Prostanoid receptors (PG-Rs: prostaglandin D, E, F, prostacyclin and thromboxane receptors (DP, EP1-4, FP, IP and TP)) are known to mediate a diverse range of biological responses, such as cardiovascular homeostasis, nociception and reproduction, via binding to their respective ligands belonging to the five classes of prostanoids (PGs: class D, E, F, I and thromboxane). The majority of these findings were reported in mammals, and despite suggestive evidence provided by previous pharmacological and physiological studies in non-mammalian vertebrates, investigations on the mechanisms behind actions of PGs were impeded by the limited information on their receptors. In the present study, the full-length cDNAs of chicken (c-) and zebrafish (z-) prostanoid receptors – cEP3, cFPs, zEP1s and zFP – were identified from respective adult ovaries and their tissue distribution examined by RT-PCR. A novel middle-truncated splice variant, cFPb, which lacks 107 amino acids between transmembrane domains 4 and 6 but otherwise identical to cFPa was first identified. Three isoforms of zEP1 – zEP1a, zEP1b, zEP1c – were found, which might have subfunctionalized in their ligand binding and G protein coupling specificity, in addition to differential tissue distribution. Using various luciferase reporter systems (pGL3-CRE, pGL-NFAT-RE, pGL4-SRE), all the cloned receptors, except cFPb, were shown to potentially couple to intracellular cAMP, Ca2+, and/or MAPK signaling pathways. Owing to the proposed roles of PGs and its potential regulation by and/or on EGFR ligands and gonadotropins in mammals and chicken, genes involved in regulation of PG functions at various levels, including biosynthesis (COX1, COX2, mPGES1, mPGES2 and cPGES), availability (PGT) and signaling (cEPs and cFPs), were also characterized in granulosa cells during hen follicular development. Lastly, using our experimental data and systematic sequence retrieval from available databases, the PG receptor cascades from representative vertebrate species were pooled and analysed using phylogenetic analyses and synteny studies. Three putative clusters (IP-like, EP4-like and EP1-like cluster) were found in lamprey genome; meanwhile, only one PG-R-like cluster was identified from the Cephalochordate lancelet (amphioxus) genome. This concurs with the 1-2-4 rule proposed in first round/second round (1R/2R) whole genome duplication in which the missing lamprey cluster was presumably lost secondarily. With support from conserved orthologs-localization, the four PG-R paralogs (proto-EP4, proto-IP/EP2/DP, proto-TP/FP/EP1 & putative proto-EP3 genes) in the ancestral vertebrates might have further diversified via either localized- (e.g. EP2 and DP) or chromosomal segmental duplication (e.g. EP1, FP and TP) which resulted in the present array of vertebrate PGRs. Additional paralogs (e.g. EP1 and EP4) were identified from fishes, by which molecular dating coincide with and hint of their origins whence the ancient fishspecific whole genome duplication (3R) occurred ~350 million years ago. The present study offers the first glimpse and a better understanding of the roles of the PG-Rs and presents a higher resolution to the evolutionary history of each PG-R family member, consolidating that particular care has to be taken when studying non-mammalian PG-R functions in which some members are absent or present in multiples and propel the investigation of adaptational changes in the coding sequence during evolution of vertebrate PG-Rs. / published_or_final_version / Biological Sciences / Doctoral / Doctor of Philosophy

CD4+ lymphocytes (T helper cells) play a crucial role in orchestrating adaptive immune responses. This project aims to characterise the CD4+ cells in zebrafish (Danio rerio) for a better understanding of adaptive immunity in teleost fish. I cloned three CD4 homologues, termed zfCD4 and zfCD4 related (zfCD4rel1 and zfCD4rel2), with both reported previously in other bony fish species. The zfCD4 and zfCD4rels transcripts were detected in immune organs in zebrafish and were most highly expressed in the lymphocyte population. A moderate induction of the zfCD4 and zfCD4rels (and other immune related genes) was seen in kidney, spleen and intestine after poly (I:C) injection. Two antibodies against CD4 and IFN-γ, respectively, have been validated using various immunostaining approaches for further functional studies. The CD4 positive cells ranged from 20-30% of lymphocytes in zebrafish, similar to what is seen in other vertebrates. The expression level of IFN-γ and other Th cell related genes were analysed in immunised fish following re-stimulation with antigen, revealing that in zfCD4+ lymphocytes an increased expression of cytokines and master transcript factors was seen when the same antigen was used for boosting. This is the first demonstration of Th-type responses effected by CD4+ lymphocytes in a poikilotherm. Lastly, I studied an aspect of Treg function in zebrafish, focused on a master transcription factor of Tregs, the FoxP3 gene. Knocking down FoxP3 genes in zebrafish resulted in modulated gene expression of cytokines and transcription factors associated with Th and Treg cells, providing some evidence that the immune tolerance function of Treg cells may exist in teleost fish, with some sub-functionalisation of the two FoxP3 paralogues apparent. This thesis extends our knowledge into teleost adaptive immune responses mediated by CD4+ Th cells and putative FoxP3+ Treg cells and may aid future studies using zebrafish as a model of vertebrate immune function.

