Shi, Yujian. / Thesis (Ph.D.)--Chinese University of Hong Kong, 2013. / Includes bibliographical references (leaves 83-101). / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Abstracts also in Chinese.

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

Temperature effects on the development of the zebrafish embryos and larvae and adults were examined. It was found that the earlier in development a temperature change was performed on an embryo, the more significant the change in survival and/or subsequent development. Thus, viable temperature ranges for zebrafish widened significantly as development proceeded. Adults reared and bred at 25oC produced embryos that were significantly more successful at the lower range of rearing temperatures compared to embryos produced from adults reared at 28oC. The majority of this study focused on the physiological effects of swim training during development in the zebrafish. The earlier in development the zebrafish larvae were trained, the greater the mortality. Trained free swimming larvae had a significantly higher routine oxygen consumption after 11 days of training, and a higher mass specific routine metabolic rate after 8 and 11 days of training. Trained free swimming larvae consumed significantly less oxygen during swimming and were more efficient at locomotion, compared to control larvae. Training enhanced survival during exposure to extreme hypoxia in all age groups. Performance aspects of training were investigated in attempt to quantify training effects and in most cases, trained fish performed significantly better than controls. As blood vessels formed during development, they decreased in cross sectional area from days two to six. It was also shown that the variability in visual stroke volume measurements could be reduced significantly by using a third dimension in the analysis with a more accurate volume equation. Finally, the ontogeny of cardiac control was evaluated. The adrenergic receptors were the first to respond to pharmacological stimulation but were closely followed by cholinergic pharmacological stimulation a few days later. There was a significant cholinergic tone present in day 15 zebrafish larvae which persisted. Although an adrenergic tone was not documented in this study, this does not prove its lack of existence.

Superior swimming stamina in adult fish is presumably passed on to their offspring, but the ontogeny of the appearance of superior stamina and the requisite enhanced cardio-respiratory support for locomotion in larval fishes has not been determined. Is the expression of the suite of parental traits enabling superior swimming stamina in their offspring dependent upon their achieving juvenile/adult morphology, or does it appear earlier in their larvae? To answer this, adults were classified into three groups based on swimming stamina, followed by measurement of length, mass, and width. Larval offspring from the two parental groups -high stamina larvae (HSL) and low stamina larvae (LSL)- were reared at 27°C in aerated water (21% O2). Routine and active heart rate, routine and active mass specific oxygen consumption were recorded through 21dpf, and cost of transport (COT) and factorial aerobic scope were derived from oxygen consumption measurements. Routine heart rate at 2dpf of LSL was 164 ± 1 b·min-1, compared to only 125 ± 2 b·min-1 for HSL. Routine heart rate subsequently peaked at 203 ± 1 b·min-1 at 5dpf in the HSL group, compared to 207 ± 1 b·min-1, at 4dpf in the LSP larvae. Active heart rate at 5 dpf of LSL was 218 ± 2 b·min-1 compared to 216 ± 2 b·min-1 for HSL. Active heart rate increased slightly to 227 ± 2 b·min-1 for LSL before decreasing again, while active heart rate remained relatively constant for HSL. Routine O2 consumption at 2dpf of HSL was 0.09 μmol·mg-1·hr-1, compared to 0.03 μmol·mg-1·hr-1 in LSL. Routine O2 consumption subsequently peaked at 0.70 μmol·mg-1·hr-1 at 9dpf in the HSL, compared to 0.71 μmol·mg-1·hr-1, at 9dpf in the LSL. These values dramatically decreased before leveling off at around 0.20 μmol·mg-1·hr-1 and 0.15 μmol·mg-1·h-1, respectively. Active O2 consumption at 5dpf for HSL was 0.38 μmol·mg-1·hr-1, compared to 0.57 μmol·mg-1·hr-1 for LSL. Active O2 consumption subsequently peaked at 0.97 μmol·mg-1·hr-1 at 10dpf in HSL, compared to 1.19 μmol·mg-1·hr-1 at 7dpf in LSL. These values also dramatically decreased and leveled off. Significant differences (p < 0.05) in heart rate and oxygen consumption persisted through 21dpf. The onset of differences observed in routine and active heart rate in early larvae, correlated with parent stamina, show that juvenile or adult features are not required as a precondition for the emergence of phenotypic physiological differences.

Bibliography: leaves 129-158. / xvii, 159, [11] leaves, [24] leaves of plates : ill. (some col.) ; 35 cm. / Title page, contents and abstract only. The complete thesis in print form is available from the University Library. / The major estrogen inducible protein in zebra fish liver has been purified to homogeneity by FPLC using anion exchange chromatography (Mono-Q Pharmacia) with purification being monitored by SDS-PAGE electrophoresis. / Thesis (Ph.D.)--University of Adelaide, Dept. of Obstetrics and Gynaecology, 1995?

Indiana University-Purdue University Indianapolis (IUPUI) / Bone growth can be impaired due to disease, such as osteoporosis. Currently, intermittent parathyroid hormone (PTH) treatment is the only approved therapy in the United States for anabolic bone growth in osteoporosis patients. The anabolic effects of PTH treatment are due, at least in part, to modulation of the Wnt/β-catenin pathway. Activation of the Wnt/ β-catenin pathway using a small molecule inhibitor of GSK3β was previously shown to increase markers of bone formation in vitro. Our study utilized a zebrafish model system to study Wnt activated fin regeneration and bone growth. Wnt signaling is the first genetically identified step in fin regeneration, and bony rays are the main structure in zebrafish fins. Thus, zebrafish fin regeneration may be a useful model to study Wnt signaling mediated bone growth. Fin regeneration experiments were conducted using various concentrations of a GSK3β inhibitor compound, LSN 2105786, for different treatment periods and regenerative outgrowth was measured at 4 and 7 days post amputation. Experiments revealed continuous low concentration (4-5 nM) treatment to be most effective at increasing regeneration. Higher concentrations inhibited fin growth, perhaps by excessive stimulation of differentiation programs. In situ hybridization experiments were performed to examine effects of GSK3β inhibitor on Wnt responsive gene expression. Experiments showed temporal and spatial changes on individual gene markers following GSK3β inhibitor treatment. Additionally, confocal microscopy and immunofluorescence labeling data indicated that the Wnt signaling intracellular signal transducer, β-catenin, accumulates throughout GSK3β inhibitor treated tissues. Finally, experiments revealed increased cell proliferation in fin regenerates following LSN 2105786 treatment. Together, these data indicate that bone growth in zebrafish fin regeneration is improved by activating Wnt signaling. Zebrafish Wnt signaling experiments provide a good model to study bone growth and bone repair mechanisms, and may provide an efficient drug discovery platform.