Kwok Hin-Fai. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2004. / Includes bibliographical references (leaves 84-100). / Abstracts in English and Chinese. / Abstract (in English) --- p.i / Abstract (in Chinese) --- p.iv / Acknowledgement --- p.vi / Table of contents --- p.vii / List of figures --- p.xi / List of tables --- p.xiv / Symbols and abbreviations --- p.xv / List of fish names mentioned in the thesis --- p.xviii / Chapter Chapter 1 --- General Introduction / Chapter 1.1 --- Gonadotropins / Chapter 1.1.1 --- Structure --- p.1 / Chapter 1.1.2 --- Function --- p.4 / Chapter 1.2 --- Gonadotropin receptor / Chapter 1.2.1 --- Structure --- p.5 / Chapter 1.2.2 --- Expression --- p.7 / Chapter 1.2.3 --- Signaling / Chapter 1.2.3.1 --- cAMP-mediated pathway --- p.7 / Chapter 1.2.3.2 --- Phospholipase C-mediated pathway --- p.9 / Chapter 1.2.4 --- Regulation of expression --- p.12 / Chapter 1.2.5 --- Desensitization of receptors / Chapter 1.2.5.1 --- Uncoupling --- p.13 / Chapter 1.2.5.2 --- Internalization --- p.13 / Chapter 1.3 --- Structure of ovarian follicles --- p.14 / Chapter 1.4 --- The project objectives and long-term significance --- p.16 / Chapter Chapter 2 --- Cloning and Characterization of Zebrafish Follicle-stimulating Hormone (FSH) and Luteinizing Hormone (LH) Receptors ´ؤ Evidence for Distinct Functions of FSH and LH in Follicle Development / Chapter 2.1 --- Introduction --- p.19 / Chapter 2.2 --- Materials and Methods / Chapter 2.2.1 --- Animals and chemicals --- p.22 / Chapter 2.2.2 --- Isolation of total RNA --- p.22 / Chapter 2.2.3 --- Cloning of zebrafish FSHR (zfFSHR) and LHR (zfLHR) cDNA fragments from the zebrafish ovary --- p.23 / Chapter 2.2.4 --- Rapid amplification of 5´ةcDNA ends (5'-RACE) and full-length cDNA --- p.24 / Chapter 2.2.5 --- Isolation of ovarian follicles --- p.25 / Chapter 2.2.6 --- Sampling of the ovaries from sexually immature zebrafish --- p.25 / Chapter 2.2.7 --- Reverse transcription-polymerase chain reaction (RT-PCR) --- p.25 / Chapter 2.2.8 --- Construction of expression plasmids --- p.26 / Chapter 2.2.9 --- Transient transfection and reporter gene assay --- p.27 / Chapter 2.2.10 --- Establishment and characterization of stable zfFSHR or zfLHR-expressing cell lines --- p.28 / Chapter 2.3 --- Results / Chapter 2.3.1 --- Cloning of FSHR and LHR cDNA from the zebrafish ovary --- p.29 / Chapter 2.3.2 --- Functional characterization of zfFSHR and zfLHR --- p.30 / Chapter 2.3.3 --- Expression of zfFSHR and zfLHR during sexual maturation --- p.31 / Chapter 2.3.4 --- Stage-dependent expression of zfFSHR and zfLHR in the ovarian follicles --- p.32 / Chapter 2.4 --- Discussion --- p.33 / Chapter Chapter 3 --- Down-regulation of FSHR and LHR Expression in the Zebrafish Follicle Ceils by Gonadotropin (hCG) and Its Sigaling Mechanism / Chapter 3.1 --- Introduction --- p.51 / Chapter 3.2 --- Materials and Methods / Chapter 3.2.1 --- Animals --- p.54 / Chapter 3.2.2 --- Chemicals and hormones --- p.54 / Chapter 3.2.3 --- Primary follicle