectivity of direct and network-mediated stimulation of the retinal ganglion cells with epi-, sub- and intraretinal electrodes

    Science.gov (United States)

    Boinagrov, David; Pangratz-Fuehrer, Susanne; Goetz, Georges; Palanker, Daniel

    2014-04-01

    Objective. Intra-retinal placement of stimulating electrodes can provide close and stable proximity to target neurons. We assessed improvement in stimulation thresholds and selectivity of the direct and network-mediated retinal stimulation with intraretinal electrodes, compared to epiretinal and subretinal placements. Approach. Stimulation thresholds of the retinal ganglion cells (RGCs) in wild-type rat retina were measured using the patch-clamp technique. Direct and network-mediated responses were discriminated using various synaptic blockers. Main results. Three types of RGC responses were identified: short latency (SL, τ 40 ms) originating in photoreceptors. Cathodic epiretinal stimulation exhibited the lowest threshold for direct RGC response and the highest direct selectivity (network/direct thresholds ratio), exceeding a factor of 3 with pulse durations below 0.5 ms. For network-mediated stimulation, the lowest threshold was obtained with anodic pulses in OPL position, and its network selectivity (direct/network thresholds ratio) increased with pulse duration, exceeding a factor of 4 at 10 ms. Latency of all three types of responses decreased with increasing strength of the stimulus. Significance. These results define the optimal range of pulse durations, pulse polarities and electrode placement for the retinal prostheses aiming at direct or network-mediated stimulation of RGCs.

  7. Temporal response properties of retinal ganglion cells in rd1 mice evoked by amplitude-modulated electrical pulse trains.

    Science.gov (United States)

    Ryu, Sang Baek; Ye, Jang Hee; Goo, Yong Sook; Kim, Chi Hyun; Kim, Kyung Hwan

    2010-12-01

    The electrophysiological properties of degenerated retinas responding to amplitude-modulated electrical pulse trains were investigated to provide a guideline for the development of a stimulation strategy for retinal prostheses. The activities of retinal ganglion cells (RGCs) in response to amplitude-modulated pulse trains were recorded from an in vitro model of retinal prosthesis, which consisted of an rd1 mouse retinal patch attached to a planar multielectrode array. The ability of the population activities of RGCs to effectively represent, or encode, the information on the visual intensity time series, when the intensity of visual input is transformed to pulse amplitudes, was investigated. An optimal pulse amplitude range was selected so that RGC firing rates increased monotonically and linearly. An approximately 10-Hz rhythm was observed in the field potentials from degenerated retinas, which resulted in a rhythmic burst of spontaneous spikes. Multiple peaks were present in poststimulus time histograms, with interpeak intervals corresponding to the oscillation frequency of the field potentials. Phase resetting of the field potential oscillation by stimulation was consistently observed. Despite a prominent alteration of the properties of electrically evoked firing with respect to normal retinas, RGC response strengths could be modulated by pulse amplitude. Accordingly, the temporal information of stimulation could be faithfully represented in the RGC firing patterns by an amplitude-modulated pulse train. The results suggest that pulse amplitude modulation is a feasible means of implementing a stimulation strategy for retinal prostheses, despite the marked change in the physiological properties of RGCs in degenerated retinas.

  8. Inducible ablation of melanopsin-expressing retinal ganglion cells reveals their central role in non-image forming visual responses.

    Directory of Open Access Journals (Sweden)

    Megumi Hatori

    Full Text Available Rod/cone photoreceptors of the outer retina and the melanopsin-expressing retinal ganglion cells (mRGCs of the inner retina mediate non-image forming visual responses including entrainment of the circadian clock to the ambient light, the pupillary light reflex (PLR, and light modulation of activity. Targeted deletion of the melanopsin gene attenuates these adaptive responses with no apparent change in the development and morphology of the mRGCs. Comprehensive identification of mRGCs and knowledge of their specific roles in image-forming and non-image forming photoresponses are currently lacking. We used a Cre-dependent GFP expression strategy in mice to genetically label the mRGCs. This revealed that only a subset of mRGCs express enough immunocytochemically detectable levels of melanopsin. We also used a Cre-inducible diphtheria toxin receptor (iDTR expression approach to express the DTR in mRGCs. mRGCs develop normally, but can be acutely ablated upon diphtheria toxin administration. The mRGC-ablated mice exhibited normal outer retinal function. However, they completely lacked non-image forming visual responses such as circadian photoentrainment, light modulation of activity, and PLR. These results point to the mRGCs as the site of functional integration of the rod/cone and melanopsin phototransduction pathways and as the primary anatomical site for the divergence of image-forming and non-image forming photoresponses in mammals.

  9. Changing coupling pattern of The ON-OFF direction-selective ganglion cells in early postnatal mouse retina.

    Science.gov (United States)

    Xu, Z; Zeng, Q; Shi, X; He, S

    2013-10-10

    In the adult rabbit and mouse retina, about 30% of the ON-OFF direction selective ganglion cells (DSGCs) are coupled via gap junctions. In early postnatal rabbit retinas, a greater proportion of morphological ON-OFF DSGCs shows coupling with a larger number of nearby somas. It is not clear whether the coupled ON-OFF DSGCs belong to the same subtype, or how coupling patterns change during development. In this study, we showed that in adult mouse retinas, all coupled ON-OFF DSGCs exhibited preferred directions (PDs) to superior, and this pattern emerged at postnatal day 15 (P15). At P13, the ON-OFF DSGCs with PDs to posterior were also coupled. Every ON-OFF DSGC in every subtype injected at P12 exhibited coupling. Therefore, a rapid decoupling process takes place in DSGCs around eye opening. Light deprivation delayed but did not halt the decoupling process. By using a transgenic mouse line in which green fluorescent protein (GFP) is selectively expressed in DSGCs with PDs to posterior and by performing in situ hybridization of cadherin-6, a marker for the DSGCs with PDs to superior and inferior, we showed that heterologous coupling existed between DSGCs with PDs to anterior and posterior till P12, but this heterologous coupling never spread to DSGCs positive for cadherin-6. Copyright © 2013 IBRO. Published by Elsevier Ltd. All rights reserved.

  10. Age-related hearing loss: prevention of threshold declines, cell loss and apoptosis in spiral ganglion neurons.

    Science.gov (United States)

    Frisina, Robert D; Ding, Bo; Zhu, Xiaoxia; Walton, Joseph P

    2016-09-23

    Age-related hearing loss (ARHL) -presbycusis - is the most prevalent neurodegenerative disease and number one communication disorder of our aged population; and affects hundreds of millions of people worldwide. Its prevalence is close to that of cardiovascular disease and arthritis, and can be a precursor to dementia. The auditory perceptual dysfunction is well understood, but knowledge of the biological bases of ARHL is still somewhat lacking. Surprisingly, there are no FDA-approved drugs for treatment. Based on our previous studies of human subjects, where we discovered relations between serum aldosterone levels and the severity of ARHL, we treated middle age mice with aldosterone, which normally declines with age in all mammals. We found that hearing thresholds and suprathreshold responses significantly improved in the aldosterone-treated mice compared to the non-treatment group. In terms of cellular and molecular mechanisms underlying this therapeutic effect, additional experiments revealed that spiral ganglion cell survival was significantly improved, mineralocorticoid receptors were upregulated via post-translational protein modifications, and age-related intrinsic and extrinsic apoptotic pathways were blocked by the aldosterone therapy. Taken together, these novel findings pave the way for translational drug development towards the first medication to prevent the progression of ARHL.

  11. Age-related hearing loss: prevention of threshold declines, cell loss and apoptosis in spiral ganglion neurons

    Science.gov (United States)

    Zhu, Xiaoxia; Walton, Joseph P.

    2016-01-01

    Age-related hearing loss (ARHL) -presbycusis - is the most prevalent neurodegenerative disease and number one communication disorder of our aged population; and affects hundreds of millions of people worldwide. Its prevalence is close to that of cardiovascular disease and arthritis, and can be a precursor to dementia. The auditory perceptual dysfunction is well understood, but knowledge of the biological bases of ARHL is still somewhat lacking. Surprisingly, there are no FDA-approved drugs for treatment. Based on our previous studies of human subjects, where we discovered relations between serum aldosterone levels and the severity of ARHL, we treated middle age mice with aldosterone, which normally declines with age in all mammals. We found that hearing thresholds and suprathreshold responses significantly improved in the aldosterone-treated mice compared to the non-treatment group. In terms of cellular and molecular mechanisms underlying this therapeutic effect, additional experiments revealed that spiral ganglion cell survival was significantly improved, mineralocorticoid receptors were upregulated via post-translational protein modifications, and age-related intrinsic and extrinsic apoptotic pathways were blocked by the aldosterone therapy. Taken together, these novel findings pave the way for translational drug development towards the first medication to prevent the progression of ARHL. PMID:27667674

  12. Glutamatergic neurotransmission from melanopsin retinal ganglion cells is required for neonatal photoaversion but not adult pupillary light reflex.

    Directory of Open Access Journals (Sweden)

    Anton Delwig

    Full Text Available Melanopsin-expressing retinal ganglion cells (mRGCs in the eye play an important role in many light-activated non-image-forming functions including neonatal photoaversion and the adult pupillary light reflex (PLR. MRGCs rely on glutamate and possibly PACAP (pituitary adenylate cyclase-activating polypeptide to relay visual signals to the brain. However, the role of these neurotransmitters for individual non-image-forming responses remains poorly understood. To clarify the role of glutamatergic signaling from mRGCs in neonatal aversion to light and in adult PLR, we conditionally deleted vesicular glutamate transporter (VGLUT2 selectively from mRGCs in mice. We found that deletion of VGLUT2 in mRGCs abolished negative phototaxis and light-induced distress vocalizations in neonatal mice, underscoring a necessary role for glutamatergic signaling. In adult mice, loss of VGLUT2 in mRGCs resulted in a slow and an incomplete PLR. We conclude that glutamatergic neurotransmission from mRGCs is required for neonatal photoaversion but is complemented by another non-glutamatergic signaling mechanism for the pupillary light reflex in adult mice. We speculate that this complementary signaling might be due to PACAP neurotransmission from mRGCs.

  13. GCaMP expression in retinal ganglion cells characterized using a low-cost fundus imaging system

    Science.gov (United States)

    Chang, Yao-Chuan; Walston, Steven T.; Chow, Robert H.; Weiland, James D.

    2017-10-01

    Objective. Virus-transduced, intracellular-calcium indicators are effective reporters of neural activity, offering the advantage of cell-specific labeling. Due to the existence of an optimal time window for the expression of calcium indicators, a suitable tool for tracking GECI expression in vivo following transduction is highly desirable. Approach. We developed a noninvasive imaging approach based on a custom-modified, low-cost fundus viewing system that allowed us to monitor and characterize in vivo bright-field and fluorescence images of the mouse retina. AAV2-CAG-GCaMP6f was injected into a mouse eye. The fundus imaging system was used to measure fluorescence at several time points post injection. At defined time points, we prepared wholemount retina mounted on a transparent multielectrode array and used calcium imaging to evaluate the responsiveness of retinal ganglion cells (RGCs) to external electrical stimulation. Main results. The noninvasive fundus imaging system clearly resolves individual (RGCs and axons. RGC fluorescence intensity and the number of observable fluorescent cells show a similar rising trend from week 1 to week 3 after viral injection, indicating a consistent increase of GCaMP6f expression. Analysis of the in vivo fluorescence intensity trend and in vitro neurophysiological responsiveness shows that the slope of intensity versus days post injection can be used to estimate the optimal time for calcium imaging of RGCs in response to external electrical stimulation. Significance. The proposed fundus imaging system enables high-resolution digital fundus imaging in the mouse eye, based on off-the-shelf components. The long-term tracking experiment with in vitro calcium imaging validation demonstrates the system can serve as a powerful tool monitoring the level of genetically-encoded calcium indicator expression, further determining the optimal time window for following experiment.

  14. Shh/Boc signaling is required for sustained generation of ipsilateral projecting ganglion cells in the mouse retina.

    Science.gov (United States)

    Sánchez-Arrones, Luisa; Nieto-Lopez, Francisco; Sánchez-Camacho, Cristina; Carreres, M Isabel; Herrera, Eloisa; Okada, Ami; Bovolenta, Paola

    2013-05-15

    Sonic Hedgehog (Shh) signaling is an important determinant of vertebrate retinal ganglion cell (RGC) development. In mice, there are two major RGC populations: (1) the Islet2-expressing contralateral projecting (c)RGCs, which both produce and respond to Shh; and (2) the Zic2-expressing ipsilateral projecting RGCs (iRGCs), which lack Shh expression. In contrast to cRGCs, iRGCs, which are generated in the ventrotemporal crescent (VTC) of the retina, specifically express Boc, a cell adhesion molecule that acts as a high-affinity receptor for Shh. In Boc(-/-) mutant mice, the ipsilateral projection is significantly decreased. Here, we demonstrate that this phenotype results, at least in part, from the misspecification of a proportion of iRGCs. In Boc(-/-) VTC, the number of Zic2-positive RGCs is reduced, whereas more Islet2/Shh-positive RGCs are observed, a phenotype also detected in Zic2 and Foxd1 null embryos. Consistent with this observation, organization of retinal projections at the dorsal lateral geniculate nucleus is altered in Boc(-/-) mice. Analyses of the molecular and cellular consequences of introducing Shh into the developing VTC and Zic2 and Boc into the central retina indicate that Boc expression alone is insufficient to fully activate the ipsilateral program and that Zic2 regulates Shh expression. Taking these data together, we propose that expression of Boc in cells from the VTC is required to sustain Zic2 expression, likely by regulating the levels of Shh signaling from the nearby cRGCs. Zic2, in turn, directly or indirectly, counteracts Shh and Islet2 expression in the VTC and activates the ipsilateral program.

