Nogo is well established for its inhibitory action on axon regeneration in the adult central nervous system. It binds to the Nogo receptor (NgR) through an extracellular active site on the protein-Nogo-66. Although it is reported that Nogo is widely expressed in the developing brain, its exact function during development of the nervous system is unclear. / The contribution of Nogo on patterning the axon routing at the optic chiasm of mouse embryo was investigated in this thesis. Using immunocytochemical staining, Nogo protein was localized on the Miller glial cells in the retina and at the optic disk. A few migrating retinal neurons also expressed Nogo. In the chiasm, Nogo was localized exclusively on the radial glia, which generate a midline domain where turning of uncrossed axons occurs. In vitro study showed expression of NgR on retinal neurites and growth cones, and neurite outgrowth from both dorsal nasal (contralaterally projecting) and ventral temporal (ipsilaterally projecting) retina was inhibited by Nogo. In the pathway, NgR expression was regionally regulated. NgR was obvious in the optic stalk and the optic tract, but not in the chiasm. Blocking Nogo function with NEP1-40, a peptide antagonist of NgR, in brain slice culture of the pathway produced significant reduction in the uncrossed projection, but had no effect on axon crossing at the midline. Furthermore, the age related fiber arrangement in the optic tract was abolished after disturbing of Nogo function. Similar abnormalities were observed in slices treated with Nogo blocking antibody. In vitro studies showed that NEP1-40 rescued the inhibition of Nogo to the retinal neurites. The downregulation of NgR at the chiasm was supported by in vitro assays showing significant reduction of receptor expression on dorsal nasal but not ventral temporal growth cones when they encountered the chiasm, thus generating a differential inhibition to ventral temporal neurites. / These results provide evidences that Nogo is a guidance molecule during the development of CNS. Interaction of Nogo and its receptor plays important role for patterning the axon divergence in the mouse optic pathway and the age related fiber order in the optic tract. / Wang Jun. / "September 2006." / Adviser: Sun-On Chan. / Source: Dissertation Abstracts International, Volume: 68-03, Section: B, page: 1474. / Thesis (Ph.D.)--Chinese University of Hong Kong, 2006. / Includes bibliographical references (p. 130-142). / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Electronic reproduction. [Ann Arbor, MI] : ProQuest Information and Learning, [200-] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Abstracts in English and Chinese. / School code: 1307.

Chung Kit Ying. / Thesis (M.Phil.)--Chinese University of Hong Kong, 1999. / Includes bibliographical references (leaves 90-104). / Abstracts in English and Chinese. / Chapter Chapter 1 --- General Introduction --- p.1-22 / Chapter Chapter 2 --- Expression of Chondroitin Sulfate Proteoglycans (CSPGs) in the Chiasm of Mouse Embryos / Introduction --- p.23-24 / Materials and Methods --- p.25 -27 / Results --- p.28-33 / Discussion --- p.34-40 / Figures --- p.41-45 / Chapter Chapter 3 --- Effects on Axon Routing after Removal of Chondroitin Sulfate Proteoglycans by Enzymatic Digestion / Introduction --- p.46 -47 / Materials and Methods --- p.48 -50 / Results --- p.57 / Discussion --- p.57-61 / Figures --- p.62-66 / Chapter Chapter 4 --- Immediate Effects of Prenatal Monocular Enucleation on the Cellular and Molecular Environment in the Development of Retinofugal Pathway / Introduction --- p.67-69 / Materials and Methods --- p.70-72 / Results --- p.73.77 / Discussion --- p.78-82 / Figures --- p.83-86 / Chapter Chapter 5 --- General Conclusion --- p.87-89 / References --- p.90 -104

