published_or_final_version / Zoology / Master / Master of Philosophy

Lau Man Tat. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2003. / Includes bibliographical references (leaves 95-126). / Abstracts in English and Chinese. / Abstract (in English) --- p.i / Abstract (in Chinese) --- p.iii / Acknowledgements --- p.v / Table of Contents --- p.vi / List of Figures --- p.xi / List of Tables --- p.xiii / Symbols and Abbreviations --- p.xiv / Scientific Names --- p.xvii / Chapter Chapter 1 --- General Introduction / Chapter 1.1 --- Gonadotropins / Chapter 1.1.1 --- Structure --- p.2 / Chapter 1.1.2 --- Function --- p.6 / Chapter 1.1.3 --- Regulation --- p.9 / Chapter 1.1.3.1 --- Hypothalamic regulators (GnRH) --- p.9 / Chapter 1.1.3.2 --- Endocrine regulators from gonads (steroids) --- p.12 / Chapter 1.1.3.3 --- Paracrine regulators (activin) --- p.14 / Chapter 1.2 --- Activin Family of Growth Factors / Chapter 1.2.1 --- Activin / Chapter 1.2.1.1 --- Structure --- p.14 / Chapter 1.2.1.2 --- Function --- p.14 / Chapter 1.2.1.3 --- Signaling --- p.18 / Chapter 1.2.2 --- Follistatin / Chapter 1.2.2.1 --- Structure --- p.21 / Chapter 1.2.2.2 --- Function --- p.21 / Chapter 1.3 --- Transcriptional regulation of pituitary gonadotropin subunit genes at the promoter level --- p.22 / Chapter 1.4 --- The project objectives and long-term significance --- p.26 / Chapter Chapter 2 --- "Cloning of Smad2, Smad3, Smad4 and Smad7 from the Goldfish Pituitary and Their Involvement in the FSHβ Transcription in LβT2 cells" / Chapter 2.1 --- Introduction --- p.28 / Chapter 2.2 --- Materials and Methods / Chapter 2.2.1 --- Chemicals --- p.31 / Chapter 2.2.2 --- Animals --- p.32 / Chapter 2.2.3 --- Isolation of total RNA --- p.32 / Chapter 2.2.4 --- "Cloning of cDNA fragments of Smad 2, 3, 4 and 7 from the goldfish pituitary" --- p.32 / Chapter 2.2.5 --- Rapid amplification of 5'-cDNA ends (5'-RACE) and full-length cDNA(3'-RACE) --- p.33 / Chapter 2.2.6 --- Primary pituitary cell culture --- p.34 / Chapter 2.2.7 --- "Validation of semi-quantitative reverse transcription-polymerase chain reaction (RT-PCR) assays for goldfish Smad 2, 3, 4 and7" --- p.35 / Chapter 2.2.8 --- Construction of the reporter plasmid containing the goldfish FSHβ promoter --- p.36 / Chapter 2.2.9 --- Construction of expression plasmids --- p.37 / Chapter 2.2.10 --- Cell culture and transient transfection --- p.38 / Chapter 2.2.11 --- SEAP reporter gene assay --- p.39 / Chapter 2.2.12 --- β-galactosidase reporter gene assay --- p.40 / Chapter 2.2.13 --- Data analysis --- p.40 / Chapter 2.3 --- Results / Chapter 2.3.1 --- "Cloning and sequence characterization of goldfish Smad 2, 3,4 and7" --- p.41 / Chapter 2.3.2 --- "Tissue distribution of Smad 2，3, 4 and 7 expression" --- p.42 / Chapter 2.3.3 --- "Validation of semi-quantitative RT-PCR assays for Smad 2, 3,4 and7" --- p.43 / Chapter 2.3.4 --- Activin regulation of Smad 2，3，4 and 7 expression in cultured goldfish pituitary cells --- p.44 / Chapter 2.3.5 --- "Smad 2, 3 and 7 regulate basal and activin-induced FSHβ transcription in LβT2 cells" --- p.44 / Chapter 2.3.6 --- Autocrine regulation of the gfFSHβ transcription by activin in LβT2 cells --- p.45 / Chapter 2.4. --- Discussion --- p.47 / Chapter Chapter 3 --- Promoter Analysis for the Smad Responsive Element (SRE) in the Goldfish Follicle Stimulating Hormone β(FSHβGene / Chapter 3.1 --- Introduction --- p.71 / Chapter 3.2 --- Materials and Methods / Chapter 3.2.1 --- Chemicals --- p.74 / Chapter 3.2.2 --- Construction of expression plasmids --- p.74 / Chapter 3.2.3 --- Construction of SEAP reporter plasmids containing different lengths of gfFSHβ promoter --- p.74 / Chapter 3.2.4 --- Cell culture and transient transfection --- p.75 / Chapter 3.2.5 --- Reporter gene assays for SEAP and β-Gal --- p.75 / Chapter 3.2.6 --- Data Analyses --- p.76 / Chapter 3.3 --- Results / Chapter 3.3.1 --- Localization of the proximal Smad-responsive Element (SRE) in the gfFHSβ promoter --- p.76 / Chapter 3.4 --- Discussion --- p.78 / Chapter Chapter 4 --- General Discussion / Chapter 4.1 --- Overview --- p.89 / Chapter 4.2 --- Contribution of the present research / Chapter 4.2.1 --- Cloning and characterization of Smad proteins from the goldfish pituitary --- p.90 / Chapter 4.2.2 --- Regulation of Smads in primary pituitary cell culture --- p.90 / Chapter 4.2.3 --- Identification of the Smad responsive element (SRE) on the gfFSHβ promoter --- p.91 / Chapter 4.3 --- Future research direction --- p.93 / References --- p.95

Helen Wei Ling Lai. / "December 2000." / Thesis (Ph.D.)--Chinese University of Hong Kong, 2000. / includes bibliographical references (p. 97-102). / 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.