  15. Single-cell resolution imaging of retinal ganglion cell apoptosis in vivo using a cell-penetrating caspase-activatable peptide probe.

    Directory of Open Access Journals (Sweden)

    Xudong Qiu

    Full Text Available Peptide probes for imaging retinal ganglion cell (RGC apoptosis consist of a cell-penetrating peptide targeting moiety and a fluorophore-quencher pair flanking an effector caspase consensus sequence. Using ex vivo fluorescence imaging, we previously validated the capacity of these probes to identify apoptotic RGCs in cell culture and in an in vivo rat model of N-methyl- D-aspartate (NMDA-induced neurotoxicity. Herein, using TcapQ488, a new probe designed and synthesized for compatibility with clinically-relevant imaging instruments, and real time imaging of a live rat RGC degeneration model, we fully characterized time- and dose-dependent probe activation, signal-to-noise ratios, and probe safety profiles in vivo. Adult rats received intravitreal injections of four NMDA concentrations followed by varying TcapQ488 doses. Fluorescence fundus imaging was performed sequentially in vivo using a confocal scanning laser ophthalmoscope and individual RGCs displaying activated probe were counted and analyzed. Rats also underwent electroretinography following intravitreal injection of probe. In vivo fluorescence fundus imaging revealed distinct single-cell probe activation as an indicator of RGC apoptosis induced by intravitreal NMDA injection that corresponded to the identical cells observed in retinal flat mounts of the same eye. Peak activation of probe in vivo was detected 12 hours post probe injection. Detectable fluorescent RGCs increased with increasing NMDA concentration; sensitivity of detection generally increased with increasing TcapQ488 dose until saturating at 0.387 nmol. Electroretinography following intravitreal injections of TcapQ488 showed no significant difference compared with control injections. We optimized the signal-to-noise ratio of a caspase-activatable cell penetrating peptide probe for quantitative non-invasive detection of RGC apoptosis in vivo. Full characterization of probe performance in this setting creates an important in

  16. Identification of the mononuclear cell infiltrate in the superior cervical ganglion of athymic nude and euthymic rats after guanethidine-induced sympathectomy

    DEFF Research Database (Denmark)

    Thygesen, P; Hougen, H P; Christensen, H B

    1990-01-01

    Guanethidine sulphate 40 mg/kg intraperitoneally for 14 days induced chromatolysis and nerve cell death in the superior cervical ganglia of athymic nude (rnu/rnu) LEW/Mol rats and their euthymic (+/rnu) LEW/Mol heterozygous littermates. Histologically the sympathetic ganglia were dominated...... rats. The number of NK-cells and monocytes/macrophages increased in both athymic and euthymic rats. The conclusion is, that guanethidine exerts a direct effect on sympathetic ganglion cells followed by a thymus-independent immune response....

  17. Mechanical tension applied to substrate films specifies location of neuritogenesis and promotes major neurite growth at the expense of minor neurite development.

    Science.gov (United States)

    Feng, Zhang-Qi; Franz, Eric W; Leach, Michelle K; Winterroth, Frank; White, Christina M; Rastogi, Arjun; Gu, Zhong-Ze; Corey, Joseph M

    2016-04-01

    One obstacle in neural repair is facilitating axon growth long enough to reach denervated targets. Recent studies show that axonal growth is accelerated by applying tension to bundles of neurites, and additional studies show that mechanical tension is critical to all neurite growth. However, no studies yet describe how individual neurons respond to tensile forces applied to cell bodies and neurites simultaneously; neither do any test motor neurons, a phenotype critical to neural repair. Here we examine the growth of dissociated motor neurons on stretchable substrates. E15 spinal motor neurons were cultured on poly-lactide-co-glycolide films stretched at 4.8, 9.6, or 14.3 mm day(-1). Morphological analysis revealed that substrate stretching has profound effects on developing motor neurons. Stretching increases major neurite length; it also forces neuritogenesis to occur nearest poles of the cell closest to the sources of tension. Stretching also reduces the number of neurites per neuron. These data show that substrate stretching affects neuronal morphology by specifying locations on the cell where neuritogenesis occurs and favoring major neurite growth at the expense of minor neurites. These results serve as a building block for development of new techniques to control and improve the growth of neurons for nerve repair purposes. © 2016 Wiley Periodicals, Inc.

  18. Multipronged approach to identify and validate a novel upstream regulator of Sncg in mouse retinal ganglion cells.

    Science.gov (United States)

    Chintalapudi, Sumana R; Morales-Tirado, Vanessa M; Williams, Robert W; Jablonski, Monica M

    2016-02-01

    Loss of retinal ganglion cells (RGCs) is one of the hallmarks of retinal neurodegenerative diseases, glaucoma being one of the most common. Mechanistic studies on RGCs are hindered by the lack of sufficient primary cells and consensus regarding their signature markers. Recently, γ-synuclein (SNCG) has been shown to be highly expressed in the somas and axons of RGCs. In various mouse models of glaucoma, downregulation of Sncg gene expression correlates with RGC loss. To investigate the role of Sncg in RGCs, we used a novel systems genetics approach to identify a gene that modulates Sncg expression, followed by confirmatory studies in both healthy and diseased retinae. We found that chromosome 1 harbors an expression quantitative trait locus that modulates Sncg expression in the mouse retina, and identified the prefoldin-2 (PFDN2) gene as the candidate upstream modulator of Sncg expression. Our immunohistochemical analyses revealed similar expression patterns in both mouse and human healthy retinae, with PFDN2 colocalizing with SNCG in RGCs and their axons. In contrast, in retinae from glaucoma subjects, SNCG levels were significantly reduced, although PFDN2 levels were maintained. Using a novel flow cytometry-based RGC isolation method, we obtained viable populations of murine RGCs. Knocking down Pfdn2 expression in primary murine RGCs significantly reduced Sncg expression, confirming that Pfdn2 regulates Sncg expression in murine RGCs. Gene Ontology analysis indicated shared mitochondrial function associated with Sncg and Pfdn2. These data solidify the relationship between Sncg and Pfdn2 in RGCs, and provide a novel mechanism for maintaining RGC health. © 2015 FEBS.

  19. Activation and inhibition of retinal ganglion cells in response to epiretinal electrical stimulation: a computational modelling study

    Science.gov (United States)

    Abramian, Miganoosh; Lovell, Nigel H.; Morley, John W.; Suaning, Gregg J.; Dokos, Socrates

    2015-02-01

    Objective. Retinal prosthetic devices aim to restore sight in visually impaired people by means of electrical stimulation of surviving retinal ganglion cells (RGCs). This modelling study aims to demonstrate that RGC inhibition caused by high-intensity cathodic pulses greatly influences their responses to epiretinal electrical stimulation and to investigate the impact of this inhibition on spatial activation profiles as well as their implications for retinal prosthetic device design. Another aim is to take advantage of this inhibition to reduce axonal activation in the nerve fibre layer. Approach. A three-dimensional finite-element model of epiretinal electrical stimulation was utilized to obtain RGC activation and inhibition threshold profiles for a range of parameters. Main results. RGC activation and inhibition thresholds were highly dependent on cell and stimulus parameters. Activation thresholds were 1.5, 3.4 and 11.3 μA for monopolar electrodes with 5, 20 and 50 μm radii, respectively. Inhibition to activation threshold ratios were mostly within the range 2-10. Inhibition significantly altered spatial patterns of RGC activation. With concentric electrodes and appropriately high levels of stimulus amplitudes, activation of passing axons was greatly reduced. Significance. RGC inhibition significantly impacts their spatial activation profiles, and therefore it most likely influences patterns of perceived phosphenes induced by retinal prosthetic devices. Thus this inhibition should be taken into account in future studies concerning retinal prosthesis development. It might be possible to utilize this inhibitory effect to bypass activation of passing axons and selectively stimulate RGCs near their somas and dendrites to achieve more localized phosphenes.

  20. Substituting mouse transcription factor Pou4f2 with a sea urchin orthologue restores retinal ganglion cell development.

    Science.gov (United States)

    Mao, Chai-An; Agca, Cavit; Mocko-Strand, Julie A; Wang, Jing; Ullrich-Lüter, Esther; Pan, Ping; Wang, Steven W; Arnone, Maria Ina; Frishman, Laura J; Klein, William H

    2016-03-16

    Pou domain transcription factor Pou4f2 is essential for the development of retinal ganglion cells (RGCs) in the vertebrate retina. A distant orthologue of Pou4f2 exists in the genome of the sea urchin (class Echinoidea) Strongylocentrotus purpuratus (SpPou4f1/2), yet the photosensory structure of sea urchins is strikingly different from that of the mammalian retina. Sea urchins have no obvious eyes, but have photoreceptors clustered around their tube feet disc. The mechanisms that are associated with the development and function of photoreception in sea urchins are largely unexplored. As an initial approach to better understand the sea urchin photosensory structure and relate it to the mammalian retina, we asked whether SpPou4f1/2 could support RGC development in the absence of Pou4f2. To answer this question, we replaced genomic Pou4f2 with an SpPou4f1/2 cDNA. In Pou4f2-null mice, retinas expressing SpPou4f1/2 were outwardly identical to those of wild-type mice. SpPou4f1/2 retinas exhibited dark-adapted electroretinogram scotopic threshold responses, indicating functionally active RGCs. During retinal development, SpPou4f1/2 activated RGC-specific genes and in S. purpuratus, SpPou4f2 was expressed in photoreceptor cells of tube feet in a pattern distinct from Opsin4 and Pax6. Our results suggest that SpPou4f1/2 and Pou4f2 share conserved components of a gene network for photosensory development and they maintain their conserved intrinsic functions despite vast morphological differences in mouse and sea urchin photosensory structures. © 2016 The Authors.

  1. TLR4 receptor expression and function in F11 dorsal root ganglion × neuroblastoma hybrid cells.

    Science.gov (United States)

    Hashemian, Sanaz; Alhouayek, Mireille; Fowler, Christopher J

    2017-11-01

    TLR4 respond to bacterial LPS to produce inflammatory cytokines. TLR4 are expressed in dorsal root ganglia and play a role in pain. F11 dorsal root ganglia × mouse neuroblastoma cells possess many of the properties seen in nociceptive dorsal root ganglia neuronal cells. Here, we investigated the effect of 2 h and 6 h treatment with LPS upon the expression of inflammatory proteins in undifferentiated and differentiated F11 cells. The cells expressed mRNA for TRL4 (mouse, not rat) and proteins involved in TLR4 signaling. TLR4 expression was confirmed using immunohistochemistry. LPS produced modest increases in mouse and rat IL-6 and in mouse cyclooxygenase-2 levels in undifferentiated cells, but did not significantly affect mouse TNF-α expression. This contrasts with the robust effects of LPS upon cyclooxygenase-2 expression in cultured dorsal root ganglia neurons. F11 cells expressed the endocannabinoid metabolizing enzymes fatty acid amide hydrolase and N-acylethanolamine acid amidase (both murine), which were functionally active. These data suggest that F11 cells are not a useful model for the study of LPS-mediated effects but may be useful for the study of endocannabinoid catabolism.

  2. Longitudinal in vivo imaging of retinal ganglion cells and retinal thickness changes following optic nerve injury in mice.

    Directory of Open Access Journals (Sweden)

    Balwantray C Chauhan

    Full Text Available Retinal ganglion cells (RGCs die in sight-threatening eye diseases. Imaging RGCs in humans is not currently possible and proof of principle in experimental models is fundamental for future development. Our objective was to quantify RGC density and retinal thickness following optic nerve transection in transgenic mice expressing cyan fluorescent protein (CFP under control of the Thy1 promoter, expressed by RGCs and other neurons.A modified confocal scanning laser ophthalmoscopy (CSLO/spectral-domain optical coherence tomography (SD-OCT camera was used to image and quantify CFP+ cells in mice from the B6.Cg-Tg(Thy1-CFP23Jrs/J line. SD-OCT circle (1 B-scan, raster (37 B-scans and radial (24 B-scans scans of the retina were also obtained. CSLO was performed at baseline (n = 11 and 3 (n = 11, 5 (n = 4, 7 (n = 10, 10 (n = 6, 14 (n = 7 and 21 (n = 5 days post-transection, while SD-OCT was performed at baseline and 7, 14 and 35 days (n = 9 post-transection. Longitudinal change in CFP+ cell density and retinal thickness were computed. Compared to baseline, the mean (SD percentage CFP+ cells remaining at 3, 5, 7, 10, 14 and 21 days post-transection was 86 (9%, 63 (11%, 45 (11%, 31 (9%, 20 (9% and 8 (4%, respectively. Compared to baseline, the mean (SD retinal thickness at 7 days post-transection was 97 (3%, 98 (2% and 97 (4% for the circle, raster and radial scans, respectively. The corresponding figures at 14 and 35 days post-transection were 96 (3%, 97 (2% and 95 (3%; and 93 (3%, 94 (3% and 92 (3%.Longitudinal imaging showed an exponential decline in CFP+ cell density and a small (≤8% reduction in SD-OCT measured retinal thickness post-transection. SD-OCT is a promising tool for detecting structural changes in experimental optic neuropathy. These results represent an important step towards translation for clinical use.