Leung Kin Mei. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2002. / Includes bibliographical references (leaves 106-125). / Abstracts in English and Chinese. / Chapter CHAPTER 1 --- GENERAL INTRODUCTION --- p.1-11 / Chapter CHAPTER 2 --- ENZYMATIC REMOVAL OF CHONDROITIN SULFATES ABOLISHES THE AGE-RELATED ORDER IN THE OPTIC TRACT OF MOUSE EMBRYOS / INTRODUCTION --- p.12-13 / MATERIALS AND METHODS --- p.13-18 / RESULTS --- p.18-24 / DISCUSSION --- p.24-29 / FIGURES --- p.30-39 / Chapter CHAPTER 3 --- EXPRESSION OF PHOSPHACAN AND NEUROCAN IN THE DEVELOPING MOUSE RETINOFUGAL PATHWAY / INTRODUCTION --- p.40-42 / MATERIALS AND METHODS --- p.42-43 / RESULTS --- p.44-49 / DISCUSSION --- p.49-55 / FIGURES --- p.56-61 / Chapter CHAPTER 4 --- HEPARAN SULFATE PROTEOGLYCAN EXPRESSION IN THE OPTIC CHIASM OF MOUSE EMBRYOS / INTRODUCTION --- p.62-63 / MATERIALS AND METHODS --- p.63-65 / RESULTS --- p.66-70 / DISCUSSION --- p.70-76 / FIGURES --- p.77-82 / Chapter CHAPTER 5 --- EXPRESSION OF NEURAL CELL ADHESION MOLECULES IN THE CHIASM OF MOUSE EMBRYOS / INTRODUCTION --- p.83-85 / MATERIALS AND METHODS --- p.85-88 / RESULTS --- p.88-92 / DISCUSSION --- p.92.95 / FIGURES --- p.96-102 / Chapter CHAPTER 6 --- GERNEAL CONCLUSION --- p.103-105 / REFERENCES --- p.106-125

Chan, Chung Kit. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2008. / Includes bibliographical references (leaves 98-110). / Abstracts in English and Chinese. / Abstract --- p.i / Abstract in Chinese --- p.iv / Acknowledgements --- p.v / Table of Abbreviations --- p.vi / Table of Contents --- p.vii / Chapter Chapter 1 --- General Introduction --- p.1 / Chapter Chapter 2 --- Functions of hyaluronan in the development of retinofugal pathway / Introduction --- p.18 / Materials and Methods --- p.19 / Results --- p.23 / Discussion --- p.26 / Figures --- p.32 / Chapter Chapter 3 --- Characterization of Nogo and its receptor in retinofugal pathway using Western blot analysis / Introduction --- p.40 / Materials and Methods --- p.42 / Results --- p.50 / Discussion --- p.52 / Figures --- p.57 / Chapter Chapter 4 --- Expression patterns and functions of Sonic hedgehogin retinofugal pathway / Introduction --- p.62 / Materials and Methods --- p.64 / Results --- p.69 / Discussion --- p.76 / Figures --- p.81 / Chapter Chapter 5 --- General Discussion --- p.91 / Figures --- p.95 / References --- p.98

Kong Fung Wong. / Thesis (M.Phil.)--Chinese University of Hong Kong, 1997. / Includes bibliographical references (leaves 59-70). / Chapter CHATPER 1 --- GENERAL INTRODUCTION --- p.1-12 / Chapter CHATPER 2 --- EXAMINATION OF THE BEHAVIOR OF GROWTH CONE IN DIFFERENT REGIONS OF THE OPTIC CHIASM / Introduction --- p.13-14 / Materials and Methods --- p.15-18 / Results --- p.19-23 / Discussion --- p.24-27 / Chapter CHATPER 3 --- STUDY OF BINOCULAR INTERACTION AFTER UNILATERAL INTRA-UTERO ENUCLEATION / Introduction --- p.28-29 / Materials and Methods --- p.30-31 / Results --- p.32-35 / Discussion --- p.36-39 / Chapter CHATPER 4 --- ISOLATION OF DIFFERENTIALLY EXPRESSED mRNA IN DIFFERENT REGIONS OF THE RETINA / Introduction --- p.40-43 / Materials and Methods --- p.44-48 / Results --- p.48-50 / Discussion --- p.51-54 / Chapter CHATPER 5 --- GENERAL DISCUSSION --- p.56-58 / REFERENCE --- p.59-70 / FIGURES / TABLES

Lam Shi Ying Joyce. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2005. / Includes bibliographical references (leaves 107-114). / Abstract in English and Chinese. / Chapter CHAPTER 1 --- GENERAL INTRODUCTION --- p.1-19 / Chapter CHAPTER 2 --- EXPRESSION OF PROTEIN KINASE C (PKC) ISOFORMS IN THE VENTRAL TEMPORAL (VT) AND DORSAL NASAL (DN) RETINAL GROWTH CONES OF MOUSE EMBRYOS / INTRODUCTION --- p.20-22 / MATERIALS AND METHODS --- p.22-24 / RESULTS --- p.24-31 / DISCUSSION --- p.31-37 / FIGURES --- p.38-46 / Chapter CHAPTER 3 --- EFFECTS ON MOUSE NASAL AND TEMPORAL RETINAL NEURITES TO CHONDROITIN SULPHATES (CS) AFTER ALTERATION OF PKC ACTIVITY / INTRODUCTION --- p.47-48 / MATERIALS AND METHODS --- p.49-51 / RESULTS --- p.51-59 / DISCUSSION --- p.60-67 / FIGURES --- p.68-74 / Chapter CHAPTER 4 --- EFFECTS ON AXON ROUTING AFTER ALTERATION OF PKC ACTIVITY ON GUIDANCE OF RETINAL GANGLION CELL AXONS AT THE OPTIC CHIASM OF MOUSE EMBRYOS / INTRODUCTION --- p.75-76 / MATERIALS AND METHODS --- p.77-80 / RESULTS --- p.80-89 / DISCUSSION --- p.89-95 / FIGURES --- p.96-103 / Chapter CHAPTER 5 --- GENERAL CONCLUSION --- p.104-106 / REFERENCES --- p.107-114

