Serial Editor Vincent Walsh Institute of Cognitive Neuroscience University College London 17 Queen Square London WC1N 3AR UK Editorial Board Mark Bear, Cambridge, USA Medicine & Translational Neuroscience Hamed Ekhtiari, Tehran, Iran Addiction Hajime Hirase, Wako, Japan Neuronal Microcircuitry Freda Miller, Toronto, Canada Developmental Neurobiology Shane O’Mara, Dublin, Ireland Systems Neuroscience Susan Rossell, Swinburne, Australia Clinical Psychology & Neuropsychiatry Nathalie Rouach, Paris, France Neuroglia Barbara Sahakian, Cambridge, UK Cognition & Neuroethics Bettina Studer, Dusseldorf, Germany Neurorehabilitation Xiao-Jing Wang, New York, USA Computational Neuroscience Elsevier Radarweg 29, PO Box 211, 1000 AE Amsterdam, Netherlands The Boulevard, Langford Lane, Kidlington, Oxford OX5 1GB, UK 225 Wyman Street, Waltham, MA 02451, USA First edition 2015 Copyright # 2015 Elsevier B.V All rights reserved No part of this publication may be reproduced or transmitted in any form or by any means, electronic or mechanical, including photocopying, recording, or any information storage and retrieval system, without permission in writing from the publisher Details on how to seek permission, further information about the Publisher’s permissions policies and our arrangements with organizations such as the Copyright Clearance Center and the Copyright Licensing Agency, can be found at our website: www.elsevier.com/permissions This book and the individual contributions contained in it are protected under copyright by the Publisher (other than as may be noted herein) Notices Knowledge and best practice in this field are constantly changing As new research and experience broaden our understanding, changes in research methods, professional practices, or medical treatment may become necessary Practitioners and researchers must always rely on their own experience and knowledge in evaluating and using any information, methods, compounds, or experiments described herein In using such information or methods they should be mindful of their own safety and the safety of others, including parties for whom they have a professional responsibility To the fullest extent of the law, neither the Publisher nor the authors, contributors, or editors, assume any liability for any injury and/or damage to persons or property as a matter of products liability, negligence or otherwise, or from any use or operation of any methods, products, instructions, or ideas contained in the material herein ISBN: 978-0-444-63566-2 ISSN: 0079-6123 For information on all Elsevier publications visit our website at store.elsevier.com Contributors Jessica Agostinone Department of Neuroscience, and Centre de Recherche du Centre Hospitalier de l’Universite´ de Montre´al (CRCHUM), University of Montreal, Montreal, QC, Canada Marta Agudo-Barriuso Departamento de Oftalmologı´a, Universidad de Murcia and Instituto Murciano de Investigacio´n Biosanitaria Virgen de la Arrixaca (IMIB-Arrixaca), Murcia, Spain Luis Alarco´n-Martı´nez Departamento de Oftalmologı´a, Universidad de Murcia and Instituto Murciano de Investigacio´n Biosanitaria Virgen de la Arrixaca (IMIB-Arrixaca), Murcia, Spain Marcelino Avile´s-Trigueros Departamento de Oftalmologı´a, Universidad de Murcia and Instituto Murciano de Investigacio´n Biosanitaria Virgen de la Arrixaca (IMIB-Arrixaca), Murcia, Spain Giacinto Bagetta Department of Pharmacy and Health and Nutritional Sciences, Section of Preclinical and Translational Pharmacology, University of Calabria, Arcavacata di Rende, Italy; University Consortium for Adaptive Disorders and Head Pain (UCHAD), Section of Neuropharmacology of Normal and Pathological Neuronal Plasticity, University of Calabria, Arcavacata di Rende, Italy Claudio Bucolo Department of Biomedical and Biotechnological Sciences, Section of Pharmacology, University of Catania, Catania, Italy Karolien Castermans Amakem Therapeutics, Diepenbeek, Belgium Shenton S.L Chew NIHR Biomedical Research Centre at Moorfields Eye Hospital NHS Foundation Trust and UCL Institute of Ophthalmology, London, UK Maria Tiziana Corasaniti Department of Health Sciences, University “Magna Graecia” of Catanzaro, Catanzaro, Italy Rosa de Hoz Instituto de Investigaciones Oftalmolo´gicas Ramo´n Castroviejo, Facultad de O´ptica y Optometrı´a, Universidad Complutense de Madrid, Spain Adriana Di Polo Department of Neuroscience, and Centre de Recherche du Centre Hospitalier de l’Universite´ de Montre´al (CRCHUM), University of Montreal, Montreal, QC, Canada v vi Contributors Filippo Drago Department of Biomedical and Biotechnological Sciences, Section of Pharmacology, University of Catania, Catania, Italy Stefano Forte IOM Ricerca srl, Catania, Italy Beatriz I Gallego Instituto de Investigaciones Oftalmolo´gicas Ramo´n Castroviejo, Universidad Complutense de Madrid, Spain Roberto Gallego-Pinazo Ophthalmic Research Unit “Santiago Grisolı´a”, University Hospital Dr Peset, and Department of Ophthalmology, University and Polytechnic Hospital la Fe, Valencia, Spain Diego Garcı´a-Ayuso Departamento de Oftalmologı´a, Universidad de Murcia and Instituto Murciano de Investigacio´n Biosanitaria Virgen de la Arrixaca (IMIB-Arrixaca), Murcia, Spain Jose´ J Garcı´a-Medina Ophthalmic Research Unit “Santiago Grisolı´a”, University Hospital Dr Peset, Valencia; Department of Ophthalmology, University Hospital Reina Sofia, and Department of Ophthalmology and Optometry, University of Murcia, Murcia, Spain Neeru Gupta Department of Ophthalmology and Vision Sciences; Department of Laboratory Medicine and Pathobiology, University of Toronto; Keenan Research Centre for Biomedical Science, and Glaucoma and Nerve Protection Unit, St Michael’s Hospital, Toronto, ON, Canada Manuel Jime´nez-Lo´pez Departamento de Oftalmologı´a, Universidad de Murcia and Instituto Murciano de Investigacio´n Biosanitaria Virgen de la Arrixaca (IMIB-Arrixaca), Murcia, Spain Nele Kindt Amakem Therapeutics, Diepenbeek, Belgium Hani Levkovitch-Verbin Glaucoma Service, Goldschleger Eye Institute, Sheba Medical Center, and Sackler Faculty of Medicine, Tel-Aviv University, Tel-Hashomer, Israel Fumihiko Mabuchi Department of Ophthalmology, Faculty of Medicine, University of Yamanashi, Chuo, Yamanashi, Japan Keith Martin John van Geest Centre for Brain