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Retinal Plasticity in Experimental Glaucoma

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Retinal Plasticity in Experimental Glaucoma Retinal plasticity in experimental glaucoma Paulina Anna Samsel Thesis submitted to Cardiff University in accordance with the requirements for the degree of Doctor of Philosophy Visual Neuroscience and Molecular Biology Group School of Optometry and Vision Sciences Cardiff University June 2010 UMI Number: U517580 All rights reserved INFORMATION TO ALL USERS The quality of this reproduction is dependent upon the quality of the copy submitted. In the unlikely event that the author did not send a complete manuscript and there are missing pages, these will be noted. Also, if material had to be removed, a note will indicate the deletion. Dissertation Publishing UMI U517580 Published by ProQuest LLC 2013. Copyright in the Dissertation held by the Author. Microform Edition © ProQuest LLC. All rights reserved. This work is protected against unauthorized copying under Title 17, United States Code. ProQuest LLC 789 East Eisenhower Parkway P.O. Box 1346 Ann Arbor, Ml 48106-1346 DECLARATION This work has not previously been accepted in substance for any degree and is not concurrently submitted in candidature for any degree. Signed , .. .^ k r r A ^ C . (candidate) Date 2 6 f o ( / ?■=>(/ STATEMENT 1 This thesissis is tbeing submitted in partial fulfillment of the requirements for the degree o f f .h M d . ......................(insert MCh, MD, MPhil, PhD etc, as appropriate) Signed (candidate) Date 7 Q /< ? ' / STATEMENT 2 This thesis is the result of my own independent work/investigation, except where otherwise stated. Other sources are acknowledged by explicit references. Signed .................... (candidate) Date ^ c y / STATEMENT 3 I hereby give consent for my thesis, if accepted, to be available for photocopying and for inter-library loan, and for the title and summary to be made available to outside organisations. Signed (candidate) Date I CONTENTS DECLARATION.......................................................................................................................................... I Acknowledgements ....................................................................................................................................VI Contributions ............................................................................................................................................ VII Abbreviations ..............................................................................................................................................IX List of Figures...........................................................................................................................................XII List of Tables....................................................................................................................................... XVIII Chapter 1: Introduction...................................................................................................................................... 1 1.1. Glaucoma................................................................................................................................................... 2 1.1.1. Background ........................................................................................................................................2 1.1.2. Forms of glaucom a.......................................................................................................................... 2 1.1.3. Diagnostic aspects.............................................................................................................................3 1.1.4. Research approaches to treatment ..................................................................................................3 1.1.5. Current treatments .............................................................................................................................5 1.1.5.1. Effect o f IOP reduction ...........................................................................................................5 1.1.5.2. Pharmacotherapy ......................................................................................................................6 1.1.5.3. Surgery and laser techniques .................................................................................................. 7 1.1.6. Mechanisms o f RGC death in glaucoma ....................................................................................... 7 1.1.6.1. Neurotrophic factor deprivation .............................................................................................8 1.1.6.2. Hypoperfusion .......................................................................................................................... 8 1.1.6.3. Glial cell activation ................................................................................................................. 9 1.1.6.4. Glutamate toxicity ..................................................................................................................10 1.1.6.5. Abnormal immune response ................................................................................................ 10 1.1.7. Morphology and function ofR G C s ............................................................................................. 11 1.1.7.1. Morphology and function of RGC in glaucoma .................................................................14 1.1.8. Models of experimental glaucoma............................................................................................... 15 1.1.8.1. Mouse model of glaucoma....................................................................................................16 1.1.8.2. Non-human primate model of IOP-induced optic nerve damage .................................... 17 1.1.8.3. Rat model of IOP-induced optic nerve damage .................................................................19 1.1.8.4. In vitro system s ..................................................................................................................... 21 1.1.8.4.1. Retinal explant culture .................................................................................................. 21 1.1.8.4.2. Retinal ganglion cell culture ........................................................................................ 21 1.2. Neuroplasticity........................................................................................................................................ 22 1.2.1. Neuroplasticity and diseases of the C N S .................................................................................... 23 1.2.2. Regulators o f neuroplasticity........................................................................................................23 1.2.3. Neuroplasticity in the retina ..........................................................................................................24 1.2.4. Neurotrophins............................................................................................................................25 1.2.5. Perineuronal nets .......................................................................................................................28 1.2.5.1. Structure and composition ...............................................................................................28 1.2.5.2. Functional roles ofPNNs in plasticity............................................................................29 1.3. Hypothesis................................................................................................................................................ 30 1.4. Project aims.............................................................................................................................................. 31 Chapter 2: Materials and Methods................................................................................................................32 2.1. Ex vivo - retinal explant culture.........................................................................................................33 2.1.1. Experimental animals .................................................................................................................... 33 2.1.2. Retinal dissection ............................................................................................................................ 33 2.1.3. Culture conditions ........................................................................................................................... 34 2.1.4. Cell viability assays ........................................................................................................................36 2.1.4.1. Image capture of labelled RGCs.......................................................................................... 36 2.1.4.2. Statistical analysis .................................................................................................................. 37 2.1.5. Gene gun processes ......................................................................................................................... 37 2.1.5.1. Biolistic labelling....................................................................................................................38 2.1.5.2. Diolistic labelling....................................................................................................................38 II 2.1.5.3. Image capture of labelled RGCs..........................................................................................39 2.1.6. Effects of BDNF and caspase inhibitors
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