  3. Effect of lycium barbarum polysaccharides on high glucose-induced retinal ganglion cell apoptosis, gene expression and delayed rectifier potassium current

    Directory of Open Access Journals (Sweden)

    Xiao-Fei Ma

    2017-05-01

    Full Text Available Objective: To study the effect of lycium barbarum polysaccharides (LBP on high glucoseinduced retinal ganglion cell apoptosis, gene expression and delayed rectifier potassium current. Methods: RGC-5 retinal ganglion cell lines were cultured and divided into control group, high glucose group and LBP group that were treated with normal DMEM, highglucose DMEM as well as high-glucose DMEM containing 500 ng/mL LBP respectively. After treatment, the Annexin V-FITC/PI kits were used to measure the number of apoptotic cells, fluorescence quantitative PCR kits were used to determine the expression of apoptosis genes and antioxidant genes, and patch clamp was used to test delayed rectifier potassium current. Results: 12, 24, 36 and 48 h after intervention, the number of apoptotic cells of high glucose group was significantly higher than that of control group, and the number of apoptotic cells of LBP group was significantly lower than that of high glucose group (P<0.05; 24 and 48 h after intervention, c-fos, c-jun, caspase-3, caspase-9, Nrf-2, NQO1 and HO-1 mRNA expression as well as potassium current amplitude (IK and maximum conductance (Gmax of high glucose group were significantly higher than those of control group while half maximum activation voltage (V1/2 was significantly lower than that of control group (P<0.05; c-fos, c-jun, caspase-3 and caspase-9 mRNA expression as well as IK and Gmax of LBP group were significantly lower than those of high glucose group, while Nrf-2, NQO1 and HO-1 mRNA expression as well as V1/2 of LBP group were significantly higher than those of high glucose group (P<0.05. Conclusions: LBP can reduce the high glucose-induced retinal ganglion cell apoptosis and inhibit the delayed rectifier potassium current amplitude.

  4. Macular ganglion cell complex and retinal nerve fiber layer comparison in different stages of age-related macular degeneration.

    Science.gov (United States)

    Zucchiatti, Ilaria; Parodi, Maurizio Battaglia; Pierro, Luisa; Cicinelli, Maria Vittoria; Gagliardi, Marco; Castellino, Niccolò; Bandello, Francesco

    2015-09-01

    To employ optical coherence tomography (OCT) to analyze the morphologic changes in the inner retina in different categories of age-related macular degeneration (AMD). Observational cross-sectional study. Single-center study. Inclusion criteria were age over 50, diagnosis of Age-Related Eye Disease Study (AREDS) category 2 and 3, naïve neovascular AMD, and atrophic AMD. Healthy patients of similar age acted as a control group. Primary outcome measures were the changes in ganglion cell complex (GCC) and retinal nerve fiber layer (RNFL). Secondary outcomes included modifications of rim area and cup-to-disc ratio. One hundred and thirty eyes of 130 patients were recruited: 26 eyes for AREDS category 2, 26 for AREDS category 3, 26 for neovascular AMD, 26 with atrophic AMD, and 26 controls. Mean peripapillary RNFL thickness was significantly lower in neovascular AMD, compared to controls (P = .004); peripapillary RNFL did not significantly vary among AREDS category 2 and 3 and atrophic AMD groups, compared to controls. Mean GCC thickness was higher in the control group, becoming progressively thinner up to neovascular and atrophic AMD groups (P < .0001). Rim area was significantly thinner in the neovascular AMD group compared with controls (P = .047); cup-to-disc ratio was higher in the neovascular AMD group compared with the control group (P = .047). This study demonstrates that eyes with neovascular AMD display reduced RNFL and GCC thickness. RNFL is partially spared in atrophic advanced AMD. The identification of alteration in RNFL and GCC thickness may reveal useful for future therapeutic implications. Copyright © 2015 Elsevier Inc. All rights reserved.

  5. Studies of Scleral Biomechanical Behavior Related to Susceptibility for Retinal Ganglion Cell Loss in Experimental Mouse Glaucoma

    Science.gov (United States)

    Nguyen, Cathy; Cone, Frances E.; Nguyen, Thao D.; Coudrillier, Baptiste; Pease, Mary E.; Steinhart, Matthew R.; Oglesby, Ericka N.; Jefferys, Joan L.; Quigley, Harry A.

    2013-01-01

    Purpose. To study anatomical changes and mechanical behavior of the sclera in mice with experimental glaucoma by comparing CD1 to B6 mice. Methods. Chronic experimental glaucoma for 6 weeks was produced in 2- to 4-month-old CD1 (43 eyes) and B6 mice (42 eyes) using polystyrene bead injection into the anterior chamber with 126 control CD1 and 128 control B6 eyes. Intraocular pressure (IOP) measurements were made with the TonoLab at baseline and after bead injection. Axial length and scleral thickness were measured after sacrifice in the CD1 and B6 animals and compared to length data from 78 eyes of DBA/2J mice. Inflation testing of posterior sclera was conducted, and circumferential and meridional strain components were determined from the displacement response. Results. Experimental glaucoma led to increases in axial length and width by comparison to fellow eyes (6% in CD1 and 10% in B6; all P glaucoma, the remainder of the sclera uniformly thinned in CD1, but thickened in B6. Peripapillary sclera in CD1 controls had significantly greater temporal meridional strain than B6 and had differences in the ratios of meridional to effective circumferential strain from B6 mice. In both CD1 and B6 mice, exposure to chronic IOP elevation resulted in stiffer pressure–strain responses for both the effective circumferential and meridional strains (multivariable regression model, P = 0.01–0.03). Conclusions. Longer eyes, greater scleral strain in some directions at baseline, and generalized scleral thinning after glaucoma were characteristic of CD1 mice that have greater tendency to retinal ganglion cell damage than B6 mice. Increased scleral stiffness after glaucoma exposure in mice mimics findings in monkey and human glaucoma eyes. PMID:23404116

  6. Retinal Ganglion Cell Protection Via Topical and Systemic Alpha-Tocopherol Administration in Optic Nerve Crush Model of Rat

    Directory of Open Access Journals (Sweden)

    Zeynep Aktaş

    2013-06-01

    Full Text Available Pur po se: The aim of our study was to investigate the neuroprotective effects of topical α-tocopherol in optic nerve crush model of rat and to compare its efficacy with that of systemic α -tocopherol. Ma te ri al and Met hod: 50 eyes of 25 Wistar albino rats were included. The eyes were divided into six groups. Optic nerve crush was performed in Groups 1, 3, 5. Additionally, systemic and topical α-tocopherol therapies were given to Groups 1 and 3, respectively. No treatment was applied in Group 5. Groups 2, 4, and 6 were the fellow eyes of the animals comprising Groups 1, 3, and 5. Eyes were enucleated at day 45 of the study. Retinal ganglion cells (RGCs were counted with light microscopy. Re sults: Mean RGC numbers were 14.5±3.7 (10.3-20 and 27.5±2.6 (24-30 in Groups 5 and 6, respectively (p: 0.001 They were measured to be 26.6±7.8 (19-45 and 24.6±3.9 (20-32 in Groups 1 and 2 and 21.1±7.1 (11-34 and 27±7.5 (18-42 in Groups 3 and 4 (p:0.659, p:0.094, respectively. There was no difference in Groups 2 and 4 compared with Group 6 (p:0.210, p:0.299, respectively. Dis cus si on: Topical α-tocopherol has a significant neuroprotective effects in optic nerve crush model of rat and may be used in the future for the treatment of optic neuropathies such as glaucoma. (Turk J Ophthalmol 2013; 43: 161-6

  7. Schwann cells genetically modified to express S100A4 increases GAP43 expression in spiral ganglion neurons in vitro.

    Science.gov (United States)

    Lei, Li; Tang, Li

    2017-07-04

    Schwann cells (SCs) have been reported as a possible source of neurotrophic support for spiral ganglion neurons (SGNs). This study was aimed to investigate whether S100A4 was contributed in the functional effects of SCs on SGNs. SCs were transfected with S100A4 vector or small interfering RNA (siRNA) against S100A4, and the transfection efficiency was verified by quantitative PCR (qPCR) and Western blot. The migration of transfected SCs was determined by Transwell assay, and the expression levels of vascular endothelial growth factor precursor (VEGF) and matrix metallopeptidase 9 (MMP-9) were measured by Western blot. Co-culture of either S100A4 overexpressed or suppressed SCs with SGNs, and the growth associated protein 43 (GAP43) expression in SGNs was detected by immunofluorescence (IF), qPCR and Western blot. The migration of SCs was significantly enhanced by S100A4 overexpression (P < 0.001), while was suppressed by S100A4 knockdown (P < 0.01). Further, the expressions of VEGF and MMP-9 were notably up-regulated by S100A4 overexpression, while were down-regulated by S100A4 knockdown. Moreover, co-culture with the S100A4 overexpressed SCs significantly increased the expression of GAP43 in SGNs (P < 0.01). As expected, co-culture with S100A4 knockdown SCs decreased GAP43 level (P < 0.05). S100A4 enhanced the migratory ability of SCs. SCs genetically modified to overexpress the S100A4 could up-regulate the GAP43 expression in SGNs.

  8. Macular Ganglion Cell Inner Plexiform Layer Thickness in Glaucomatous Eyes with Localized Retinal Nerve Fiber Layer Defects.

    Directory of Open Access Journals (Sweden)

    Chunwei Zhang

    Full Text Available To investigate macular ganglion cell-inner plexiform layer (mGCIPL thickness in glaucomatous eyes with visible localized retinal nerve fiber layer (RNFL defects on stereophotographs.112 healthy and 149 glaucomatous eyes from the Diagnostic Innovations in Glaucoma Study (DIGS and the African Descent and Glaucoma Evaluation Study (ADAGES subjects had standard automated perimetry (SAP, optical coherence tomography (OCT imaging of the macula and optic nerve head, and stereoscopic optic disc photography. Masked observers identified localized RNFL defects by grading of stereophotographs.47 eyes had visible localized RNFL defects on stereophotographs. Eyes with visible localized RNFL defects had significantly thinner mGCIPL thickness compared to healthy eyes (68.3 ± 11.4 μm versus 79.2 ± 6.6 μm respectively, P<0.001 and similar mGCIPL thickness to glaucomatous eyes without localized RNFL defects (68.6 ± 11.2 μm, P = 1.000. The average mGCIPL thickness in eyes with RNFL defects was 14% less than similarly aged healthy controls. For 29 eyes with a visible RNFL defect in just one hemiretina (superior or inferior mGCIPL was thinnest in the same hemiretina in 26 eyes (90%. Eyes with inferior-temporal RNFL defects also had significantly thinner inferior-temporal mGCIPL (P<0.001 and inferior mGCIPL (P = 0.030 compared to glaucomatous eyes without a visible RNFL defect.The current study indicates that presence of a localized RNFL defect is likely to indicate significant macular damage, particularly in the region of the macular that topographically corresponds to the location of the RNFL defect.

  9. Protection of neurons in the retinal ganglion cell layer against excitotoxicity by the N-acylethanolamine, N-linoleoylethanolamine

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    Duncan RS

    2011-04-01

    Full Text Available R. Scott Duncan1,*, Hua Xin1,*, Daryl L Goad1, Kent D Chapman2,3, Peter Koulen1,31Vision Research Center and Departments of Ophthalmology and Basic Medical Science, School of Medicine, University of Missouri, Kansas City, MO, USA; 2Department of Biological Sciences, University of North Texas, Denton, TX, USA; 3Center for Plant Lipid Research, University of North Texas, Denton, TX, USA *Authors contributed equallyAbstract: Retinal ganglion cell (RGC death is a hallmark of neurodegenerative diseases and disease processes of the eye, including glaucoma. The protection of RGCs has been an important strategy for combating glaucoma, but little clinical success has been reported to date. One pathophysiological consequence of glaucoma is excessive extracellular glutamate subsequently leading to excitotoxicity in the retina. Endocannabinoids, such as the N-acylethanolamine (NAE, arachidonylethanolamine (NAE 20:4, exhibit neuroprotective properties in some models of neurodegenerative disease. The majority of NAEs, however, are not cannabinoids, and their physiological function is not clear. Here, we determined whether the noncannabinoid NAE, linoleoylethanolamine (NAE18:2, protects neurons in the RGC layer against glutamate excitotoxicity in ex-vivo retina cultures. Using a terminal deoxynucleotidyl transferase-mediated dUTP (2´-deoxyuridine 5´-triphosphate nick-end labeling (TUNEL assay, we determined that NAE18:2 reduces the number of apoptotic RGC layer neurons in response to glutamate and conclude that NAE18:2 is a neuroprotective compound with potential for treating glaucomatous retinopathy.Keywords: neuroprotection, glutamate, calcium signaling, immunocytochemistry, eye, vision, glaucoma.