Repair, University of Cambridge; Cambridge NIHR Biomedical Research Centre, and Wellcome Trust Medical Research Council Cambridge Stem Cell Institute, Cambridge, UK Contributors Alessandra Martins Discipline of Clinical Ophthalmology and Eye Health, University of Sydney, and Sydney Eye Hospital, Sydney, NSW, Australia Lieve Moons Research Group of Neural Circuit Development and Regeneration, KU Leuven, Leuven, Belgium Luigi Antonio Morrone Department of Pharmacy and Health and Nutritional Sciences, Section of Preclinical and Translational Pharmacology, University of Calabria, Arcavacata di Rende, Italy; University Consortium for Adaptive Disorders and Head Pain (UCHAD), Section of Neuropharmacology of Normal and Pathological Neuronal Plasticity, University of Calabria, Arcavacata di Rende, Italy Francisco M Nadal-Nicola´s Departamento de Oftalmologı´a, Universidad de Murcia and Instituto Murciano de Investigacio´n Biosanitaria Virgen de la Arrixaca (IMIB-Arrixaca), Murcia, Spain Carlo Nucci Ophthalmology Unit, Department of Experimental Medicine and Surgery, University of Rome Tor Vergata, Rome, Italy Arturo Ortı´n-Martı´nez Departamento de Oftalmologı´a, Universidad de Murcia and Instituto Murciano de Investigacio´n Biosanitaria Virgen de la Arrixaca (IMIB-Arrixaca), Murcia, Spain Craig Pearson John van Geest Centre for Brain Repair, University of Cambridge; Cambridge NIHR Biomedical Research Centre, Cambridge, UK, and National Heart, Lung and Blood Institute, National Institutes of Health, Bethesda, MD, USA Maria D Pinazo-Dura´n Ophthalmic Research Unit “Santiago Grisolı´a”, University Hospital Dr Peset, and Department of Surgery/Ophthalmology, Faculty of Medicine and Odontology, University of Valencia, Valencia, Spain Chiara Bianca Maria Platania Department of Biomedical and Biotechnological Sciences, Section of Pharmacology, University of Catania, Catania, Italy Harry A Quigley Glaucoma Center of Excellence, Wilmer Institute, Johns Hopkins University School of Medicine, Baltimore, MD, USA Ana I Ramı´rez Instituto de Investigaciones Oftalmolo´gicas Ramo´n Castroviejo, Facultad de O´ptica y Optometrı´a, Universidad Complutense de Madrid, Spain Jose´ M Ramirez Instituto de Investigaciones Oftalmolo´gicas Ramo´n Castroviejo, Departamento de Oftalmologı´a y ORL, Facultad de Medicina, Universidad Complutense de Madrid, Spain vii viii Contributors Blanca Rojas Instituto de Investigaciones Oftalmolo´gicas Ramo´n Castroviejo, Departamento de Oftalmologı´a y ORL, Facultad de Medicina, Universidad Complutense de Madrid, Spain Giovanni Luca Romano Department of Biomedical and Biotechnological Sciences, Section of Pharmacology, University of Catania, Catania, Italy Laura Rombola` Department of Pharmacy, Health and Nutritional Sciences, Section of Preclinical and Translational Pharmacology, University of Calabria, Cosenza, Italy Rossella Russo Department of Pharmacy, Health and Nutritional Sciences, Section of Preclinical and Translational Pharmacology, University of Calabria, Cosenza, Italy Yoichi Sakurada Department of Ophthalmology, Faculty of Medicine, University of Yamanashi, Chuo, Yamanashi, Japan Juan J Salazar Instituto de Investigaciones Oftalmolo´gicas Ramo´n Castroviejo, Facultad de O´ptica y Optometrı´a, Universidad Complutense de Madrid, Spain Manuel Salinas-Navarro Departamento de Oftalmologı´a, Universidad de Murcia and Instituto Murciano de Investigacio´n Biosanitaria Virgen de la Arrixaca (IMIB-Arrixaca), Murcia, Spain Elena Salobrar-Garcı´a Instituto de Investigaciones Oftalmolo´gicas Ramo´n Castroviejo, Universidad Complutense de Madrid, Spain Salvatore Salomone Department of Biomedical and Biotechnological Sciences, Section of Pharmacology, University of Catania, Catania, Italy Ingeborg Stalmans Laboratory of Ophthalmology, KU Leuven, and Department of Ophthalmology, University Hospitals Leuven (UZ Leuven), Leuven, Belgium Nicholas Strouthidis NIHR Biomedical Research Centre at Moorfields Eye Hospital NHS Foundation Trust and UCL Institute of Ophthalmology, London, UK; Discipline of Clinical Ophthalmology and Eye Health, University of Sydney, Sydney, NSW, Australia, and Singapore Eye Research Institute, Singapore, Singapore Alberto Trivin˜o Instituto de Investigaciones Oftalmolo´gicas Ramo´n Castroviejo, Departamento de Oftalmologı´a y ORL, Facultad de Medicina, Universidad Complutense de Madrid, Spain Contributors Francisco J Valiente-Soriano Laboratorio de Oftalmologı´a Experimental, Departamento de Oftalmologı´a, Facultad de Medicina, Universidad de Murcia, and Departamento de Oftalmologı´a, Universidad de Murcia and Instituto Murciano de Investigacio´n Biosanitaria Virgen de la Arrixaca (IMIB-Arrixaca), Murcia, Spain Tine Van Bergen Laboratory of Ophthalmology, KU Leuven, Leuven, Belgium Sarah Van de Velde Laboratory of Ophthalmology, KU Leuven, Leuven, Belgium Evelien Vandewalle Laboratory of Ophthalmology, KU Leuven, and Department of Ophthalmology, University Hospitals Leuven (UZ Leuven), Leuven, Belgium Manuel Vidal-Sanz Departamento de Oftalmologı´a, Universidad de Murcia and Instituto Murciano de Investigacio´n Biosanitaria Virgen de la Arrixaca (IMIB-Arrixaca), Murcia, Spain Maria P Villegas-Pe´rez Departamento de Oftalmologı´a, Universidad de Murcia and Instituto Murciano de Investigacio´n Biosanitaria Virgen de la Arrixaca (IMIB-Arrixaca), Murcia, Spain Yeni Yucel Department of Ophthalmology and Vision Sciences; Department of Laboratory Medicine and Pathobiology, University of Toronto; Keenan Research Centre for Biomedical Science; Ophthalmic Pathology Laboratory, University of Toronto, St Michael’s Hospital, and Faculty of Engineering & Architectural Science, Ryerson University, Toronto, ON, Canada Vicente Zano´n-Moreno Ophthalmic Research Unit “Santiago Grisolı´a”, University Hospital Dr Peset, and Department of Surgery/Ophthalmology, Faculty of Medicine and Odontology, University of Valencia, Valencia, Spain ix Preface: New Trends in Basic and Clinical Research of Glaucoma: A Neurodegenerative Disease of the Visual System Part A Glaucoma, the second leading cause of blindness in the world, is characterized by progressive retinal ganglion cell (RGC) axons degeneration and death leading to typical optic nerve head damage and distinctive visual field defects This disease