No description available.

Metabolic power budgeting, the regulation of metabolism with respect ot metabolic scope, was studied in the laboratory in zebra fish using respirometry, and in the field on pike using heart-rate telemetry. Increased food consumption by zebra fish resulted in higher growth, mortality and metabolism. The magnitudes of the components of metabolism: maximum metabolism, standard metabolism, routine metabolism and feeding metabolism were measured. Power budgets for zebra fish fed high and low rations were constructed. Fish fed high rations worked harder than fish fed lower rations, but were apparently not working near the upper limit of the metabolic scope. Possible mechanisms for growth-related mortality are considered. Biological information on the populations of pike in Lochs Kinord and Davan (Grampian Highlands) were gathered. The population of L. Kinord was dominated by young, small fish; apparently due to exploitation. L. Davan is unexploited and had a pike population consisting of a much wider range of ages and sizes. Methods for assessing regurgitation by pike were developed. Effects of long and short-term temperature fluctuations, and feeding on heart rate of captive pike were studied. Resting heart rate increased exponentially with increasing temperature; heart rate appeared to accommodate all changes in resting metabolism. Post-prandial heart-rate records could be used to accurately estimate meal size. Gastric evacuation rates corresponded to digestion times estimated from heart-rate records. Heart-rate telemetry was used to study metabolic power budgeting, feeding and activity of wild pike from Lochs Kinord and Davan in June 1988. Pike worked mainly at low power levels relative to metabolic scope. Tachycardias associated with localized movement were frequent, and such movement was accurately recorded by heart-rate telemetry but frequently undetected by conventional means. Feeding events were identified and the metabolic costs of survival estimated. Some unusually energetically-expensive localized movements were recorded; the possible reasons for this are discussed. Intraperitoneal implantation techniques were developed for transmitter attachment on pike. Experments using dummy transmitters on pike and rainbow trout showed no effect on growth, survival or reproduction, but tissue reactions differed. Male and female pike, location-tracked with implanted transmitters before, during and after spawning time exhibited increased overall activity during the apparent spawning period, as well as changes in diet activity. Males were significantly more active than females in three out of seven weeks. Spawning appears to be a period of high energy expenditure for pike.

Thesis (M.S. in Biological Sciences)--Vanderbilt University, May 2008. / Title from title screen. Includes bibliographical references.

Thesis (Ph.D.)--University of Nebraska-Lincoln, 2007. / Title from title screen (site viewed Feb. 19, 2008). PDF text: xiii, 118 p. : ill. (some col.) ; 2 Mb. UMI publication number: AAT 3271922. Includes bibliographical references. Also available in microfilm and microfiche formats.

Thesis (M.S.)--Texas State University-San Marcos, 2008. / Vita. Appendix: leaves 34-43. Includes bibliographical references (leaves 44-48). Also available on microfilm.

Thesis (Ph. D.)--University of Iowa, 2007. / Thesis supervisors: Val C. Sheffield, Diane C. Slusarski. Includes bibliographical references (leaves 139-151).

Thesis (Ph.D.)--York University, 2007. Graduate Programme in Biology. / Typescript. Includes bibliographical references. Also available on the Internet. MODE OF ACCESS via web browser by entering the following URL: http://gateway.proquest.com/openurl?url_ver=Z39.88-2004&res_dat=xri:pqdiss&rft_val_fmt=info:ofi/fmt:kev:mtx:dissertation&rft_dat=xri:pqdiss:NR29321

Honors Project--Smith College, Northampton, Mass., 2009. / Includes bibliographical references (p. 125)