cell culture --- p.55 / Chapter 3.2.4 --- Total RNA isolation --- p.55 / Chapter 3.2.5 --- "Validation of semi-quantitative RT-PCR assays for FSHR, LHR and GAPDH" --- p.56 / Chapter 3.2.6 --- Data analysis --- p.57 / Chapter 3.3 --- Results / Chapter 3.3.1 --- Validation of semi-quantitative RT-PCR assays --- p.57 / Chapter 3.3.2 --- Gonadotropin regulation of FSHR and LHR expression in cultured zebrafish ovarian follicle cells --- p.58 / Chapter 3.3.3 --- Effect of db-cAMP and forskolin on FSHR and LHR expression --- p.59 / Chapter 3.3.4 --- Effects of H89 on hCG-induced suppression of FSHR and LHR expression --- p.60 / Chapter 3.4 --- Discussion --- p.60 / Chapter Chapter 4 --- General Discussion --- p.75 / Chapter 4.1 --- Cloning of zebrafish FSHR and LHR cDNAs and demonstration of receptor specificity --- p.77 / Chapter 4.2 --- Evidence for the differential expression of FSHR and LHR in the zebrafish ovarian and follicle development --- p.78 / Chapter 4.3 --- Down-regulation of FSHR and LHR expression in the zebrafish follicle cells by gonadotropin (hCG) --- p.79 / Chapter 4.4 --- Future research direction --- p.80 / References --- p.84

So Wai-Kin. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2004. / Includes bibliographical references (leaves 100-127). / Abstracts in English and Chinese. / Acknowledgement --- p.I / Abstract (in English) --- p.II / Abstract (in Chinese) --- p.IV / Table of contents --- p.VI / List of Figures --- p.X / Symbols and Abbreviations --- p.XII / List of fish names mentioned in the thesis --- p.XIV / Chapter Chapter 1 --- General Introduction / Chapter 1.1 --- Pituitary --- p.1 / Chapter 1.2 --- Gonadotropins --- p.1 / Chapter 1.2.1 --- Structure --- p.2 / Chapter 1.2.2 --- Signaling --- p.3 / Chapter 1.2.3 --- Expression --- p.5 / Chapter 1.2.4 --- Functions --- p.7 / Chapter 1.2.4.1 --- Gonadotropin actions on gametogenesis --- p.7 / Chapter 1.2.4.2 --- Gonadotropin actions on steroidogenesis --- p.8 / Chapter 1.2.5 --- Regulation --- p.9 / Chapter 1.2.5.1 --- Neuroendocrine control --- p.10 / Chapter 1.2.5.1.1 --- Gonadotropin-releasing hormone (GnRH) --- p.10 / Chapter 1.2.5.1.2 --- Dopamine (DA) --- p.12 / Chapter 1.2.5.2 --- Gonadal steroid feedback --- p.12 / Chapter 1.2.5.2.1 --- Positive feedback --- p.13 / Chapter 1.2.5.2.2 --- Negative feedback --- p.14 / Chapter 1.2.5.3 --- Paracrine regulators within pituitary --- p.15 / Chapter 1.3 --- Objectives of the present study --- p.16 / Chapter Chapter 2 --- "Molecular Cloning and Functional Characterization of Zebrafish FSHβ, LHβ and GTHα subunits" / Chapter 2.1 --- Introduction --- p.19 / Chapter 2.2 --- Materials and methods --- p.21 / Chapter 2.2.1 --- Chemicals --- p.21 / Chapter 2.2.2 --- Animals --- p.21 / Chapter 2.2.3 --- Genomic DNA isolation --- p.22 / Chapter 2.2.4 --- Total RNA isolation --- p.22 / Chapter 2.2.5 --- Cloning of zebrafish FSHp，LHβ and GTHa fragments --- p.23 / Chapter 2.2.5.1 --- LHβ and GTHα --- p.23 / Chapter 2.2.5.2 --- FSHβ --- p.23 / Chapter 2.2.6 --- "5'- and 3'-RACE of zebrafish FSHp, LHβ and GTHα subunits" --- p.24 / Chapter 2.2.7 --- Construction of expression constructs --- p.25 / Chapter 2.2.8 --- Cell culture and transfection of Flp-In´ёØ CHO cell --- p.26 / Chapter 2.2.9 --- Recombinant production of zebrafish FSH and LH --- p.27 / Chapter 2.2.10 --- Reverse transcription-polymerase chain reaction (RT-PCR) analysis --- p.27 / Chapter 2.2.11 --- Northern blot hybridization --- p.28 / Chapter 2.2.12 --- SEAP reporter gene assay --- p.28 / Chapter 2.2.13 --- Data analysis --- p.29 / Chapter 2.3 --- Results --- p.30 / Chapter 2.3.1 --- "Cloning of zebrafish FSHβ, LHβ and GTHα subunits" --- p.30 / Chapter 2.3.2 --- "Expression of zebrafish FSHp, LHβ and GTHα in the zebrafish pituitary" --- p.31 / Chapter 2.3.3 --- Recombinant production of zebrafish FSH and LH --- p.32 / Chapter 2.3.4 --- Functional analysis of zebrafish FSH and LH --- p.33 / Chapter 2.4 --- Discussion --- p.34 / Chapter Chapter 3 --- "Spatial Expression Patterns of Zebrafish FSHβ, LHβ and GTHα Subunits in the Pituitary and Their Temporal Expression Profiles during Sexual Maturation and Ovulatory Cycle" / Chapter 3.1 --- Introduction --- p.58 / Chapter 3.2 --- Materials and methods --- p.61 / Chapter 3.2.1 --- Chemicals --- p.61 / Chapter 3.2.2. --- Animals --- p.62 / Chapter 3.2.3 --- Total RNA isolation from zebrafish pituitaries and reverse transcription --- p.62 / Chapter 3.2.4 --- Validation of RT-PCR on single pituitary --- p.63 / Chapter 3.2.5 --- Real-time PCR --- p.64 / Chapter 3.2.6 --- Tissue preparation for in situ hybridization --- p.64 / Chapter 3.2.7 --- In situ hybridization --- p.65 / Chapter 3.2.8 --- Data analysis --- p.66 / Chapter 3.3 --- Results --- p.66 / Chapter 3.3.1 --- "PCR amplification of FSHβ, LHβ and GTHα and GAPDH in single zebrafish pituitary" --- p.67 / Chapter 3.3.2 --- "Establishement of real-time RT-PCR for zebrafish FSHβ, LHβ and GTHa and GAPDH" --- p.67 / Chapter 3.3.3 --- "Temporal expression profiles of zebrafish FSHβ, LHβ and GTHα subunits during sexual maturation" --- p.67 / Chapter 3.3.4 --- "Temporal expression profiles of zebrafish FSHp, LHβ and GTHα subunits during ovulatory cycle" --- p.68 / Chapter 3.3.5 --- "In situ hybridization of zebrafish FSHβ, LHβ and GTHα" --- p.69 / Chapter 3.4 --- Discussion --- p.70 / Chapter Chapter 4 --- General Discussion / Chapter 4.1 --- Cloning of zebrafish gonadotropin subunit cDNAs --- p.91 / Chapter 4.2 --- Bioactivity and receptor specificity of recombinant zebrafish FSH and LH --- p.91 / Chapter 4.3 --- Expression of gonadotropin subunits during zebrafish sexual maturation and ovulatory cycle --- p.92 / Chapter 4.4 --- "Localization of FSHβ, LHβ and GTHα subunits in zebrafish pituitary" --- p.93 / Chapter 4.5 --- Contributions of the present study --- p.94 / Chapter 4.6 --- Future prospects --- p.95 / References --- p.100

Thesis (Ph.D.) -- University of Texas Southwestern Medical Center at Dallas, 2005. / Not embargoed. Vita. References located at the end of each chapter.