No description available.

Reorganization of the intermediate filament (IF) network during axonal regeneration is accompanied by changes in the expression of various IF proteins. An increase in expression of the neuronal IF subunit gefiltin in goldfish retinal ganglion cells (RGCs) has been linked to the unique ability of the goldfish optic nerve to regenerate following injury. Evidence suggests that the optic tectum may regulate the expression of gefiltin during regeneration. The goal of this thesis was to determine the function of the tectum in the regulation of gefiltin mRNA expression during optic fiber regeneration in the goldfish. It was found that gefiltin mRNA levels in the RGCs of animals that received an optic nerve crush (ONC group) increased by 10 days, peaked from 20 to 38 days at around 5.5-fold over normal, and declined to near normal by 115 days. In animals that had the entire tectum removed and an optic nerve crush (ETR group), gefiltin mRNA levels increased by 10 days, peaked at 20 days at around 5.5- to 6.5-fold over normal, and although they dropped slightly thereafter, they remained elevated at around 5-fold over normal for at least 115 days. When axons regenerated to the ipsilateral tectal lobe as a result of a left tectal lobe removal and left eye removal surgery the expression pattern of gefiltin mRNA paralleled that of the ONC group. It was also found that the abundance of gefiltin subunits in the retina was elevated at 30 days of regeneration in ONC and ETR animals, and that levels in the nerve were reconstituted to 80% of normal by 30 days. These results demonstrate that increases in gefiltin mRNA and protein levels during optic nerve regeneration are independent of the tectum, whereas the downregulation of gefiltin mRNA levels is entirely dependent upon the tectum. (Abstract shortened by UMI.)

Thesis (M. Phil.)--University of Hong Kong, 2000. / Includes bibliographical references (leaves 62-76).

Thesis (M.Phil.)--University of Hong Kong, 2000. / Includes bibliographical references (leaves 62-76) Also available in print.

Nestin is a type VI intermediate filament protein that marks proliferative cells in the central and peripheral nervous system of vertebrates during development and adulthood. Nestin is not only expressed in progenitor cells of neuronal tissues but is also present in muscle, heart, lung, pancreas and skin follicle tissues. The goal of this thesis is to investigate and characterize the nestin protein in goldfish and relate nestin expression to neuroregeneration and brain plasticity events in the adult goldfish forebrain. Currently little is known about nestin function and regulation in vertebrates, especially in fish. In this study we used Rapid amplification of cDNA ends PCR (RACE-PCR) to isolate goldfish nestin mRNA. We uncovered several different mRNA transcripts. PCR analysis and sequencing further identified three different nestin transcripts of 4003, 2446, and 2126 nucleotides with a predicted protein length of 860, 274, and 344 amino acids respectively. We next applied a multiple-antigenic peptide (MAP) strategy to generate a polyclonal goldfish-specific nestin antibody against a 23 amino acid sequence located at the N-terminal end of goldfish nestin. Western blotting revealed the existence of three different nestin protein isoforms (nestin A, B and C); the first report of nestin isoforms in teleost species. Nestin expression and distribution in the goldfish brain is complex and revealed both individual and tissue-dependent variations. The most remarkable finding following principal component analysis of the western blot data was the uniqueness of the pituitary, hypothalamus and telencephalon. These tissues are proliferative in nature containing progenitor and proliferative cellular pools that are involved in important biological axes such as the motor and reproductive axis. Interestingly, all three tissues were able to change their proliferative cellular profile of nestin protein expression to alleviate the detrimental effects of the neurotoxin 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) upon administration. The toxin MPTP destroys dopamine neurons in the fish brain leading to motor deficits and reproductive difficulties. The incorporation of 5-bromo-2’-deoxyuriding (BrdU) into newly synthesized DNA revealed an upregulation of BrdU immunolabeling following MPTP administration in the area telencephali pars dorsalis (Vd) and along the ventricular surface area of the telencephalon suggesting the generation of new neurons in the adult central nervous system. This thesis reports novel nestin isoforms and illustrates regenerative events occurring in the goldfish telencephalon following a neurotoxic insult. This work provides a framework for future investigations of the differential roles and regulation of the nestins to better understand seasonal neuronal plasticity, neuronal regeneration and neuronal circuitry in teleost.

No description available.

Starch gel electrophoresis and DNA fingerprinting were used to assess molecular genetic variation within and between strains of goldfish, Carassius auratus. Genetic variation observed both within and between strains was low using both methods. using isozyme markers, mean heterozygosity (0.0% - 4.4%) and percent polymorphic loci (0.0% - 30.6%) within strains were low compared to other vertebrate species. Analysis of the isozyme data using Chi-square tests against Hardy-Weinberg genotype expectations and wright's (1943, 1951) F statistics indicated modest heterogeneity in genotype frequencies between strains. DNA fingerprint band sharing values were determined both within and between strains of goldfish. Mean band sharing values within strains were high (68.0% -97.6%), comparable to values observed in highly inbred populations of other vertebrates. Levels of band sharing between strains were determined using a DNA mixing procedure. Band sharing values between strains (57.7% - 100%) were higher than those observed between strains of other domesticated species. Analysis of DNA fingerprint data using ANOVA indicated no significant differences between strains, suggesting that the fancy strains did not exhibit significant differentiation as manifested in DNA fingerprint phenotypes. / Master of Science