is a chronic optic neuropathy most often associated with increased intraocular pressure and age as main risk factors Defective axonal transport, trophic factor withdrawal, and neuroinflammation are emerging as important pathophysiological factors Despite the limited value of the animal models in recapitulating the pathophysiology of the disease, these have allowed determinants involved in RGC apoptosis to be dissected Under conditions of glutamate homeostasis disruption, excitotoxicity ensues and this causes neuronal damage implicating oxidative stress Free radical species accumulation can cause RGC death by inhibition of key enzymes of the tricarboxylic acid cycle, the mitochondrial electron transport chain, and mitochondrial calcium homeostasis, leading to defective energy metabolism Accordingly, in glaucomatous patients a significant decrease in the total antioxidant capacity has been reported along with increased end-products of lipid peroxidation, among other putative markers Several interventions find their rational in the causative role of oxidative stress in RGC death, though these have limited or no clinical proof Experimental data indicate that axonal injury triggers rapid structural alterations in RGC dendritic arbors, prior to manifest axonal loss, leading to synaptic rearrangements and functional deficits Tissue remodeling occurring in glaucoma may cause biomechanical and microstructural changes that are likely to alter the mechanical environment of the optic nerve head and may contribute to axonal damage Indeed, experimental evidence following laser photocoagulation demonstrates that the volume occupied by retinotectal afferents is halved, ocular hypertension affects selectively projecting neurons (e.g., RGC), and intraocular administration of BDNF results in increased RGC survival These data are at variance with changes in other cells/sectors of the retina for the proportion of the cell loss, for its diffuse and not sectorial topography, for it does not respond to BDNF neuroprotection, and for progressive functional and morphological alterations there occur Most of the data in the literature have been gathered employing experimental models of unilateral glaucoma and using the normotensive contralateral eye as the normal control Interestingly, some studies have recently reported the activation xix xx Preface: New trends in basic and clinical research of glaucoma of the retinal macroglia and microglia in the uninjured eye along with important observations implicating innate and adaptive immunity The latter data support a role for blood–retina barrier disruption in the pathophysiology of glaucoma-associated neurodegenerative process other than simply suggesting that the eye contralateral to experimental glaucoma cannot be a true control Experimental data support the hypothesis that autophagy might participate in the process leading to RGC death though the precise role awaits to be clarified In fact, evidence shows that downregulation of autophagy-related genes (Atg5, Atg7, and BECN1) in normal human aging brain has been reported On the contrary, a recent study analyzing LC3 and p62 levels in fresh TM from human donors reported lower levels of p62 and increased LC3II/LC3I ratio in subjects older than 60 years suggesting an age-related upregulation of autophagy in the TM A marked reduction in macroautophagy activity in the aged retina that is associated, in vitro and in vivo, with a sustained upregulation of the chaperone-mediated autophagy in the compromised cells has been recently noticed Accordingly, age-related dysfunction of autophagy in the retina might represent another determinant for glaucoma progression Indeed, association of glaucoma with age-related neurodegenerative diseases stems from these sharing similar miRNAs regulated transduction pathways (see also Part B for additional evidence) In fact, by means of in silico approaches and access to bioinformatic resources, deregulated miRNAs in glaucoma, in age-related macular degeneration (AMD) and Alzheimer’s disease (AD), respectively, have been found Actually, 88 predicted miRNAs are common to glaucoma and AMD; 19 are common to glaucoma and AD; and are common to AMD and AD These findings provide a valuable hint to assess deregulation of specific miRNA as potential biomarkers and therapeutic targets, in glaucoma and other neurodegenerative diseases by means of preclinical and clinical studies The wealth of the above-mentioned data in conjunction with important news emerging from clinical genetics and cell therapy technology is deeply discussed by authoritative, world-widely recognized, scientists in this issue (Part A) of Progress in Brain Research dedicated to glaucoma To them is addressed our sincere acknowledgment for making the issue a success Also, our thanks go to the skillful technical collaboration of individuals belonging to the Production Department of Elsevier We are especially indebted to Shellie Bryant for her continuous and highly qualified editorial assistance from the very beginning of this venture The Editors Giacinto Bagetta and Carlo Nucci CHAPTER Retinal neurodegeneration in experimental glaucoma Manuel Vidal-Sanz1,2, Francisco J Valiente-Soriano1, Arturo Ortı´n-Martı´nez1, Francisco M Nadal-Nicola´s1, Manuel Jime´nez-Lo´pez, Manuel Salinas-Navarro, Luis Alarco´n-Martı´nez, Diego Garcı´a-Ayuso, Marcelino Avile´s-Trigueros, Marta Agudo-Barriuso, Maria P Villegas-Pe´rez Departamento de Oftalmologı´a, Universidad de Murcia and Instituto Murciano de Investigacio´n Biosanitaria Virgen de la Arrixaca (IMIB-Arrixaca), Murcia, Spain Corresponding author: Tel.: +34-868-884330; Fax: +34-868-883962, e-mail address: manuel.vidal@um.es Abstract In rats and mice, limbar tissues of the left eye were laser-photocoagulated (LP) and ocular hypertension (OHT) effects were investigated week to months later To investigate the innermost layers, retinas were examined in wholemounts using tracing from the superior colliculi to identify retinal ganglion cells (RGCs) with intact retrograde axonal transport, melanopsin immunodetection to identify intrinsically photosensitive RGCs (m+RGC), Brn3a immunodetection to identify most RGCs but not m+RGCs, RECA1 immunodetection to examine the inner retinal vessels, and DAPI staining to