  10. Neuroprotective effects of tempol acyl esters against retinal ganglion cell death in a rat partial optic nerve crush model.

    Science.gov (United States)

    Thaler, Sebastian; Fiedorowicz, Michal; Grieb, Pawel; Wypych, Zbigniew; Knap, Narcyz; Borowik, Tomasz; Zawada, Katarzyna; Kaminski, Jaroslaw; Wozniak, Michal; Rejdak, Robert; Zrenner, Eberhart; Schuettauf, Frank

    2011-11-01

    The aim of this study is to search for more effective derivatives of the superoxide dismutase mimetic tempol (4-hydroxy-2,2,6,6-tetramethylpiperidine-1-oxyl). Although tempol is neuroprotective in a rat partial optic nerve crush (PONC) model, relatively high doses are required to exert this effect. Tempol acyl esters with different-length fatty acids (tempol-C4, tempol-C8, tempol-C12 and tempol-C16) were synthesized and the following properties were evaluated: water-octanol partition coefficient, liposome-liposome energy transfer, and electron paramagnetic resonance (EPR). Brown Norway rats underwent PONC and received tempol or acyl esters intraperitoneally once daily for 7 consecutive days. We then compared the effects of tempol and its four esters on retinal ganglion cell (RGC) damage using a retrograde labelling method. The water-octanol partition coefficient increased with increasing length of attached acyl chain. However, the energy of the liposome-liposome transfer seemed to be optimal for tempol-C8 and tempol-C12. The EPR signal was very similar for all tested compounds, suggesting similar efficiency of superoxide scavenging. Partial optic nerve crush in vehicle-treated animals reduced RGC numbers by approx. 59% when compared with sham-operated eyes. Tempol did not affect RGC loss at a dose of 1 mg/kg. In contrast, at molar doses equivalent to 1 mg/kg of tempol, tempol-C8 showed a significant neuroprotective effect, whereas tempol-C4, tempol-C12 and tempol-C16 did not act neuroprotectively. Manipulating the hydrophobicity of tempol seems to be a promising tool for developing more potent neuroprotectants in the PONC degeneration model. However, the resulting compounds need further pharmacological evaluation. © 2011 The Authors. Acta Ophthalmologica © 2011 Acta Ophthalmologica Scandinavica Foundation.

  11. Induction of Heat Shock Protein-72 by Magnetic Nanofluid Hyperthermia in Cultured Retinal Ganglion Cells for Neuroprotective Treatment in Glaucoma

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    Jin Wook Jeoung

    2015-01-01

    Full Text Available Background. Magnetic hyperthermia using superparamagnetic nanoparticle (SPNP agents is considered a promising biotechnological approach to induce heat shock proteins (HSPs in a target tissue because it can generate accurately controllable localized heating. Objectives. The main objective of this study is to demonstrate induction of HSPs in cultured retinal ganglion cells (RGCs by using engineered Mn0.5Zn0.5Fe2O4 SPNP agents coated with polyethylene glycol (PEG 500. Methods. The Mn0.5Zn0.5Fe2O4 nanoparticles were synthesized using a high temperature thermal decomposition method. The AC heating characteristics of PEG 500-coated Mn0.5Zn0.5Fe2O4 nanoparticles were investigated using an AC solenoid coil-capacitor system. Results. PEG 500-coated SPNPs efficiently penetrated into the cytoplasm of RGCs without causing obvious cytological changes and showed stable and well-saturated self-heating temperature rise characteristics. Immunofluorescent staining images showed that AC magnetic hyperthermia successfully induced HSP72 in RGCs incubated with Mn0.5Zn0.5Fe2O4 nanoparticles. In Western blot analysis, a significant increase in immunoreactivity was observed for RGCs incubated with SPNPs in a fixed AC magnetic field (fappl=140 kHz and Happl=140 Oe. Conclusion. Our results demonstrate that the induction of HSP72 with a magnetic nanofluid hyperthermia could potentially be used as a neuroprotective treatment modality by way of enhancing a natural cytoprotective response.

  12. The Photopic Negative Response: An Objective Measure of Retinal Ganglion Cell Function in Patients With Leber's Hereditary Optic Neuropathy.

    Science.gov (United States)

    Karanjia, Rustum; Berezovsky, Adriana; Sacai, Paula Yuri; Cavascan, Nivea Nunes; Liu, Henry Yuheng; Nazarali, Samir; Moraes-Filho, Milton Nunes; Anderson, Kirsten; Tran, Jeffrey Show; Watanabe, Sung EunSong; Moraes, Milton Nunes; Sadun, Federico; DeNegri, Anna Maria; Barboni, Piero; do Val Ferreira Ramos, Carolina; La Morgia, Chiara; Carelli, Valerio; Belfort, Rubens; Coupland, Stuart Glenn; Salomao, Solange Rios; Sadun, Alfredo A

    2017-05-01

    The photopic negative response (PhNR) is a slow negative component of a flash photopic full-field ERG that has been shown to be specific for retinal ganglion cell (RGC) activity. Direct evaluation of RGC function is desirable in patients with Leber's hereditary optic neuropathy (LHON) in which the loss of central acuity can make it difficult to monitor patients with standard metrics. The purpose of this study was to evaluate the use of PhNR as an objective noninvasive clinical metric in LHON. Full-field photopic ERG recordings were collected in subjects with the mt.11778G>A/ND4 LHON mutation using a red on blue stimulus. The PhNR was identified using a computer-based automated detection system, and data were manually examined to remove movement artifacts. The PhNR amplitude was compared between controls (n = 13), carriers (n = 17), and affected (n = 6). Mean PhNR amplitude decreased significantly across groups (P < 0.0001). Post hoc Tukey's test revealed a significant decrease in PhNR amplitude between carriers and controls (P < 0.05) and between carriers and affected (P < 0.01). We are able to demonstrate that the PhNR amplitude is significantly decreased in patients affected by LHON compared to carriers in a well-described pedigree. Surprisingly, there was also a decrease in PhNR in carriers, suggesting potential subclinical RGC dysfunction in some carriers. This is important in patients affected with LHON who typically have a dense central scotoma. The PhNR may be a useful objective outcome measure for future clinical trials.

  13. Transplanted skin-derived precursor stem cells generate enteric ganglion-like structures in vivo.

    Science.gov (United States)

    Wagner, Justin P; Sullins, Veronica F; Dunn, James C Y

    2014-08-01

    Hirschsprung's disease is characterized by a developmental arrest of neural crest cell migration, causing distal aganglionosis. Transplanted cells derived from the neural crest may regenerate enteric ganglia in this condition. We investigated the potential of skin-derived precursor cells (SKPs) to engraft and to differentiate into enteric ganglia in aganglionic rat intestine in vivo. Adult Lewis rat jejunal segments were separated from intestinal continuity and treated with benzalkonium chloride to induce aganglionosis. Ganglia were identified via immunohistochemical stains for S100 and β-III tubulin (TUJ1). SKPs were procured from neonatal Lewis rats expressing enhanced green fluorescent protein (GFP) and cultured in neuroglial-selective media. SKP cell line expansion was quantified, and immunophenotypes were assessed by immunocytochemistry. Aganglionic segments underwent SKP transplantation 21-79days after benzalkonium chloride treatment. The presence of GFP+cells, mature neurons, and mature glia was evaluated at posttransplant days 1, 6, and 9. Benzalkonium chloride-induced aganglionosis persisted for at least 85days. Prior to differentiation, SKPs expressed S100, denoting neural crest lineage, and nestin, a marker of neuronal precursors. Differentiated SKPs in vitro expressed GFAP, a marker of glial differentiation, as well as TUJ1 and several enteric neurotransmitters. After transplantation, GFP+structures resembling ganglia were identified between longitudinal and circular smooth muscle layers. SKPs are capable of engraftment, migration, and differentiation within aganglionic rodent intestine in vivo. Differentiated SKPs generate structures that resemble enteric ganglia. Our observations suggest that SKPs represent a potential gangliogenic therapeutic agent for Hirschsprung's disease. Copyright © 2014 Elsevier Inc. All rights reserved.

  14. Glial membranes at the node of Ranvier prevent neurite outgrowth

    DEFF Research Database (Denmark)

    Huang, Jeffrey K; Phillips, Greg R; Roth, Alejandro D

    2005-01-01

    of neurite outgrowth, including the oligodendrocyte myelin glycoprotein (OMgp). In rat spinal cord, OMgp was not localized to compact myelin, as previously thought, but to oligodendroglia-like cells, whose processes converge to form a ring that completely encircles the nodes. In OMgp-null mice, CNS nodes...

  15. Membrane progesterone receptor beta (mPRβ/Paqr8) promotes progesterone-dependent neurite outgrowth in PC12 neuronal cells via non-G protein-coupled receptor (GPCR) signaling.

    Science.gov (United States)

    Kasubuchi, Mayu; Watanabe, Keita; Hirano, Kanako; Inoue, Daisuke; Li, Xuan; Terasawa, Kazuya; Konishi, Morichika; Itoh, Nobuyuki; Kimura, Ikuo

    2017-07-12

    Recently, sex steroid membrane receptors garnered world-wide attention because they may be related to sex hormone-mediated unknown rapid non-genomic action that cannot be currently explained by their genomic action via nuclear receptors. Progesterone affects cell proliferation and survival via non-genomic effects. In this process, membrane progesterone receptors (mPRα, mPRβ, mPRγ, mPRδ, and mPRε) were identified as putative G protein-coupled receptors (GPCRs) for progesterone. However, the structure, intracellular signaling, and physiological functions of these progesterone receptors are still unclear. Here, we identify a molecular mechanism by which progesterone promotes neurite outgrowth through mPRβ (Paqr8) activation. Mouse mPRβ mRNA was specifically expressed in the central nervous system. It has an incomplete GPCR topology, presenting 6 transmembrane domains and did not exhibit typical GPCR signaling. Progesterone-dependent neurite outgrowth was exhibited by the promotion of ERK phosphorylation via mPRβ, but not via other progesterone receptors such as progesterone membrane receptor 1 (PGRMC-1) and nuclear progesterone receptor in nerve growth factor-induced neuronal PC12 cells. These findings provide new insights of regarding the non-genomic action of progesterone in the central nervous system.

  16. Transgenic inhibition of astroglial NF-κB protects from optic nerve damage and retinal ganglion cell loss in experimental optic neuritis

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    Brambilla Roberta

    2012-09-01

    Full Text Available Abstract Background Optic neuritis is an acute, demyelinating neuropathy of the optic nerve often representing the first appreciable symptom of multiple sclerosis. Wallerian degeneration of irreversibly damaged optic nerve axons leads to death of retinal ganglion cells, which is the cause of permanent visual impairment. Although the specific mechanisms responsible for triggering these events are unknown, it has been suggested that a key pathological factor is the activation of immune-inflammatory processes secondary to leukocyte infiltration. However, to date, there is no conclusive evidence to support such a causal role for infiltrating peripheral immune cells in the etiopathology of optic neuritis. Methods To dissect the contribution of the peripheral immune-inflammatory response versus the CNS-specific inflammatory response in the development of optic neuritis, we analyzed optic nerve and retinal ganglion cells pathology in wild-type and GFAP-IκBα-dn transgenic mice, where NF-κB is selectively inactivated in astrocytes, following induction of EAE. Results We found that, in wild-type mice, axonal demyelination in the optic nerve occurred as early as 8 days post induction of EAE, prior to the earliest signs of leukocyte infiltration (20 days post induction. On the contrary, GFAP-IκBα-dn mice were significantly protected and showed a nearly complete prevention of axonal demyelination, as well as a drastic attenuation in retinal ganglion cell death. This correlated with a decrease in the expression of pro-inflammatory cytokines, chemokines, adhesion molecules, as well as a prevention of NAD(PH oxidase subunit upregulation. Conclusions Our results provide evidence that astrocytes, not infiltrating immune cells, play a key role in the development of optic neuritis and that astrocyte-mediated neurotoxicity is dependent on activation of a transcriptional program regulated by NF-κB. Hence, interventions targeting the NF-κB transcription

  17. ADF/cofilin-mediated actin retrograde flow directs neurite formation in the developing brain.

    Science.gov (United States)

    Flynn, Kevin C; Hellal, Farida; Neukirchen, Dorothee; Jacob, Sonja; Tahirovic, Sabina; Dupraz, Sebastian; Stern, Sina; Garvalov, Boyan K; Gurniak, Christine; Shaw, Alisa E; Meyn, Liane; Wedlich-Söldner, Roland; Bamburg, James R; Small, J Victor; Witke, Walter; Bradke, Frank

    2012-12-20

    Neurites are the characteristic structural element of neurons that will initiate brain connectivity and elaborate information. Early in development, neurons are spherical cells but this symmetry is broken through the initial formation of neurites. This fundamental step is thought to rely on actin and microtubule dynamics. However, it is unclear which aspects of the complex actin behavior control neuritogenesis and which molecular mechanisms are involved. Here, we demonstrate that augmented actin retrograde flow and protrusion dynamics facilitate neurite formation. Our data indicate that a single family of actin regulatory proteins, ADF/Cofilin, provides the required control of actin retrograde flow and dynamics to form neurites. In particular, the F-actin severing activity of ADF/Cofilin organizes space for the protrusion and bundling of microtubules, the backbone of neurites. Our data reveal how ADF/Cofilin organizes the cytoskeleton to drive actin retrograde flow and thus break the spherical shape of neurons. Copyright © 2012 Elsevier Inc. All rights reserved.