Chiasm cells, which include glia and neurons, are generated early before any retinal axon arrives at the midline of the mouse ventral diencephalon. These cells have been shown to affect retinal axon growth and patterning in the optic chiasm. In this study, we used EdU (5-ethyny1-2'-deoxyuridine) for birthdating these chiasm cells, aiming to find out when these cells are generated; then we tried to trace their fates at later stages of development. EdU injection at embryonic day (E) 9.5 to El 1 labeled a number of chiasmatic neurons and radial glial cells at E13, which were immunoreactive for SSEA-1 and RC2, respectively. After colocalization studies, we found that most of these neurons were born as early as E9.5, while a large number of radial glial cells were born as from El 1. Both E9.5-born chiasmatic neurons and Ell-born radial glia decreased by E14-E16; the radial glia even disappeared finally from the midline. Furthermore, we found that some chiasmatic neurons underwent apoptotic cell death as from El 4, and that the radial glia likely differentiated into other cell types after finishing their retinal axon guidance mission at the midline. So it is reasonable that some of the earliest born chiasm cells disappear during development. / During development, retinal ganglion cell axons grow from the eye to the ventral diencephalon, where axons from the two eyes converge and segregate into crossed and uncrossed projections, forming the optic chiasm. This pattern is critical for binocular vision. Although significant progress has been obtained over the past decades, how retinal axon growth and guidance are regulated at the chiasm is largely unknown. Our research will focus on those problems. / In the last part of this thesis, we investigated the retinal axon pathway in the ventral diencephalon of the Sox10Dom mutant embryos and gamma-crystallin mutant embryos. Our findings indicate that Sox10 may not contribute to axon guidance in the developing optic pathway whereas gammaA-crystallin may only play a role in the later uncrossed axons. / N-methyl-D-aspartate (NMDA) receptor is one of the ionotropic glutamate receptors, which are important in synaptic plasticity, apart from implications in dendritic spine remodeling, neurite outgrowth, elongation and branching and glutamate neurotoxicity. There are several subtypes of NMDA receptor channel subunits, NR1, NR2A-D, NR3A&B. The functional diversity of NMDA receptor resides in the different assembly of subunits. In this study, we used RT-PCR to analyze the mRNA expression of all the NMDA receptor subunits in mouse embryos. After that we chose the NR1, NR2B and NR3A antibodies to investigate NMDA receptor subunit expression in the optic pathway during mouse optic pathway development. Using immunohistochemistry, we found that NR1, NR2B and NR3A were expressed in the mouse retina and optic pathway as from E13 when the optic chiasm is forming. Expression of the NMDA receptor subunits were found in the inner cell layers and along retinal axons. Colocalization studies showed that NR1, NR2B and NR3A were localized on the ganglion cells and their axons. In the ventral diencephalon, these subunits were expressed extensively, but NR1 and NR3A were particularly strong along the optic nerve and optic tract. Furthermore, to identify the function of NMDA receptor during optic chiasm development, we cultured E14 retinal explants on laminin and poly-D-ornithine in the presence of the NMDA receptor antagonists MK-801 or Dextrorphan-D-tartrate. These two antagonists can significantly inhibit the retinal axon outgrowth, suggesting that the NMDA receptor promotes retinal axon outgrowth in the retinofugal pathway during optic chiasm development. / Li, Jia. / Adviser: Chan Sun On. / Source: Dissertation Abstracts International, Volume: 73-02, Section: B, page: . / Thesis (Ph.D.)--Chinese University of Hong Kong, 2010. / Includes bibliographical references (leaves 145-158). / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Electronic reproduction. [Ann Arbor, MI] : ProQuest Information and Learning, [201-] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Abstract also in Chinese.