detect all nuclei in the GC layer The outer retinal layers (ORLs) were examined in cross sections analyzed morphometrically or in wholemounts to study S- and L-cones Innervation of the superior colliculi was examined 10 days to 14 weeks after LP with orthogradely transported cholera toxin subunit B By weeks, OHT resulted in pie-shaped sectors devoid of FG+RGCs or Brn3a+RGCs but with large numbers of DAPI+nuclei Brn3a+RGCs were significantly greater than FG+RGCs, indicating the survival of large numbers of RGCs with their axonal transport impaired The inner retinal vasculature showed no abnormalities that could account for the sectorial loss of RGCs m+RGCs decreased to approximately 50–51% in a diffuse loss across the retina Cross sections showed focal areas of degeneration in the ORLs RGC loss at m diminished to 20–25% and did not progress further with time, whereas the S- and L-cone populations diminished progressively up to m The retinotectal projection was reduced by 10 days and did not progress further LP-induced OHT results in retrograde degeneration of RGCs and m+RGCs, severe damage to the ORL, and loss of retinotectal terminals Equally contributed to this work Progress in Brain Research, Volume 220, ISSN 0079-6123, http://dx.doi.org/10.1016/bs.pbr.2015.04.008 © 2015 Elsevier B.V All rights reserved Results 3.2.1 AMA0526 Improves Surgical Outcome in a Rabbit Model of Glaucoma Filtration Surgery To determine the effect of AMA0526 glaucoma filtration surgery outcome, bleb area, IOP, and bleb survival were assessed in a rabbit model of glaucoma filtration surgery Topical instillation with AMA0526 0.3% significantly improved bleb area compared to vehicle-treated control animals (ANOVA; P < 0.0001; Fig 3A) IOP was also significantly lower in the group treated with the ROCK inhibitor (ANOVA; P < 0.0001; Fig 3B) As shown in the Kaplan–Meier survival curve, AMA0526 also significantly prolonged bleb survival compared to the control group (log-rank test; P ¼ 0.0025; Fig 3C) All vehicle-treated blebs failed by day 21 after surgery, whereas blebs treated with AMA0526 survived up to day 35 Figure shows representative pictures of blebs treated with AMA0526 or vehicle Compared to treatment with the ROCK inhibitor, vehicle-treated blebs were characterized by a more pronounced vascularity in the early period after filtration surgery Immunohistological staining was performed to evaluate if AMA0526 affected inflammation, angiogenesis, and collagen deposition after filtration surgery in this model Compared to vehicle treatment, ROCK inhibition significantly decreased FIGURE Clinical investigation of rabbit eyes after glaucoma filtration surgery and treatment with AMA0526 (A) Bleb area of treated eyes was significantly improved compared to vehicle-treated control eyes (ANOVA; P < 0.0001; n ¼ 15) (B) A significant difference in postoperative IOP was shown between the AMA0526-treated group and the control group (ANOVA; P < 0.0001; n ¼ 15) As shown in the Kaplan–Meier survival curve, AMA0526 significantly prolonged bleb survival after filtration surgery (C) (log-rank test; P ¼ 0.0025; n ¼ 5) 291 292 CHAPTER 14 Rho kinase inhibitor AMA0526 FIGURE Representative pictures of blebs treated with vehicle or AMA0526 at several time points after surgery Compared to AMA0526 treatment, vehicle-treated blebs were characterized by a more pronounced vascularity at the early time points after filtration surgery Bleb failure was defined as the appearance of an avascular flat bleb the amount of CD45-positive inflammatory cells by 43%, 37%, and 44% at PD8, 14, and 30, respectively (t-test; P < 0.05; Fig 5A) A twofold reduction of blood vessel density was observed at all time points after surgery (t-test; P < 0.05; 49%, 42%, and 52%, respectively; Fig 5B) On surgical days 14 and 30, postoperative collagen I deposition at the site of filtration was significantly reduced by 23% and 48%, respectively (Fig 5C) Thus, AMA0526 improved surgical outcome by increasing bleb area and survival In addition, treatment with a ROCK inhibitor reduced inflammation, blood vessel density, and collagen deposition at the site of filtration surgery DISCUSSION Failure of glaucoma filtration surgery due to excessive postoperative wound healing with subsequent scarring is still a major problem in the management of glaucoma Surgical failure results in poor postoperative control of IOP and consequent progression of visual field loss (Addicks et al., 1983; Borisuth et al., 1999) One of the most important developments to improve glaucoma filtration surgery outcome is the use of antimetabolites, such as MMC and 5-FU However, by causing widespread cell death, the use of these nonspecific compounds as anti-scarring agents can be associated with sight-threatening complications (Crowston et al., 1998; Khaw et al., 1993) Hence, there is need to develop adjuvant strategies with more physiological action that specifically target the wound healing process to improve patient outcome following glaucoma surgery (Van Bergen et al., 2014) Rho kinase inhibitors have already been widely investigated for their IOPlowering properties by directly targeting trabecular meshwork contractility (Fukunaga et al., 2009; Honjo et al., 2001a,b; Isobe et al., 2014; Nishio et al., 2009; Ramachandran et al., 2011; Rao et al., 2001; Tokushige et al., 2007; Van De Velde et al., 2014) Additionally, this chapter contains increasing evidence that ROCK inhibitors, next to their IOP-lowering capacities, may possess beneficial effects for the management of glaucoma by improving glaucoma filtration surgery outcome Our in vitro results confirmed that the ROCK inhibitor, AMA0526, inhibited Discussion FIGURE Inflammation, blood vessel density, and collagen deposition in rabbit eyes after glaucoma filtration surgery The number of leukocytes, density of blood vessels, and collagen deposition were determined by counting the CD45-positive cells and calculating the CD31and collagen I-positive area as a proportion of the total wound area, respectively (A and B) At PD8, 14, and 30, AMA0526 significantly decreased the amount of CD45-positive cells and blood vessel density compared to vehicle-treated control eyes (C) Postoperative collagen I deposition at the site of filtration was significantly reduced at 