  18. Neuroprotective copper bis(thiosemicarbazonato complexes promote neurite elongation.

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    Laura Bica

    Full Text Available Abnormal biometal homeostasis is a central feature of many neurodegenerative disorders including Alzheimer's disease (AD, Parkinson's disease (PD, and motor neuron disease. Recent studies have shown that metal complexing compounds behaving as ionophores such as clioquinol and PBT2 have robust therapeutic activity in animal models of neurodegenerative disease; however, the mechanism of neuroprotective action remains unclear. These neuroprotective or neurogenerative processes may be related to the delivery or redistribution of biometals, such as copper and zinc, by metal ionophores. To investigate this further, we examined the effect of the bis(thiosemicarbazonato-copper complex, Cu(II(gtsm on neuritogenesis and neurite elongation (neurogenerative outcomes in PC12 neuronal-related cultures. We found that Cu(II(gtsm induced robust neurite elongation in PC12 cells when delivered at concentrations of 25 or 50 nM overnight. Analogous effects were observed with an alternative copper bis(thiosemicarbazonato complex, Cu(II(atsm, but at a higher concentration. Induction of neurite elongation by Cu(II(gtsm was restricted to neurites within the length range of 75-99 µm with a 2.3-fold increase in numbers of neurites in this length range with 50 nM Cu(II(gtsm treatment. The mechanism of neurogenerative action was investigated and revealed that Cu(II(gtsm inhibited cellular phosphatase activity. Treatment of cultures with 5 nM FK506 (calcineurin phosphatase inhibitor resulted in analogous elongation of neurites compared to 50 nM Cu(II(gtsm, suggesting a potential link between Cu(II(gtsm-mediated phosphatase inhibition and neurogenerative outcomes.

  19. Erythropoietin protects adult retinal ganglion cells against NMDA-, trophic factor withdrawal-, and TNF-α-induced damage.

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    Zhi-Yang Chang

    Full Text Available PURPOSE: This study aimed to evaluate the neuroprotective effect of EPO in the presence of N-methyl-d-aspartate (NMDA-, trophic factor withdrawal (TFW-, and tumor necrosis factor-alpha (TNF-α-induced toxicity on total, small, and large retinal ganglion cells (RGCs. METHODS: Retinal cells from adult rats were cultured in a medium containing brain-derived neurotrophic factor (BDNF, ciliary neurotrophic factor (CNTF, basic fibroblast growth factor (bFGF, and forskolin. Expression of RGC markers and EPOR was examined using immunocytochemistry. RGCs were classified according to their morphological properties. Cytotoxicity was induced by NMDA, TFW, or TNF-α. RGC survival was assessed by counting thy-1 and neurofilament-l double-positive cells. RESULTS: EPO offered dose-dependent (EC₅₀ = 5.7 ng/mL protection against NMDA toxicity for small RGCs; protection was not significant for large RGCs. Time-course analysis showed that the presence of EPO either before or after NMDA exposure gave effective protection. For both small and large RGCs undergoing trophic factor withdrawal, EPO at concentrations of 1, 10, or 100 ng/mL improved survival. However, EPO had to be administered soon after the onset of injury to provide effective protection. For TNF-α-induced toxicity, survival of small RGCs was seen only for the highest examined concentration (100 ng/mL of EPO, whereas large RGCs were protected at concentrations of 1, 10, or 100 ng/mL of EPO. Time-course analysis showed that pretreatment with EPO provided protection only for large RGCs; early post-treatment with EPO protected both small and large RGCs. Inhibitors of signal transduction and activators of transcription such as (STAT-5, mitogen-activated protein kinases (MAPK/extracellular-regulated kinase (ERK, and phosphatidyl inositol-3 kinase (PI3K/Akt impaired the protective effect of EPO on RGCs exposed to different insults. CONCLUSION: EPO provided neuroprotection to cultured adult rat RGCs

  20. Neural stem cell-based intraocular administration of ciliary neurotrophic factor attenuates the loss of axotomized ganglion cells in adult mice.

    Science.gov (United States)

    Flachsbarth, Kai; Kruszewski, Katharina; Jung, Gila; Jankowiak, Wanda; Riecken, Kristoffer; Wagenfeld, Lars; Richard, Gisbert; Fehse, Boris; Bartsch, Udo

    2014-09-30

    To analyze the neuroprotective effect of intravitreally grafted neural stem (NS) cells genetically modified to secrete ciliary neurotrophic factor (CNTF) on intraorbitally lesioned retinal ganglion cells (RGCs) in adult mice. Adherently cultivated NS cells were genetically modified to express a secretable variant of mouse CNTF together with the fluorescent reporter protein Venus. Clonal CNTF-secreting NS cell lines were established using fluorescence activated cell sorting, and intravitreally grafted into adult mice 1 day after an intraorbital crush of the optic nerve. Brn-3a-positive RGCs were counted in flat-mounted retinas at different postlesion intervals to evaluate the neuroprotective effect of the CNTF-secreting NS cells on the axotomized RGCs. Anterograde axonal tracing experiments were performed to analyze the regrowth of the injured RGC axons in CNTF-treated retinas. Intravitreally grafted NS cells preferentially differentiated into astrocytes that survived in the host eyes, stably expressed CNTF, and significantly attenuated the loss of the axotomized RGCs over a period of at least 4 months, the latest postlesion time point analyzed. Depending on the postlesion interval analyzed, the number of RGCs in eyes with grafted CNTF-secreting NS cells was 2.8-fold to 6.4-fold higher than in eyes with grafted control NS cells. The CNTF-secreting NS cells additionally induced long-distance regrowth of the lesioned RGC axons. Genetically modified clonal NS cell lines may serve as a useful tool for preclinical studies aimed at evaluating the therapeutic potential of a sustained cell-based intravitreal administration of neuroprotective factors in mouse models of glaucoma. Copyright 2014 The Association for Research in Vision and Ophthalmology, Inc.

  1. Neuronal injury external to the retina rapidly activates retinal glia, followed by elevation of markers for cell cycle re-entry and death in retinal ganglion cells.

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    Alba Galan

    Full Text Available Retinal ganglion cells (RGCs are neurons that relay visual signals from the retina to the brain. The RGC cell bodies reside in the retina and their fibers form the optic nerve. Full transection (axotomy of the optic nerve is an extra-retinal injury model of RGC degeneration. Optic nerve transection permits time-kinetic studies of neurodegenerative mechanisms in neurons and resident glia of the retina, the early events of which are reported here. One day after injury, and before atrophy of RGC cell bodies was apparent, glia had increased levels of phospho-Akt, phospho-S6, and phospho-ERK1/2; however, these signals were not detected in injured RGCs. Three days after injury there were increased levels of phospho-Rb and cyclin A proteins detected in RGCs, whereas these signals were not detected in glia. DNA hyperploidy was also detected in RGCs, indicative of cell cycle re-entry by these post-mitotic neurons. These events culminated in RGC death, which is delayed by pharmacological inhibition of the MAPK/ERK pathway. Our data show that a remote injury to RGC axons rapidly conveys a signal that activates retinal glia, followed by RGC cell cycle re-entry, DNA hyperploidy, and neuronal death that is delayed by preventing glial MAPK/ERK activation. These results demonstrate that complex and variable neuro-glia interactions regulate healthy and injured states in the adult mammalian retina.

  2. A feed-forward regulation of endothelin receptors by c-Jun in human non-pigmented ciliary epithelial cells and retinal ganglion cells.

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    Junming Wang

    Full Text Available c-Jun, c-Jun N-terminal kinase(JNK and endothelin B (ETB receptor have been shown to contribute to the pathogenesis of glaucoma. Previously, we reported that an increase of c-Jun and CCAAT/enhancer binding protein β (C/EBPβ immunohistostaining is associated with upregulation of the ETB receptor within the ganglion cell layer of rats with elevated intraocular pressure (IOP. In addition, both transcription factors regulate the expression of the ETB receptor in human non-pigmented ciliary epithelial cells (HNPE. The current study addressed the mechanisms by which ET-1 produced upregulation of ET receptors in primary rat retinal ganglion cells (RGCs and HNPE cells. Treatment of ET-1 and ET-3 increased the immunocytochemical staining of c-Jun and C/EBPβ in primary rat RGCs and co-localization of both transcription factors was observed. A marked increase in DNA binding activity of AP-1 and C/EBPβ as well as elevated protein levels of c-Jun and c-Jun-N-terminal kinase (JNK were detected following ET-1 treatment in HNPE cells. Overexpression of ETA or ETB receptor promoted the upregulation of c-Jun and also elevated its promoter activity. In addition, upregulation of C/EBPβ augmented DNA binding and mRNA expression of c-Jun, and furthermore, the interaction of c-Jun and C/EBPβ was confirmed using co-immunoprecipitation. Apoptosis of HNPE cells was identified following ET-1 treatment, and overexpression of the ETA or ETB receptor produced enhanced apoptosis. ET-1 mediated upregulation of c-Jun and C/EBPβ and their interaction may represent a novel mechanism contributing to the regulation of endothelin receptor expression. Reciprocally, c-Jun was also found to regulate the ET receptors and C/EBPβ appeared to play a regulatory role in promoting expression of c-Jun. Taken together, the data suggests that ET-1 triggers the upregulation of c-Jun through both ETA and ETB receptors, and conversely c-Jun also upregulates endothelin receptor expression

  3. Long-term gene therapy causes transgene-specific changes in the morphology of regenerating retinal ganglion cells.

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    Jennifer Rodger

    Full Text Available Recombinant adeno-associated viral (rAAV vectors can be used to introduce neurotrophic genes into injured CNS neurons, promoting survival and axonal regeneration. Gene therapy holds much promise for the treatment of neurotrauma and neurodegenerative diseases; however, neurotrophic factors are known to alter dendritic architecture, and thus we set out to determine whether such transgenes also change the morphology of transduced neurons. We compared changes in dendritic morphology of regenerating adult rat retinal ganglion cells (RGCs after long-term transduction with rAAV2 encoding: (i green fluorescent protein (GFP, or (ii bi-cistronic vectors encoding GFP and ciliary neurotrophic factor (CNTF, brain-derived neurotrophic factor (BDNF or growth-associated protein-43 (GAP43. To enhance regeneration, rats received an autologous peripheral nerve graft onto the cut optic nerve of each rAAV2 injected eye. After 5-8 months, RGCs with regenerated axons were retrogradely labeled with fluorogold (FG. Live retinal wholemounts were prepared and GFP positive (transduced or GFP negative (non-transduced RGCs injected iontophoretically with 2% lucifer yellow. Dendritic morphology was analyzed using Neurolucida software. Significant changes in dendritic architecture were found, in both transduced and non-transduced populations. Multivariate analysis revealed that transgenic BDNF increased dendritic field area whereas GAP43 increased dendritic complexity. CNTF decreased complexity but only in a subset of RGCs. Sholl analysis showed changes in dendritic branching in rAAV2-BDNF-GFP and rAAV2-CNTF-GFP groups and the proportion of FG positive RGCs with aberrant morphology tripled in these groups compared to controls. RGCs in all transgene groups displayed abnormal stratification. Thus in addition to promoting cell survival and axonal regeneration, vector-mediated expression of neurotrophic factors has measurable, gene-specific effects on the morphology of injured

  4. Long-Term Gene Therapy Causes Transgene-Specific Changes in the Morphology of Regenerating Retinal Ganglion Cells

    Science.gov (United States)

    Rodger, Jennifer; Drummond, Eleanor S.; Hellström, Mats; Robertson, Donald; Harvey, Alan R.