Hao Yanli. / "July 2004." / Thesis (Ph.D.)--Chinese University of Hong Kong, 2004. / Includes bibliographical references (p. 113-134). / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Mode of access: World Wide Web. / Abstracts in English and Chinese.

by Ling Lin. / "November 2002." / Thesis (Ph.D.)--Chinese University of Hong Kong, 2002. / Includes bibliographical references (p. 149-168). / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Mode of access: World Wide Web. / Abstracts in English and Chinese.

Chez les animaux à vision binoculaire, la vision tridimensionnelle permet la perception de la profondeur grâce à l'intégration de l'information visuelle en provenance des deux yeux. La première étape de cette intégration est rendue possible anatomiquement par la ségrégation des axones controlatéraux et ipsilatéraux des cellules ganglionnaires de la rétine (CGR) au niveau du chiasma optique. Les axones controlatéraux croisent la ligne médiane au chiasma en route du nerf optique vers le cerveau. À l’inverse, les axones ipsilatéraux s'écartent du chiasma et continuent dans le tractus optique ipsilatéral, en évitant la ligne médiane vers leurs cibles cérébrales. Les mécanismes moléculaires à la base de ce phénomène ne sont pas complètement compris. Les études présentées dans cette thèse montrent que Boc, le récepteur de Sonic Hedgehog (Shh) dans le guidage axonal, est enrichi dans les CGRs ipsilatérales de la rétine en développement. La présence de Shh sur la ligne médiane, et le mode d'expression complémentaire du récepteur nous ont conduit à émettre l'hypothèse que Shh pourrait repousser les axones ipsilatéraux au niveau du chiasma en activant le récepteur Boc. Conformément à cette hypothèse, nous avons constaté que seulement les CGR exprimant Boc se rétractent in vitro en réponse à Shh et que cette réponse est perdue dans les CGR mutantes pour Boc. In vivo, nous démontrons que Boc est requis pour la ségrégation normale des axones ipsilatéraux au niveau du chiasma optique et, inversement, que l'expression ectopique de Boc dans les CGR contralatérales empêche leurs axones de traverser le chiasma optique. Dans l’ensemble, ces résultats suggèrent que Shh repousse les axones ipsilatéraux au niveau du chiasma optique par son récepteur Boc. Cette première partie de notre travail identifie un nouveau couple ligand-récepteur requis pour la ségrégation des axones au niveau du chiasma optique. Une interaction moléculaire impliquée dans cette ségrégation implique l’éphrine-B2 et ses récepteurs EphB (EphB1). Dans la deuxième partie de notre travail, nous montrons, in vivo, en utilisant des souris doubles et quadruples mutantes pour les récepteurs Boc, EphB1 ou les trois récepteurs EphB, que l’abrogation des deux voies de signalisation Shh et éphrine-B2 conduit à l'absence de projections ipsilatérales. Ceci indique que les deux signalisations agissent de façon indépendante dans des voies parallèles. De manière intéressante, ces souris mutantes ont été utilisées comme modèle génétique pour démontrer des défauts dans la perception de la profondeur de champs chez des animaux dépourvus de projections visuelles ipsilatérales. Ainsi, les travaux présentés dans cette thèse démontrent pour la première fois que la formation des projections rétiniennes ipsilatérales est essentielle à l’établissement de la vision binoculaire et dépend des voies induites par les récepteurs d’éphrine-B2 et Shh. / In animals with binocular vision, three dimensional vision allows perception of depth through the integration of visual information from both eyes. The first step of this integration is possible anatomically with the segregation of contralateral and ipsilateral axons at the optic chiasm. 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. The molecular mechanism underlying this phenomenon is not completely understood. The studies presented in this thesis 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. Taken together, these results suggest that Shh repels ipsilateral RGC axons at the optic chiasm via its receptor Boc. This first part of this thesis identifies a novel receptor required for the segregation of axons at the optic chiasm. The other couple ligand-receptor involved in this segregation is the Ephrin-B2/EphB signalling. In the second part of this thesis, I show that in vivo, the abrogation of both signalling pathways using quadruple knockout mice of the receptor Boc and three EphB receptors led to the absence of ipsilateral projections, indicating that Shh and ephrinB2 signalling act independently in two parallel pathways. More importantly, these animals, used as a new genetic model to perform visual tests, had a diminished ability to perceive depth. Thus, this thesis demonstrates for the first time that the establishment of ipsilateral retinal projections, essential for accurate binocular vision and perception of depth, is made possible by the combination of EphB and Shh signalling.