14 and 30 days after surgery (t-test; P < 0.05; n ¼ 5/time point) (D) Representative pictures of CD45, CD31, and collagen I staining at different time points after glaucoma filtration surgery proliferation of HTF These fibroblasts are known to be the key players in ocular wound healing and should be considered as the main target for the development of anti-scarring strategies to improve surgery outcome (Khaw et al., 1994) Normally, fibroblasts exist within the subconjunctival connective tissue as quiescent cells After surgery, several growth factors stimulate fibroblast proliferation to generate a sufficient amount of cells at the site of injury (Khaw et al., 1994) Moreover, these cells can be activated and be differentiated into myofibroblasts TGF-b is considered as a pivotal mediator of fibroblast-to-myofibroblast transition which is characterized by a-SMA expression Myofibroblasts are at the core of the fibrotic process because they deposit extracellular matrix proteins in particular collagen which will obstruct the flow of aqueous humor through the created channel (Desmouliere et al., 1993; Seong et al., 2009) In this study, we investigated TGF-b1-induced a-SMA expression in HTF to reveal the direct role of Rho-ROCK signaling on this process Indeed, exposure of HTF to TGF-b1 induced a positive a-SMA staining Coincubation with AMA0526 inhibited this TGF-b1-induced a-SMA staining in HTF An explanation of how ROCK inhibitors prevent this transdifferentiation is postulated by 293 294 CHAPTER 14 Rho kinase inhibitor AMA0526 Meyer-Ter-Vehn et al They showed that a rapid contractile response is induced by TGF-b in HTF which precedes myofibroblast transdifferentiation ROCK inhibitors are able to counteract this contraction and subsequently prevent TGF-b-induced fibroblast-to-myofibroblast differentiation and may therefore modulate postoperative scarring after glaucoma filtration surgery (Meyer-Ter-Vehn et al., 2006) A rabbit model of glaucoma filtration surgery was used to further investigate the antifibrotic effect Collagen I is the most abundant collagen type correlated with the process of wound healing (Tahery and Lee, 1989) In vivo, AMA0526 was able to significantly decrease collagen I deposition at the site of filtration surgery, as compared to vehicle treatment Because wound healing and subsequent fibrosis are the result of a complex interplay of different processes such as inflammation and angiogenesis, we also investigated the effect of ROCK inhibition on these processes after glaucoma filtration surgery Experimental evidence already indicates the antiinflammatory and antiangiogenic effects of ROCK inhibition (Doe et al., 2007; He et al., 2008; Okamoto et al., 2010; Zhao et al., 2006) Indeed, AMA0526 also significantly reduced the amount of CD45-positive inflammatory cells and blood vessel density after glaucoma filtration surgery Importantly, this effect of AMA0526 was translated into an improved bleb area and IOP control compared to the control group One of the most important risk factors related to surgical outcome is bleb vascularity Indeed, severe vascularity is an important predictive factor for early bleb failure Therefore, vascularity of the bleb is a good measure for surgical outcome (Khaw et al., 2007) Compared to vehicle treatment, AMA0526-treated blebs were characterized by less pronounced vascularity in the early period after filtration surgery These results indicate that AMA0526 interferes at different levels in the process of wound healing, leading to improved surgery outcome Therefore, ROCK inhibitors may be considered as more physiological agents, which more specifically target the wound healing process compared to antimitotics Further studies need to be performed to directly compare AMA0526 to MMC Taken together, our data indicate that AMA0526 effectively inhibited proliferation and transdifferentiation of HTF in vitro Additionally, AMA0526 improved glaucoma filtration surgery outcome in rabbits by a reduction of inflammation, angiogenesis, and collagen deposition Therefore, ROCK inhibitors might be useful agents for improving the success rate of glaucoma filtration surgery and thereby upgrade the prognosis of glaucoma patients after surgery ACKNOWLEDGMENTS The authors wish to thank Sofie Beckers, Ann Verbeek, and Martine Leijssen for their technical support AMA0526 was kindly provided by Amakem Ophthalmics Financial Disclosure: Supported by the IWT (Agentschap voor Innovatie door Wetenschap en Technologie Vlaanderen), FWO (Fonds voor Wetenschappelijk Onderzoek Vlaanderen), FRO (Fund for Research in Ophthalmology), and Amakem Ophthalmics (Diepenbeek, Belgium) 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(Pt 2), 188–195 Khaw, P., Grehn, F., Hollo, G., Overton, B., Wilson, R., Vogel, R., Smith, Z., 2007 A phase III study of subconjunctival human anti-transforming growth factor beta(2) monoclonal antibody (CAT-152) to prevent scarring after first-time trabeculectomy Ophthalmology 114, 1822–1830 Li, Z., Van Bergen, T., Van De Veire, S., Van De Vel, I., Moreau, H., Dewerchin, M., Maudgal, P.C., Zeyen, T., Spileers, W., Moons, L., Stalmans, I., 2009 Inhibition of vascular endothelial growth factor reduces scar formation after glaucoma filtration surgery Invest Ophthalmol Vis Sci 50, 5217–5225 Liao, J.K., Seto, M., Noma, K., 2007 Rho kinase (ROCK) inhibitors J Cardiovasc Pharmacol 50, 17–24 Meyer-Ter-Vehn, T., Sieprath, S., Katzenberger, B., Gebhardt, S., Grehn, F., Schlunck, G., 2006 Contractility as a prerequisite for TGF-beta-induced myofibroblast transdifferentiation in human tenon fibroblasts Invest Ophthalmol Vis Sci 47, 4895–4904 Nishio, M., Fukunaga, T., Sugimoto, M., Ikesugi, K., Sumi, K., Hidaka, H., Uji, Y., 2009 The effect of the H-1152P, a potent Rho-associated coiled coil-formed protein kinase inhibitor, in rabbit normal and ocular hypertensive eyes Curr Eye Res 34, 282–286 Okamoto, H., Yoshio, T., Kaneko, H., Yamanaka, H., 2010 Inhibition of NF-kappaB signaling by fasudil as a potential therapeutic strategy for rheumatoid arthritis Arthritis Rheum 62, 82–92 Ramachandran, C., Patil, R.V., Combrink, K., Sharif, N.A., Srinivas, S.P., 2011 Rho-Rho kinase pathway in the actomyosin contraction and cell-matrix adhesion in immortalized human trabecular meshwork