    2012-01-01

    Recombinant adeno-associated viral (rAAV) vectors can be used to introduce neurotrophic genes into injured CNS neurons, promoting survival and axonal regeneration. Gene therapy holds much promise for the treatment of neurotrauma and neurodegenerative diseases; however, neurotrophic factors are known to alter dendritic architecture, and thus we set out to determine whether such transgenes also change the morphology of transduced neurons. We compared changes in dendritic morphology of regenerating adult rat retinal ganglion cells (RGCs) after long-term transduction with rAAV2 encoding: (i) green fluorescent protein (GFP), or (ii) bi-cistronic vectors encoding GFP and ciliary neurotrophic factor (CNTF), brain-derived neurotrophic factor (BDNF) or growth-associated protein-43 (GAP43). To enhance regeneration, rats received an autologous peripheral nerve graft onto the cut optic nerve of each rAAV2 injected eye. After 5–8 months, RGCs with regenerated axons were retrogradely labeled with fluorogold (FG). Live retinal wholemounts were prepared and GFP positive (transduced) or GFP negative (non-transduced) RGCs injected iontophoretically with 2% lucifer yellow. Dendritic morphology was analyzed using Neurolucida software. Significant changes in dendritic architecture were found, in both transduced and non-transduced populations. Multivariate analysis revealed that transgenic BDNF increased dendritic field area whereas GAP43 increased dendritic complexity. CNTF decreased complexity but only in a subset of RGCs. Sholl analysis showed changes in dendritic branching in rAAV2-BDNF-GFP and rAAV2-CNTF-GFP groups and the proportion of FG positive RGCs with aberrant morphology tripled in these groups compared to controls. RGCs in all transgene groups displayed abnormal stratification. Thus in addition to promoting cell survival and axonal regeneration, vector-mediated expression of neurotrophic factors has measurable, gene-specific effects on the morphology of injured adult

  5. Evaluation of Ganglion Cell-Inner Plexiform Layer Thickness after Vitreoretinal Surgery with Internal Limiting Membrane Peeling in Cases with Idiopathic Macular Hole

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    Sibel Demirel

    2017-06-01

    Full Text Available Objectives: To evaluate macular retinal ganglion cell-inner plexiform layer (GCIPL thickness after vitrectomy with internal limiting membrane (ILM peeling for idiopathic macular holes using spectral domain optical coherence tomography (SD-OCT. Materials and Methods: Eighteen eyes of 18 patients with unilateral idiopathic macular hole who underwent vitrectomy with ILM peeling were retrospectively analyzed. Healthy fellow eyes of the patients and 18 eyes of 18 age-matched healthy individuals constituted the control group. The patients were evaluated at 1 day, 1 week, 1 month, and 3 months after surgery. The best corrected visual acuity (BCVA measurements, biomicroscopic examination findings and SD-OCT measurements were recorded. Ganglion cell-inner plexiform layer thickness was evaluated with ganglion cell analysis software of Cirrus HD-OCT before surgery and at 1 month and 3 months after surgery and compared with control groups. Presence of dissociated optic nerve fiber layer (DONFL was evaluated with C-scan mode. Results: Of the 18 patients, 9 were male and 9 were female with a mean age of 65.6±5.6 (55-77 years. Preoperative BCVA was 0.75±0.19 logMAR, while it was 0.44±0.17 logMAR and 0.36±0.15 logMAR at postoperative 1 and 3 months, respectively (p0.05. Conclusion: Internal limiting membrane peeling during macular hole surgery may cause functional and /or structural changes that may be associated with visual acuity. Significant GCIPL thinning and DONLF appearance may occur postoperatively.

  6. Differentiation of retinal ganglion cells and photoreceptor precursors from mouse induced pluripotent stem cells carrying an Atoh7/Math5 lineage reporter.

    Directory of Open Access Journals (Sweden)

    Bin-Bin Xie

    Full Text Available The neural retina is a critical component of the visual system, which provides the majority of sensory input in humans. Various retinal degenerative diseases can result in the permanent loss of retinal neurons, especially the light-sensing photoreceptors and the centrally projecting retinal ganglion cells (RGCs. The replenishment of lost RGCs and the repair of optic nerve damage are particularly challenging, as both RGC specification and their subsequent axonal growth and projection involve complex and precise regulation. To explore the developmental potential of pluripotent stem cell-derived neural progenitors, we have established mouse iPS cells that allow cell lineage tracing of progenitors that have expressed Atoh7/Math5, a bHLH transcription factor required for RGC production. These Atoh7 lineage reporter iPS cells encode Cre to replace one copy of the endogenous Atoh7 gene and a Cre-dependent YFP reporter in the ROSA locus. In addition, they express pluripotent markers and are capable of generating teratomas in vivo. Under anterior neural induction and neurogenic conditions in vitro, the Atoh7-Cre/ROSA-YFP iPS cells differentiate into neurons that co-express various RGC markers and YFP, indicating that these neurons are derived from Atoh7-expressing progenitors. Consistent with previous in vivo cell lineage studies, the Atoh7-Cre/ROSA-YFP iPS cells also give rise to a subset of Crx-positive photoreceptor precursors. Furthermore, inhibition of Notch signaling in the iPSC cultures results in a significant increase of YFP-positive RGCs and photoreceptor precursors. Together, these results show that Atoh7-Cre/ROSA-YFP iPS cells can be used to monitor the development and survival of RGCs and photoreceptors from pluripotent stem cells.

  7. Both systemic and local application of Granulocyte-colony stimulating factor (G-CSF is neuroprotective after retinal ganglion cell axotomy

    Directory of Open Access Journals (Sweden)

    Dietz Gunnar PH

    2009-05-01

    Full Text Available Abstract Background The hematopoietic Granulocyte-Colony Stimulating Factor (G-CSF plays a crucial role in controlling the number of neutrophil progenitor cells. Its function is mediated via the G-CSF receptor, which was recently found to be expressed also in the central nervous system. In addition, G-CSF provided neuroprotection in models of neuronal cell death. Here we used the retinal ganglion cell (RGC axotomy model to compare effects of local and systemic application of neuroprotective molecules. Results We found that the G-CSF receptor is robustly expressed by RGCs in vivo and in vitro. We thus evaluated G-CSF as a neuroprotectant for RGCs and found a dose-dependent neuroprotective effect of G-CSF on axotomized RGCs when given subcutaneously. As stem stell mobilization had previously been discussed as a possible contributor to the neuroprotective effects of G-CSF, we compared the local treatment of RGCs by injection of G-CSF into the vitreous body with systemic delivery by subcutaneous application. Both routes of application reduced retinal ganglion cell death to a comparable extent. Moreover, G-CSF enhanced the survival of immunopurified RGCs in vitro. Conclusion We thus show that G-CSF neuroprotection is at least partially independent of potential systemic effects and provide further evidence that the clinically applicable G-CSF could become a treatment option for both neurodegenerative diseases and glaucoma.

  8. miRNA-141 attenuates UV-induced oxidative stress via activating Keap1-Nrf2 signaling in human retinal pigment epithelium cells and retinal ganglion cells

    Science.gov (United States)

    Cheng, Li-Bo; Li, Ke-ran; Yi, Nan; Li, Xiu-miao; Wang, Feng; Xue, Bo; Pan, Ying-shun; Yao, Jin; Jiang, Qin; Wu, Zhi-feng

    2017-01-01

    Activation of NF-E2-related factor 2 (Nrf2) signaling could protect cells from ultra violet (UV) radiation. We aim to provoke Nrf2 activation via downregulating its inhibitor Keap1 by microRNA-141 (“miR-141”). In both human retinal pigment epithelium cells (RPEs) and retinal ganglion cells (RGCs), forced-expression of miR-141 downregulated Keap1, causing Nrf2 stabilization, accumulation and nuclear translocation, which led to transcription of multiple antioxidant-responsive element (ARE) genes (HO1, NOQ1 and GCLC). Further, UV-induced reactive oxygen species (ROS) production and cell death were significantly attenuated in miR-141-expressing RPEs and RGCs. On the other hand, depletion of miR-141 via expressing its inhibitor antagomiR-141 led to Keap1 upregulation and Nrf2 degradation, which aggravated UV-induced death of RPEs and RGCs. Significantly, Nrf2 shRNA knockdown almost abolished miR-141-mediated cytoprotection against UV in RPEs. These results demonstrate that miR-141 targets Keap1 to activate Nrf2 signaling, which protects RPEs and RGCs from UV radiation. PMID:28061435

  9. Bioactive compounds in green tea leaves attenuate the injury of retinal ganglion RGC-5 cells induced by H2O2 and ultraviolet radiation.

    Science.gov (United States)

    Jin, Jianchang; Ying, Hao; Huang, Meirong; Du, Qizhen

    2015-11-01

    The Chinese commonly believe that tea helps maintain clear vision. This viewpoint has been recorded in Chinese medical books also. The key bioactive compounds in green tea leaves, (-)-epigallocatechin gallate (EGCG), L-theanine (theanine) and caffeine, were investigated for their abilities to attenuate the injury of retinal ganglion cells (RGC-5) induced by H2O2 and ultraviolet radiation. Theanine and caffeine promoted cell growth while concentrations of EGCG greater than 10μg/ml inhibited cell growth. The nine and caffeine both protected RGC-5 cells from injury as well as enhanced their recovery, while EGCG only protected the cells from injury and did not help them to recover. Tea is a unique drink, which is simultaneously enriched with EGCG, theanine and caffeine. The role of these compounds in optic nerve protection may partially explain why some tea drinkers feel enhanced vision.

  10. Assessment of Rod, Cone, and Intrinsically Photosensitive Retinal Ganglion Cell Contributions to the Canine Chromatic Pupillary Response

    Science.gov (United States)

    Yeh, Connie Y.; Koehl, Kristin L.; Harman, Christine D.; Iwabe, Simone; Guzman, José M.; Petersen-Jones, Simon M.; Kardon, Randy H.; Komáromy, András M.

    2017-01-01

    Purpose The purpose of this study was to evaluate a chromatic pupillometry protocol for specific functional assessment of rods, cones, and intrinsically photosensitive retinal ganglion cells (ipRGCs) in dogs. Methods Chromatic pupillometry was tested and compared in 37 dogs in different stages of primary loss of rod, cone, and combined rod/cone and optic nerve function, and in 5 wild-type (WT) dogs. Eyes were stimulated with 1-s flashes of dim (1 cd/m2) and bright (400 cd/m2) blue light (for scotopic conditions) or bright red (400 cd/m2) light with 25-cd/m2 blue background (for photopic conditions). Canine retinal melanopsin/Opn4 was cloned, and its expression was evaluated using real-time quantitative reverse transcription-PCR and immunohistochemistry. Results Mean ± SD percentage of pupil constriction amplitudes induced by scotopic dim blue (scDB), scotopic bright blue (scBB), and photopic bright red (phBR) lights in WT dogs were 21.3% ± 10.6%, 50.0% ± 17.5%, and 19.4% ± 7.4%, respectively. Melanopsin-mediated responses to scBB persisted for several minutes (7.7 ± 4.6 min) after stimulus offset. In dogs with inherited retinal degeneration, loss of rod function resulted in absent scDB responses, followed by decreased phBR responses with disease progression and loss of cone function. Primary loss of cone function abolished phBR responses but preserved those responses to blue light (scDB and scBB). Although melanopsin/Opn4 expression was diminished with retinal degeneration, melanopsin-expressing ipRGCs were identified for the first time in both WT and degenerated canine retinas. Conclusions Pupil responses elicited by light stimuli of different colors and intensities allowed differential functional assessment of canine rods, cones, and ipRGCs. Chromatic pupillometry offers an effective tool for diagnosing retinal and optic nerve diseases. PMID:28061512

  11. [The neuroprotective effect of erigeron breviscapus (vant) hand-mazz on retinal ganglion cells after optic nerve crush injury].

    Science.gov (United States)

    Jiang, Bing; Jiang, You-qin

    2003-08-01

    To investigate whether a Chinese herbal medicine, erigeron breviscapus (vant) hand-mazz (EBHM), can protect the retinal ganglion cells (RGC) damaged by calibrated optic nerve crush injury. Forty-two Sprague-Dawley rats were randomly divided into two groups. Calibrated optic nerve crush injury model was induced in the right eyes by a special designed optic nerve clip. The left eyes served as a control. All 42 rats were randomly divided into 2 groups. Group A consisted of the rats with calibrated optic nerve crush injury and group B consisted of rats with calibrated optic nerve crush injury treated with EBHM. In group B, EBHM solution was given once after the crush injury. According to the time interval between the optic nerve crush and the sacrifice, both groups A and B were further divided into three subgroups (day 4, day 14 and day 21). Therefore, there were 7 rats in each subgroup. Three days before sacrifice, 3% fast blue was injected into superior colliculi bilaterally. The eyes were enucleated after the rat was sacrificed, and flat mounts of the retina from both eyes were prepared on a slide and observed under a fluorescence microscope. Four photos with 400 x magnification were taken from each of the four quadrants of the retina 1 mm away from the optic disc. The labeled RGC were counted by a computerized image analyzer. The labeled RGC rate was used for statistical analysis (the labeled RGC rate = number of RGC in injured eye/control eye x 100%). In group A, the labeled RGC rate was (77.79 +/- 7.11)%, (63.76 +/- 3.79)% and (54.66 +/- 4.75)% on day 4, day 14 and day 21, respectively. In group B, the labeled RGC rate was (80.13 +/- 12.03)%, (78.17 +/- 9.19)% and (83.59 +/- 12.61)% on day 4, day 14 and day 21, respectively. In group B, which was treated with EBHM after injury, the labeled RGC rate was significantly higher than that of group A on day 14 and day 21. In the experimental optic nerve crush model in rats, EBHM therapy can increase the survival rate of

  12. ANALYSIS OF THE STRUCTURE OF MAGNETIC FIELDS THAT INDUCED INHIBITION OF STIMULATED NEURITE OUTGROWTH

    Science.gov (United States)

    The important experiments showing nonlinear amplitude dependences of the neurite outgrowth in pheochromocytoma nerve cells due to ELF magnetic field exposure had been carried out in a nonuniform ac magnetic field. The nonuniformity entailed larger than expected variances in magne...