cells Mol Vis 17, 1877–1890 Rao, P.V., Deng, P.F., Kumar, J., Epstein, D.L., 2001 Modulation of aqueous humor outflow facility by the Rho kinase-specific inhibitor Y-27632 Invest Ophthalmol Vis Sci 42, 1029–1037 Roth, S.M., Spaeth, G.L., Starita, R.J., Birbillis, E.M., Steinmann, W.C., 1991 The effects of postoperative corticosteroids on trabeculectomy and the clinical course of glaucoma: fiveyear follow-up study Ophthalmic Surg 22, 724–729 Seong, G.J., Hong, S., Jung, S.A., Lee, J.J., Lim, E., Kim, S.J., Lee, J.H., 2009 TGF-betainduced interleukin-6 participates in transdifferentiation of human Tenon’s fibroblasts to myofibroblasts Mol Vis 15, 2123–2128 Skuta, G.L., Beeson, C.C., Higginbotham, E.J., Lichter, P.R., Musch, D.C., Bergstrom, T.J., Klein, T.B., Falck Jr., F.Y., 1992 Intraoperative mitomycin versus postoperative 5-fluorouracil in high-risk glaucoma filtering surgery Ophthalmology 99, 438–444 Tahery, M.M., Lee, D.A., 1989 Review: pharmacologic control of wound healing in glaucoma filtration surgery J Ocul Pharmacol 5, 155–179 Tokushige, H., Inatani, M., Nemoto, S., Sakaki, H., Katayama, K., Uehata, M., Tanihara, H., 2007 Effects of topical administration of y-39983, a selective rho-associated protein kinase inhibitor, on ocular tissues in rabbits and monkeys Invest Ophthalmol Vis Sci 48, 3216–3222 References Tura, A., Grisanti, S., Petermeier, K., Henke-Fahle, S., 2007 The Rho-kinase inhibitor H-1152P suppresses the wound-healing activities of human Tenon’s capsule fibroblasts in vitro Invest Ophthalmol Vis Sci 48, 2152–2161 Van Bergen, T., Van De Velde, S., Vandewalle, E., Moons, L., Stalmans, I., 2014 Improving patient outcomes following glaucoma surgery: state of the art and future perspectives Clin Ophthalmol 8, 857–867 Van De Velde, S., Van Bergen, T., Sijnave, D., Hollanders, K., Castermans, K., Defert, O., Leysen, D., Vandewalle, E., Moons, L., Stalmans, I., 2014 AMA0076, a novel, locally acting rho kinase inhibitor, potently lowers intraocular pressure in New Zealand white rabbits with minimal hyperemia Invest Ophthalmol Vis Sci 55 (2), 1006–1016 Van Nieuw Amerongen, G.P., Koolwijk, P., Versteilen, A., Van Hinsbergh, V.W., 2003 Involvement of RhoA/Rho kinase signaling in VEGF-induced endothelial cell migration and angiogenesis in vitro Arterioscler Thromb Vasc Biol 23, 211–217 Weinreb, R.N., 1995 Riding the trojan horse of glaucoma surgery J Glaucoma 4, 2–4 Zhao, L., Xu, G., Zhou, J., Xing, H., Wang, S., Wu, M., Lu, Y.P., Ma, D., 2006 The effect of RhoA on human umbilical vein endothelial cell migration and angiogenesis in vitro Oncol Rep 15, 1147–1152 297 Index Note: Page numbers followed by f indicate figures and t indicate tables A AD See Alzheimer’s disease (AD) Adaptive optics confocal scanning laser tomography (AOCSLT), 180–181 Age-related macular degeneration (AMD), 242 gene association studies for, 222t inflammation, 232 microRNA (miRNA), 220, 221t, 221f oxidative stress, 139 Venn diagram, 220, 221f Alzheimer’s disease (AD), 88, 206, 218–219 inflammation, 232 miRTarBase-predicted pathways, 223–224, 224f Venn diagram, 220, 221f AMA0526, glaucoma filtration surgery fibroblast-to-myofibroblast differentiation, 290–292, 290f HTFs, inhibits proliferation of, 289–290, 289f materials and methods cells and culture conditions, 285–286 in vitro assays, 286–289 rabbit model, 291–292, 291–293f AMD See Age-related macular degeneration (AMD) 2-Aminoethylsulfonic acid, 268–270 Amyloid b (Ab) deposition, 218–219 Analysis of variance (ANOVA), 289 Anterior chamberassociated immune deviation (ACAID), 166–167 Antioxidants and glaucoma animal models, 139–140, 140t in human studies, 140–143 interventional studies, 143, 144t observational studies, 140–142, 141t Apoptosis, 223–225 Apoptosis proteins, caspases and inhibitor of, 47–48 Aqueous humor dynamics and IOP conventional outflow/TM pathway, 187 drainage of aqueous humor, 186, 187f unconventional outflow/UVS pathway, 187–188 Aqueous outflow modulation, stem cells function and pathology, 243 TM repair, 244 Aqueous outflow pathway, 243 Astrocytes (AROA), 160 Astroglial cells, reactivation of, 163–164 Atg5 genes, 98 Atg7 genes, 98 Autoimmunity, 222–223 Autophagic markers, 92, 93t Autophagolysosomal system, 89 Autophagy dysregulation aging and glaucoma, 98 mechanisms, regulation, and functions, 89–91 phosphoinositide-3 kinase (PI3K) inhibitor, 97 retinal ganglion cells (RGCs) death of, 91–98 B BCL-2 family, 45–47, 46f BECN1 genes, 98 Blood brain barrier (BBB), 222–223 Blood retinal barrier (BRB), 222–223 Brain-derived neurotrophic factor (BDNF) gene, 3, 9–10, 245 Brn3a+RGCs, 21–23, 22f C Cannabinoids, 258–259 Causative POAG disease genes, 108–110, 108t Cellular protein catabolism, 89 Central nervous system (CNS), 156 Ciliary neurotrophic factor (CNTF), 245 c-Jun N-terminal kinases (JNKs), 40–43 CNS See Central nervous system (CNS) CNTF See Ciliary neurotrophic factor (CNTF) Coenzyme Q (CoQ10), 259–260 Confocal scanning laser ophthalmoscope (CSLO), 203 Confocal scanning laser tomography (CSLT), 174–175 Contralateral eye, 157, 163–167 Contralateral superior colliculus, 11f Conventional pathway, 186 Cytokine-cytokine receptor interaction, 223–224 D Damage-associated molecular patterns (DAMPs), 226 Data interpretation caveats, 97–98 Deformations, intraocular pressure (IOP), 72, 73f Diabetic retinopathy (DR), 138 299 300 Index E Enhanced depth imaging (EDI), 180 Erigeron breviscapus (vant.) Hand Mazz (EBHM), 260–262 ERKs See Extracellular signal- regulated protein kinases (ERKs) Erythropoietin (EPO), 262–263 Estrogens, 263–264 Experimental glaucoma (EG), 174–175 Extracellular matrix (ECM), 243 Extracellular signal- regulated protein kinases (ERKs), 44–45 Eye, lymphatic drainage aqueous humor dynamics and IOP conventional outflow/TM pathway, 187 drainage of aqueous humor, 186, 187f unconventional outflow/UVS pathway, 187–188 glaucoma treatment, 190, 191–192f ocular lymphatics, 188–189, 189f F Flavonoids, 264–265 Forskolin, 265–266 G Ganglion cell layer (GCL), GDx nerve fiber analyzer, 176 Genome-wide association studies (GWAS) approach primary angleclosure glaucoma (PACG), 116 primary open-angle glaucoma (POAG), 111, 112t, 114–115 GFAP See Glial fibrillary acidic protein (GFAP) GFAP-labeled retinal area (GFAP-RA), 160–162, 161f Glaucoma animal models, 67–68 connective tissue-based therapies noncellular components, 75–76 remodeling, alter cellular behavior in, 76–77 scleral/ONH protective therapy, 77–78 gene association studies for, 222t genetic models of, 96–97 inflammation, 232 microRNA (miRNA) in, 220, 220t, 221f ONH, remodeling of, 72–74, 73–74f sclera and ONH in mouse glaucoma, 68–72, 69–70f pathological changes, 64–67, 65–66f stem cell, 242 Venn diagram, 220, 221f vision loss, dendritic pathology, 199–200 Glaucoma genetics familial linkage analysis causative POAG disease genes, 108–110, 108t myocilin (MYOC), 108t, 109 optineurin (OPTN), 108t, 109–110 WDR36 (WD repeat domain 36), 108t, 110 genes, 111t genome-wide association studies (GWAS) approach primary angleclosure glaucoma (PACG), 116 primary open-angle glaucoma (POAG), 111, 112t, 114–115 perspective, 116 Glaucoma pathogenesis antioxidants and glaucoma animal models, 139–140, 140t in human studies, 140–143, 141t interventional studies, 143, 144t observational studies, 140–142, 141t mitochondrial involvement in glaucoma disease, 133–134 mitochondrial failure and glaucoma, genetic links, 134–136, 135t mitochondrial-induced cell damage, 130f, 132–133, 133t neurodegeneration, 130f, 132–133, 133t reactive oxygen species (ROS), 129–131, 130f oxidative stress age-related macular degeneration (AMD), 139 diabetic retinopathy (DR), 138 primary open-angle glaucoma, 138 pro-oxidants and antioxidants sources, 137f reactive oxygen species, 137f Glaucomatous eyes contralateral eye, 157, 163–167 optic-nerve crush, 157–158 optic-nerve section, 157–158 RGC damage after axonal injury, 157–158 after IOP elevation, 158–163, 161–162f Glaucomatous optic neuropathies (GONs), 2, 156 autophagy aging and glaucoma, 98 mechanisms, regulation, and functions, 89–91 retinal ganglion cells (RGCs) death, 91–98 intraocular pressure (IOP), 88 Glial fibrillary acidic protein (GFAP), 156 GWAS approach See Genome-wide association studies (GWAS) approach H Heidelberg Retina Tomograph (HRT), 174 Hematic involvement, 164–165 Index Human miRNA Disease Database (HMDD), 219 Human Tenon Fibroblast (HTF), 284–285 See also AMA0526, glaucoma filtration surgery Huntington’s disease, 88 Hypoxia-inducible transcription factor (HIF), 228–229 I Immune system, 165 Induced pluripotent stem (iPS), 242–243 Intraocular pressure (IOP), 2–3, 59–64, 62f, 186, 200, 224–225, 241–242 In vitro assays, AMA0526 caspase 3/7 activity assay, 286–287 fibroblast-to-myofibroblast differentiation, 287 filtration surgery, rabbit model of, 287–288 histological evaluation, 288–289 proliferation assay, 286 statistical analysis, 289 treatment regimen and postoperative clinical examination, 288 IOP See Intraocular pressure (IOP) J JNKs See c-Jun N-terminal kinases (JNKs) Junctional adhesion molecule B, 201–202 K Kaplan–Meier survival analysis, 289 KEGG pathways, 223–224 L Lamina cribrosa, 61–64, 63f Laser photocoagulation (LP), 2–3 limbal and episcleral veins results, Lateral geniculate nucleus (LGN), 247–248 Lycium barbarum, 266–267 Lymphatic drainage, eye aqueous humor dynamics and IOP conventional outflow/TM pathway, 187 drainage of aqueous humor, 186, 187f unconventional outflow/UVS pathway, 187–188 glaucoma treatment, 190, 191–192f ocular lymphatics, 188–189, 189f Lymphatic vessel endothelial hyaluronan receptor-1 (LYVE-1), 188 Lysyl oxidase-like protein (LOXL1), 67 M Mammalian target of rapamycin (mTOR) kinase, 89, 208–209 MAP-kinase pathway, 40, 43f Melanopsin, 21–23, 201–202 Mesenchymal stem cells (MSCs), 243 MicroRNA (miRNA) in age-related macular degeneration (AMD), 220, 221t, 221f apoptosis, 224–225 cytokine-cytokine receptor interaction, 225–226 data mining of, 219 in glaucoma, 220, 220t, 221f HIF-1 pathway, 228–229 microT-CDS, 222–223, 223t miRTarBase-predicted pathways, 224f NF-kB pathway, 227–228 NT pathway, 229–230 pathways, 220 target prediction, 219 TLR signaling pathway, 226–227 miR2Disease, 219 Mitochondrial involvement, glaucoma pathogenesis glaucoma disease, 133–134 mitochondrial failure and glaucoma, genetic links, 134–136, 135t mitochondrial-induced cell damage, 130f, 132–133, 133t neurodegeneration, 130f, 132–133, 133t reactive oxygen species (ROS), 129–131, 130f m+RGCs, 21–23, 22f Multipotent, 242–243 Myocilin (MYOC), 97, 108t, 109 N Natural compounds, retinal ganglion cell neuroprotection cannabinoids, 258–259 coenzyme Q (CoQ10), 259–260 Erigeron breviscapus (vant.) Hand Mazz (EBHM), 260–262 erythropoietin (EPO), 262–263 estrogens, 263–264 flavonoids, 264–265 forskolin, 265–266 Lycium barbarum, 266–267 neuroglobin, 267 prostaglandins (PGs), 267–268 taurine (2-aminoethylsulfonic acid), 268–270 Neurofilament-H (NF-H), 201–202 Neuroglobin, 267 Neurotrophic factors (NTFs), 242 301 302 Index Neurotrophin (NT), 223–224 N-methyl-D-aspartate (NMDA), 257–258 Normal tension glaucoma (NTG), 48–49, 88 O OAG See Open-angle glaucoma (OAG) Ocular hypertension (OHT), 95–96, 156 GCL, 12f, 16–17 ORLs, 12f, 17–21 outer retinal layers focal degeneration after, 18f retinal ganglion cells, 12f retinal injury, 21–23 retinal vessels, 15f Open-angle glaucoma (OAG), 59–60 Optic-nerve crush, 157–158 Optic nerve crush/axotomy, 92–94 Optic nerve head (ONH), 59–60, 62f, 64–67, 74f, 173–174, 200 Optic-nerve section, 157–158 Optineurin (OPTN), 96, 108t, 109–110 Outer retinal layers (ORLs), Oxidative stress age-related macular degeneration (AMD), 139 diabetic retinopathy (DR), 138 primary open-angle glaucoma, 138 pro-oxidants and antioxidants sources, 137f reactive oxygen species, 137f P P38, 43–44 Parkinson’s disease (PD), 88 Peripapillary sclera (PPS), 60–64 PI-3 kinase/Akt pathway, 44–45 Post-chiasmatic contralateral axons, 164 Primary angleclosure glaucoma (PACG), 116 Primary open-angle glaucoma (POAG), 111, 112t, 114–115, 128 Prostaglandins (PGs), 190, 267–268 R Reactive oxygen species (ROS), 137f, 228–229 Retinal degeneration, 9–10 Retinal ganglion cell apoptotic pathway apoptosis proteins, caspases and inhibitor of, 47–48 BCL-2 family, 45–47, 46f BDNF and neurotrophic factor deprivation, 38–40, 39f c-Jun N-terminal kinases (JNKs), 40–43 extracellular signal- regulated protein kinases (ERKs), 44–45 limitations, 49–50 MAP-kinase pathway, 40, 43f normal tension glaucoma (NTG), 48–49 P38, 43–44 PI-3 kinase/Akt pathway, 44–45 signaling pathways, 41t Retinal