  13. Paxillin facilitates timely neurite initiation on soft-substrate environments by interacting with the endocytic machinery.

    Science.gov (United States)

    Chang, Ting-Ya; Chen, Chen; Lee, Min; Chang, Ya-Chu; Lu, Chi-Huan; Lu, Shao-Tzu; Wang, De-Yao; Wang, Aijun; Guo, Chin-Lin; Cheng, Pei-Lin

    2017-12-22

    Neurite initiation is the first step in neuronal development and occurs spontaneously in soft tissue environments. Although the mechanisms regulating the morphology of migratory cells on rigid substrates in cell culture are widely known, how soft environments modulate neurite initiation remains elusive. Using hydrogel cultures, pharmacologic inhibition, and genetic approaches, we reveal that paxillin-linked endocytosis and adhesion are components of a bistable switch controlling neurite initiation in a substrate modulus-dependent manner. On soft substrates, most paxillin binds to endocytic factors and facilitates vesicle invagination, elevating neuritogenic Rac1 activity and expression of genes encoding the endocytic machinery. By contrast, on rigid substrates, cells develop extensive adhesions, increase RhoA activity and sequester paxillin from the endocytic machinery, thereby delaying neurite initiation. Our results highlight paxillin as a core molecule in substrate modulus-controlled morphogenesis and define a mechanism whereby neuronal cells respond to environments exhibiting varying mechanical properties.

  14. Analysis of gene expression during neurite outgrowth and regeneration

    Directory of Open Access Journals (Sweden)

    Tai Yu

    2007-11-01

    Full Text Available Abstract Background The ability of a neuron to regenerate functional connections after injury is influenced by both its intrinsic state and also by extrinsic cues in its surroundings. Investigations of the transcriptional changes undergone by neurons during in vivo models of injury and regeneration have revealed many transcripts associated with these processes. Because of the complex milieu of interactions in vivo, these results include not only expression changes directly related to regenerative outgrowth and but also unrelated responses to surrounding cells and signals. In vitro models of neurite outgrowth provide a means to study the intrinsic transcriptional patterns of neurite outgrowth in the absence of extensive extrinsic cues from nearby cells and tissues. Results We have undertaken a genome-wide study of transcriptional activity in embryonic superior cervical ganglia (SCG and dorsal root ganglia (DRG during a time course of neurite outgrowth in vitro. Gene expression observed in these models likely includes both developmental gene expression patterns and regenerative responses to axotomy, which occurs as the result of tissue dissection. Comparison across both models revealed many genes with similar gene expression patterns during neurite outgrowth. These patterns were minimally affected by exposure to the potent inhibitory cue Semaphorin3A, indicating that this extrinsic cue does not exert major effects at the level of nuclear transcription. We also compared our data to several published studies of DRG and SCG gene expression in animal models of regeneration, and found the expression of a large number of genes in common between neurite outgrowth in vitro and regeneration in vivo. Conclusion Many gene expression changes undergone by SCG and DRG during in vitro outgrowth are shared between these two tissue types and in common with in vivo regeneration models. This suggests that the genes identified in this in vitro study may represent new

  15. Characterization of intravitreally delivered capsid mutant AAV2-Cre vector to induce tissue-specific mutations in murine retinal ganglion cells.

    Science.gov (United States)

    Langouet-Astrie, Christophe J; Yang, Zhiyong; Polisetti, Sraavya M; Welsbie, Derek S; Hauswirth, William W; Zack, Donald J; Merbs, Shannath L; Enke, Raymond A

    2016-10-01

    Targeted expression of Cre recombinase in murine retinal ganglion cells (RGCs) by viral vector is an effective strategy for creating tissue-specific gene knockouts for investigation of genetic contribution to RGC degeneration associated with optic neuropathies. Here we characterize dosage, efficacy and toxicity for sufficient intravitreal delivery of a capsid mutant Adeno-associated virus 2 (AAV2) vector encoding Cre recombinase. Wild type and Rosa26 (R26) LacZ mice were intravitreally injected with capsid mutant AAV2 viral vectors. Murine eyes were harvested at intervals ranging from 2 weeks to 15 weeks post-injection and were assayed for viral transduction, transgene expression and RGC survival. 10(9) vector genomes (vg) were sufficient for effective in vivo targeting of murine ganglion cell layer (GCL) retinal neurons. Transgene expression was observed as early as 2 weeks post-injection of viral vectors and persisted to 11 weeks. Early expression of Cre had no significant effect on RGC survival, while significant RGC loss was detected beginning 5 weeks post-injection. Early expression of viral Cre recombinase was robust, well-tolerated and predominantly found in GCL neurons suggesting this strategy can be effective in short-term RGC-specific mutation studies in experimental glaucoma models such as optic nerve crush and transection experiments. RGC degeneration with Cre expression for more than 4 weeks suggests that Cre toxicity is a limiting factor for targeted mutation strategies in RGCs. Copyright © 2016 Elsevier Ltd. All rights reserved.

  16. Age-dependent neuroprotection of retinal ganglion cells by tempol-C8 acyl ester in a rat NMDA toxicity model.

    Science.gov (United States)

    Fiedorowicz, Michal; Rejdak, Robert; Schuettauf, Frank; Wozniak, Michal; Grieb, Pawel; Thaler, Sebastian

    2014-01-01

    The efficacy of tempol and its acyl derivative tempol-C8 as retinoprotective agents was compared in a rat model of NMDA-induced retinal ganglion cell (RGC) damage. Tempol or tempol-C8 in different doses was administered intraperitoneally to 6 weeks old (pre-adolescent) and 9-10 weeks old (young adult) rats before and after an intravitreous NMDA injection. Retinal ganglion cell were retrogradely labeled with the fluorescent tracer hydroxystilbamidine and RGC counting was performed on retinal flatmounts. Intravitreal NMDA reduced RGC counts by about 90%, independently of age tempol-C8, but not tempol unmodified, showed a significant, dose-dependent RGC rescue effect, with peak activity at 5.8 µmol/kg (p tempol or tempol-C8. In contrast to tempol itself, tempol-C8 acyl ester was neuroprotective in pre-adolescent rats in the NMDA- induced RGC damage model. Therefore, neuroprotection by tempol acyl esters seems to be superior to that of tempol under certain conditions.

  17. Variations of retinal nerve fiber layer thickness and ganglion cell-inner plexiform layer thickness according to the torsion direction of optic disc.

    Science.gov (United States)

    Lee, Kang Hoon; Kim, Chan Yun; Kim, Na Rae

    2014-02-20

    To examine the relationship between the optic disc torsion and peripapillary retinal nerve fiber layer (RNFL) thickness through a comparison with the macular ganglion cell inner plexiform layer complex (GCIPL) thickness measured by Cirrus optical coherence tomography (OCT). Ninety-four eyes of 94 subjects with optic disc torsion and 114 eyes of 114 subjects without optic disc torsion were enrolled prospectively. The participants underwent fundus photography and OCT imaging in peripapillary RNFL mode and macular GCIPL mode. The participants were divided into groups according to the presence or absence of optic disc torsion. The eyes with optic disc torsion were further divided into supranasal torsion and inferotemporal torsion groups according to the direction of optic disc torsion. The mean RNFL and GCIPL thicknesses for the quadrants and subsectors were compared. The superior and inferior peak locations of the RNFL were also measured according to the torsion direction. The temporal RNFL thickness was significantly thicker in inferotemporal torsion, whereas the GCIPL thickness at all segments was unaffected. The inferotemporal optic torsion had more temporally positioned superior peak locations of the RNFL than the nontorsion and supranasal-torted optic disc. Thickening of the temporal RNFL with a temporal shift in the superior peak within the eyes with inferotemporal optic disc torsion can lead to interpretation errors. The ganglion cell analysis algorithm can assist in differentiating eyes with optic disc torsion.

  18. Vesicular glutamate transporter 2 (VGLUT2) is co-stored with PACAP in projections from the rat melanopsin-containing retinal ganglion cells

    DEFF Research Database (Denmark)

    Engelund, Anna Iversen; Fahrenkrug, Jan; Harrison, Adrian Paul

    2010-01-01

    transcardially perfusion-fixated, after which the brains and eyes were removed for double immunohistochemical staining using a polyclonal anti-VGLUT2 antibody and a mouse monoclonal anti-PACAP antibody. Results revealed that VGLUT2- and PACAP-immunoreactivity (-ir) were present in ipRGCs and co......The retinal ganglion cell layer of the eye comprises a subtype of cells characterized by their intrinsic photosensitivity and expression of melanopsin (ipRGCs). These cells regulate a variety of non-image-forming (NIF) functions such as light entrainment of circadian rhythms, acute suppression......-localization of vesicular glutamate transporter 2 (VGLUT2; a marker of glutamate signaling) and PACAP in ipRGCs and their projections in the brain. Nine adult male Wistar rats were assigned to one of three groups; anterograde tracing (n = 3), eye enucleation (n = 3), and untreated (n = 3). Under anaesthesia, rats were...

  19. Intracerebroventricular gene therapy that delays neurological disease progression is associated with selective preservation of retinal ganglion cells in a canine model of CLN2 disease.

    Science.gov (United States)

    Whiting, Rebecca E H; Jensen, Cheryl A; Pearce, Jacqueline W; Gillespie, Lauren E; Bristow, Daniel E; Katz, Martin L

    2016-05-01

    CLN2 disease is one of a group of lysosomal storage disorders called the neuronal ceroid lipofuscinoses (NCLs). The disease results from mutations in the TPP1 gene that cause an insufficiency or complete lack of the soluble lysosomal enzyme tripeptidyl peptidase-1 (TPP1). TPP1 is involved in lysosomal protein degradation, and lack of this enzyme results in the accumulation of protein-rich autofluorescent lysosomal storage bodies in numerous cell types including neurons throughout the central nervous system and the retina. CLN2 disease is characterized primarily by progressive loss of neurological functions and vision as well as generalized neurodegeneration and retinal degeneration. In children the progressive loss of neurological functions typically results in death by the early teenage years. A Dachshund model of CLN2 disease with a null mutation in TPP1 closely recapitulates the human disorder with a progression from disease onset at approximately 4 months of age to end-stage at 10-11 months. Delivery of functional TPP1 to the cerebrospinal fluid (CSF), either by periodic infusion of the recombinant protein or by a single administration of a TPP1 gene therapy vector to the CSF, significantly delays the onset and progression of neurological signs and prolongs life span but does not prevent the loss of vision or modest retinal degeneration that occurs by 11 months of age. In this study we found that in dogs that received the CSF gene therapy treatment, the degeneration of the retina and loss of retinal function continued to progress during the prolonged life spans of the treated dogs. Eventually the normal cell layers of the retina almost completely disappeared. An exception was the ganglion cell layer. In affected dogs that received TPP1 gene therapy to the CSF and survived an average of 80 weeks, ganglion cell axons were present in numbers comparable to those of normal Dachshunds of similar age. The selective preservation of the retinal ganglion cells suggests

  20. Neuroprotection by α2-Adrenergic Receptor Stimulation after Excitotoxic Retinal Injury: A Study of the Total Population of Retinal Ganglion Cells and Their Distribution in the Chicken Retina.

    Science.gov (United States)

    Galindo-Romero, Caridad; Harun-Or-Rashid, Mohammad; Jiménez-López, Manuel; Vidal-Sanz, Manuel; Agudo-Barriuso, Marta; Hallböök, Finn

    2016-01-01

    We have studied the effect of α2-adrenergic receptor stimulation on the total excitotoxically injured chicken retinal ganglion cell population. N-methyl-D-aspartate (NMDA) was intraocularly injected at embryonic day 18 and Brn3a positive retinal ganglion cells (Brn3a+ RGCs) were counted in flat-mounted retinas using automated routines. The number and distribution of the Brn3a+ RGCs were analyzed in series of normal retinas from embryonic day 8 to post-hatch day 11 retinas and in retinas 7 or 14 days post NMDA lesion. The total number of Brn3a+ RGCs in the post-hatch retina was approximately 1.9x106 with a density of approximately 9.2x103 cells/mm2. The isodensity maps of normal retina showed that the density decreased with age as the retinal size increased. In contrast to previous studies, we did not find any specific region with increased RGC density, rather the Brn3a+ RGCs were homogeneously distributed over the central retina with decreasing density in the periphery and in the region of the pecten oculli. Injection of 5-10 μg NMDA caused 30-50% loss of Brn3a+ cells and the loss was more severe in the dorsal than in the ventral retina. Pretreatment with brimonidine reduced the loss of Brn3a+ cells both 7 and 14 days post lesion and the protective effect was higher in the dorsal than in the ventral retina. We conclude that α2-adrenergic receptor stimulation reduced the impact of the excitotoxic injury in chicken similarly to what has been shown in mammals. Furthermore, the data show that the RGCs are evenly distributed over in the retina, which challenges previous results that indicate the presence of specific high RGC-density regions of the chicken retina.