ganglion cells (RGCs), 2, 10–16, 59–60, 241–242, 257–258 axonal injury acute axonal injury, 203–204 Alzheimer’s disease (AD), 206 amyotrophic lateral sclerosis, 206 ocular hypertension models, 204–206 Parkinson’s disease, 206 death of autophagic markers, 92, 93t data interpretation caveats, 97–98 glaucoma, genetic models of, 96–97 ocular hypertension, 95–96 optic nerve crush/axotomy, 92–94 retinal ischemia/reperfusion, 94–95 dendrite and synapse stability GTPases, 207–208 rapamycin pathway, 208–209 dendritic pathology, 199–200 morphological diversity, 200–202, 201f stem cells axon projection and synapse formation, 247–248 differentiation, 247 migration and integration, 246–247 neural replacement vs neuroprotection, 248 surviving percentage, 20f topological analysis of, 19f Retinal ischemia/reperfusion, 94–95 Retinal nerve fiber layer (RNFL), 173–174 Retinal neurodegeneration image acquisition, 7–8 image analysis, immunodetection and DAPI staining, 6–7 methods animal handling, 4–5 animal manipulations, tissue processing, ocular hypertension (OHT) GCL, 16–17 ORLs, 17–21 retinal injury, 21–23 results and discussion BDNF, 10–16 laser-photocoagulated (LP), ocular hypertension (OHT), retinotectal innervation, 9–10 Index statistical analysis, Retinal neuroprotection obstacles, 246 stem cell-mediated neuroprotection, 245–246 stem cell sources and NTFS, 245 Retinal pigment epithelium (RPE), 157 RGCs See Retinal ganglion cells (RGCs) Rho-associated kinases (ROCK), 284–285 Rho-family of GTPases, 207–210 Rho kinase inhibitor See AMA0526, glaucoma filtration surgery ROS See Reactive oxygen species (ROS) S Scanning laser polarimetry (SLP), 175–176 Sclera, 64–67 biomechanical testing of, 67 Signaling pathways, 41t Spectral domain optical coherence tomography (SDOCT), 177–180 Stargardt’s disease, 242 Stem cells aqueous outflow modulation function and pathology, 243 TM repair, 244 clinical application and translation, 249–250 definition, 242–243 delivery methods, 248–249 endogenous stem cells, 243 induced pluripotent stem (iPS), 242–243 mesenchymal stem cells (MSCs), 243 retinal ganglion cells (RGCs) axon projection and synapse formation, 247–248 differentiation, 247 migration and integration, 246–247 neural replacement vs neuroprotection, 248 retinal neuroprotection obstacles, 246 stem cell-mediated neuroprotection, 245–246 stem cell sources and NTFS, 245 Superior colliculus (SC), T Taurine (2-aminoethylsulfonic acid), 268–270 Thrombospondins (TSPs), 71 TM See Trabecular meshwork (TM) Toll-like receptor (TLR) signaling pathway, 223–224 Topographical change analysis (TCA), 175 Totipotent, 242–243 Trabecular meshwork (TM), 97, 186, 186, 242 U Ubiquitin-proteasome system (UPS), 89 Uveoscleral (UVS) pathway, 187–188 V Vascular endothelial growth factor (VEGF), 284–285 W WDR36 (WD repeat domain 36), 108t, 110 Y Yellow fluorescent protein (YFP), 201–202 303 Other volumes in PROGRESS IN BRAIN RESEARCH Volume 167: Stress Hormones and Post Traumatic Stress Disorder: Basic Studies and Clinical Perspectives, by E.R de Kloet, M.S Oitzl and E Vermetten (Eds.) – 2008, ISBN 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Neuro-Historical Dimensions, by Stanley Finger, Dahlia W Zaidel, Franc¸ois Boller and Julien Bogousslavsky (Eds.) – 2013, 978-0-444-62730-8 Volume 204: The Fine Arts, Neurology, and Neuroscience: New Discoveries and Changing Landscapes, by Stanley Finger, Dahlia W Zaidel, Franc¸ois Boller and Julien Bogousslavsky (Eds.) – 2013, 978-0-444-63287-6 Volume 205: Literature, Neurology, and Neuroscience: Historical and Literary Connections, by Anne Stiles, Stanley Finger and Franc¸ois Boller (Eds.) – 2013, 978-0-444-63273-9 Volume 206: Literature, Neurology, and Neuroscience: Neurological and Psychiatric Disorders, by Stanley Finger, Franc¸ois Boller and Anne Stiles (Eds.) – 2013, 978-0-444-63364-4 Volume 207: Changing Brains: Applying Brain Plasticity to Advance and Recover Human Ability, by Michael M Merzenich, Mor Nahum and Thomas M Van Vleet (Eds.) – 2013, 978-0-444-63327-9 Volume 208: Odor Memory and Perception, by Edi Barkai and Donald A Wilson (Eds.) – 2014, 978-0-444-63350-7 Volume 209: The Central Nervous System Control of Respiration, by Gert Holstege, Caroline M Beers and Hari H Subramanian (Eds.) – 2014, 978-0-444-63274-6 Volume 210: Cerebellar Learning, Narender Ramnani (Ed.) – 2014, 978-0-444-63356-9 Volume 211: Dopamine, by Marco Diana, Gaetano Di Chiara and Pierfranco Spano (Eds.) – 2014, 978-0-444-63425-2 Volume 212: Breathing, Emotion and Evolution, by Gert Holstege, Caroline M Beers and Hari H Subramanian (Eds.) – 2014, 978-0-444-63488-7 Volume 213: Genetics of Epilepsy, by Ortrud K Steinlein (Ed.) – 2014, 978-0-444-63326-2 Volume 214: Brain Extracellular Matrix in Health and Disease, by Asla Pitkaănen, Alexander Dityatev and Bernhard Wehrle-Haller (Eds.) – 2014, 978-0-444-63486-3 Other volumes in PROGRESS IN BRAIN RESEARCH Volume 215: The History of the Gamma Knife, by Jeremy C Ganz (Ed.) – 2014, 978-0-444-63520-4 Volume 216: Music, Neurology, and Neuroscience: Historical Connections and Perspectives, by Francáois Boller, Eckart Altenmuăller, and Stanley Finger (Eds.) – 2015, 978-0-444-63399-6 Volume 217: Music, Neurology, and Neuroscience: Evolution, the Musical Brain, Medical Conditions, and Therapies, by Eckart Altenmuăller, Stanley Finger, and Franc¸ois Boller (Eds.) – 2015, 978-0-444-63551-8 Volume 218: Sensorimotor Rehabilitation: At the Crossroads of Basic and Clinical Sciences, by Numa Dancause, Sylvie Nadeau, and Serge Rossignol (Eds.) – 2015, 978-0-444-63565-5 Volume 219: The Connected Hippocampus, by Shane O’Mara and Marian Tsanov (Eds.) – 2015, 978-0-444-63549-5 307 ... on the retinal pathology induced by OHT In certain glaucoma patients despite the efforts to maintain IOP below certain levels, RGC loss keeps progressing to blindness This has prompted investigators... abnormal RT97 staining in bundles of axons and RGCs that were mainly located outside the areas of the retina containing surviving RGCs This abnormal RT97 staining consisted of axonal beadings and varicosities... anterograde tracer, we have investigated the fate of the retinal terminals in their main target in the brain, the contralateral SC METHODS 2.1 ANIMAL HANDLING Experiments were prepared in accordance with