  1. Neuroprotection by α2-Adrenergic Receptor Stimulation after Excitotoxic Retinal Injury: A Study of the Total Population of Retinal Ganglion Cells and Their Distribution in the Chicken Retina

    Science.gov (United States)

    Galindo-Romero, Caridad; Harun-Or-Rashid, Mohammad; Jiménez-López, Manuel; Vidal-Sanz, Manuel; Agudo-Barriuso, Marta

    2016-01-01

    We have studied the effect of α2-adrenergic receptor stimulation on the total excitotoxically injured chicken retinal ganglion cell population. N-methyl-D-aspartate (NMDA) was intraocularly injected at embryonic day 18 and Brn3a positive retinal ganglion cells (Brn3a+ RGCs) were counted in flat-mounted retinas using automated routines. The number and distribution of the Brn3a+ RGCs were analyzed in series of normal retinas from embryonic day 8 to post-hatch day 11 retinas and in retinas 7 or 14 days post NMDA lesion. The total number of Brn3a+ RGCs in the post-hatch retina was approximately 1.9x106 with a density of approximately 9.2x103 cells/mm2. The isodensity maps of normal retina showed that the density decreased with age as the retinal size increased. In contrast to previous studies, we did not find any specific region with increased RGC density, rather the Brn3a+ RGCs were homogeneously distributed over the central retina with decreasing density in the periphery and in the region of the pecten oculli. Injection of 5–10 μg NMDA caused 30–50% loss of Brn3a+ cells and the loss was more severe in the dorsal than in the ventral retina. Pretreatment with brimonidine reduced the loss of Brn3a+ cells both 7 and 14 days post lesion and the protective effect was higher in the dorsal than in the ventral retina. We conclude that α2-adrenergic receptor stimulation reduced the impact of the excitotoxic injury in chicken similarly to what has been shown in mammals. Furthermore, the data show that the RGCs are evenly distributed over in the retina, which challenges previous results that indicate the presence of specific high RGC-density regions of the chicken retina. PMID:27611432

  2. The dark phase intraocular pressure elevation and retinal ganglion cell degeneration in a rat model of experimental glaucoma☆

    Science.gov (United States)

    Kwong, Jacky M.K.; Vo, Nancy; Quan, Ann; Nam, Michael; Kyung, Haksu; Yu, Fei; Piri, Natik; Caprioli, Joseph

    2013-01-01

    Intraocular pressure (IOP) elevation is considered as a major risk factor causing the progression of vision deterioration in glaucoma. Although it is known that the IOP level changes widely throughout the day and night, how the dark or light phase IOP elevation contributes to retinal ganglion cell (RGC) degeneration is still largely unclear. To examine the profile of IOP, modified laser photocoagulation was applied to the trabecular meshwork of Brown Norway rats and both light and dark phase IOPs were monitored approximately 1–2 times a week. The relationship between IOP elevation and RGC degeneration was investigated while RGC body loss was analyzed with Rbpms immunolabeling on retinal wholemount and axonal injury in the optic nerve was semi-quantified. The baseline awake dark and light IOPs were 30.4 ± 2.7 and 20.2 ± 2.1 mmHg respectively. The average dark IOP was increased to 38.2 ± 3.2 mmHg for five weeks after the laser treatment on 270° trabecular meshwork. However, there was no significant loss of RGC body and axonal injury. After laser treatment on 330° trabecular meshwork, the dark and light IOPs were significantly increased to 43.8 ± 4.6 and 23 ± 3.7 mmHg respectively for 5 weeks. The cumulative dark and light IOP elevations were 277 ± 86 and 113 ± 50 mmHg days respectively while the cumulative total (light and dark) IOP elevation was 213 ± 114 mmHg days. After 5 weeks, regional RGC body loss of 29.5 ± 15.5% and moderate axonal injury were observed. Axonal injury and loss of RGC body had a high correlation with the cumulative total IOP elevation (R2 = 0.60 and 0.65 respectively). There was an association between the cumulative dark IOP elevation and RGC body loss (R2 = 0.37) and axonal injury (R2 = 0.51) whereas the associations between neuronal damages and the cumulative light IOP elevation were weak (for RGC body loss, R2 = 0.01; for axonal injury, R2 = 0.26). Simple linear regression model analysis showed statistical significance for the

  3. Neuronal Survival, Morphology and Outgrowth of Spiral Ganglion Neurons Using a Defined Growth Factor Combination.

    Directory of Open Access Journals (Sweden)

    Jana Schwieger

    Full Text Available The functionality of cochlear implants (CI depends, among others, on the number and excitability of surviving spiral ganglion neurons (SGN. The spatial separation between the SGN, located in the bony axis of the inner ear, and the CI, which is inserted in the scala tympani, results in suboptimal performance of CI patients and may be decreased by attracting the SGN neurites towards the electrode contacts. Neurotrophic factors (NTFs can support neuronal survival and neurite outgrowth.Since brain-derived neurotrophic factor (BDNF is well known for its neuroprotective effect and ciliary neurotrophic factor (CNTF increases neurite outgrowth, we evaluated if the combination of BDNF and CNTF leads to an enhanced neuronal survival with extended neurite outgrowth. Both NTFs were added in effective high concentrations (BDNF 50 ng/ml, CNTF 100 ng/ml, alone and in combination, to cultured dissociated SGN of neonatal rats for 48 hours.The neuronal survival and neurite outgrowth were significantly higher in SGN treated with the combination of the two NTFs compared to treatment with each factor alone. Additionally, with respect to the morphology, the combination of BDNF and CNTF leads to a significantly higher number of bipolar neurons and a decreased number of neurons without neurites in culture.The combination of BDNF and CNTF shows a great potential to increase the neuronal survival and the number of bipolar neurons in vitro and to regenerate retracted nerve fibers.

  4. Unbiased estimates of number and size of rat dorsal root ganglion cells in studies of structure and cell survival

    DEFF Research Database (Denmark)

    Lamm, Trine Tandrup

    Neurodegenerative sygdomme er karakteriseret ved tab af nervefibre og nervecellelegemer. Tilstande med fysiske eller toksikologiske beskadigelser af de primære sensoriske nerveceller hos rotten har ofte været anvendt som model for forståelse af de processer, der fører til celledød eller -overleve...

  5. Neurite outgrowth on fluorinated polyimide film micropatterned by ion irradiation

    Energy Technology Data Exchange (ETDEWEB)

    Okuyama, Y.; Sato, M.; Nagaoka, S.; Kawakami, H. E-mail: kawakami-hiroyoshi@c.metro-u.ac.jp; Suzuki, Y.; Iwaki, M

    2003-05-01

    In this study, we investigated neurite outgrowth on a fluorinated polyimide film micropatterned by ion irradiation. We used the fluorinated polyimide because of its excellent thermal and mechanical properties and biocompatibility. Rattus norvegicus chromaphin (PC12) cells were used for in vitro studies. The polyimide films were irradiated with He{sup +}, Ne{sup +} or Kr{sup +} at 1 x 10{sup 14} ions/cm{sup 2} using an ion-beam mask. The lines in the mask were 120 and 160 {mu}m wide and 120-160 {mu}m apart. PC12 cells were selectively adhered on the polyimide film micropatterned by Kr{sup +}-irradiation. However, the neurite length on the film irradiated by Kr{sup +} was shorter than that determined in the film irradiated by He{sup +}. On the other hand, neurite outgrowth on the polyimide film micropatterned by He{sup +}-irradiation was at least 100 {mu}m in length. This initial study indicated the enhanced outgrowth of PC12 cells on the fluorinated polyimide film micropatterned by ion irradiation.

  6. No dramatic age-related loss of hair cells and spiral ganglion neurons in Bcl-2 over-expression mice or Bax null mice

    Directory of Open Access Journals (Sweden)

    Ohlemiller Kevin K

    2010-07-01

    Full Text Available Abstract Age-related decline of neuronal function is associated with age-related structural changes. In the central nervous system, age-related decline of cognitive performance is thought to be caused by synaptic loss instead of neuronal loss. However, in the cochlea, age-related loss of hair cells and spiral ganglion neurons (SGNs is consistently observed in a variety of species, including humans. Since age-related loss of these cells is a major contributing factor to presbycusis, it is important to study possible molecular mechanisms underlying this age-related cell death. Previous studies suggested that apoptotic pathways were involved in age-related loss of hair cells and SGNs. In the present study, we examined the role of Bcl-2 gene in age-related hearing loss. In one transgenic mouse line over-expressing human Bcl-2, there were no significant differences between transgenic mice and wild type littermate controls in their hearing thresholds during aging. Histological analysis of the hair cells and SGNs showed no significant conservation of these cells in transgenic animals compared to the wild type controls during aging. These data suggest that Bcl-2 overexpression has no significant effect on age-related loss of hair cells and SGNs. We also found no delay of age-related hearing loss in mice lacking Bax gene. These findings suggest that age-related hearing loss is not through an apoptotic pathway involving key members of Bcl-2 family.

  7. Evaluation of Macular Ganglion Cell-inner Plexiform Layer and Choroid in Psoriasis Patients Using Enhanced Depth Imaging Spectral Domain Optical Coherence Tomography.

    Science.gov (United States)

    Ersan, Ismail; Kilic, Sevilay; Arikan, Sedat; Kara, Selcuk; Işik, Selda; Gencer, Baran; Ogretmen, Zerrin

    2017-08-01

    To evaluate changes in the thickness of the central macula, macular ganglion cell-inner plexiform layer (mGCIPL), and subfoveal choroid in patients with psoriasis using spectral domain optical coherence tomography (SD-OCT). The measurements of macular, mGCIPL thicknesses and subfoveal choroidal thickness (SFCT) obtained by SD-OCT of psoriasis patients (n = 46). These measurements were compared with those of 50 healthy controls. The macular, mGCIPL, and choroidal thicknesses did not differ between the controls and psoriatic subjects (p>0.05). When the patients were divided into two distinct groups, only the SFCT was significantly thicker in the severe psoriasis group compared with the mild psoriasis group (p = 0.003). These findings suggest that choroidal alterations are seen without macular changes in patients with psoriasis. Severe psoriasis appears to be related to increases in SFCT as a consequence of possible inflammatory cascades that are part of the disease's pathogenesis.

  8. Chronic, low-dose rotenone reproduces Lewy neurites found in early stages of Parkinson's disease, reduces mitochondrial movement and slowly kills differentiated SH-SY5Y neural cells

    Directory of Open Access Journals (Sweden)

    Liu Lei

    2008-12-01

    Full Text Available Abstract Background Parkinson's disease, the most common adult neurodegenerative movement disorder, demonstrates a brain-wide pathology that begins pre-clinically with alpha-synuclein aggregates ("Lewy neurites" in processes of gut enteric and vagal motor neurons. Rostral progression into substantia nigra with death of dopamine neurons produces the motor impairment phenotype that yields a clinical diagnosis. The vast majority of Parkinson's disease occurs sporadically, and current models of sporadic Parkinson's disease (sPD can utilize directly infused or systemic neurotoxins. Results We developed a differentiation protocol for human SH-SY5Y neuroblastoma that yielded non-dividing dopaminergic neural cells with long processes that we then exposed to 50 nM rotenone, a complex I inhibitor used in Parkinson's disease models. After 21 days of rotenone, ~60% of cells died. Their processes retracted and accumulated ASYN-(+ and UB-(+ aggregates that blocked organelle transport. Mitochondrial movement velocities were reduced by 8 days of rotenone and continued to decline over time. No cytoplasmic inclusions resembling Lewy bodies were observed. Gene microarray analyses showed that the majority of genes were under-expressed. qPCR analyses of 11 mtDNA-encoded and 10 nDNA-encoded mitochondrial electron transport chain RNAs' relative expressions revealed small increases in mtDNA-encoded genes and lesser regulation of nDNA-encoded ETC genes. Conclusion Subacute rotenone treatment of differentiated SH-SY5Y neuroblastoma cells causes process retraction and partial death over several weeks, slowed mitochondrial movement in processes and appears to reproduce the Lewy neuritic changes of early Parkinson's disease pathology but does not cause Lewy body inclusions. The overall pattern of transcriptional regulation is gene under-expression with minimal regulation of ETC genes in spite of rotenone's being a complex I toxin. This rotenone-SH-SY5Y model in a

  9. The meniscus ganglion

    International Nuclear Information System (INIS)

    Schaefer, H.

    1982-01-01

    Normal dimensions of the meniscus quoted in the literature vary somewhat; measurements were therefore carried out on the height and width on standardised arthrograms. This made it possible to evaluate changes in the height of the meniscus objectively and to diagnose degeneration with a gang