In the second part of our study, cDNA of AQP-1 and pro-vasotocin were cloned from the silver sea bream. An AQP-1 full clone was isolated from kidney and intestine and it consists of 904 bp with an open reading frame of 774 bp. The deduced amino sequence of sea bream AQP-1 shares highest identity with AQP-1a of gilthead sea bream (97.7%) and AQP-1a of other fish species (83.6% to 95.8%), however, considerably low identity was found between the silver sea bream AQP-1 and AQP-1b of gilthead sea bream (56%). The silver sea bream AQP-1 possesses basic features of a functional aquaporin and AQP-1, which includes two channel-forming asparagine-proline-alanine (NPA) signature motifs, six transmembrane domains, residues of the pore-forming region and a potential mercurial inhibiting site (Cys-178). The water channel was ubiquitously expressed in gills, liver, intestine, rectum, kidney, heart, urinary bladder and blood cells. A partial fragment of pro-vasotocin was isolated from hypothalamus of silver sea bream and consists of 184 bp, including encoding regions for the processing and amidation signal, vasotocin hormone and part of the neurophysin. / Lastly, single doses of cortisol (50 microg/g tissue) or vasotocin (1 microg/g tissue) were administered to seawater-acclimated sea bream with further three-day stabilizing period in seawater followed by an abrupt 6‰ exposure or administered to seawater transfer controls for three days. Cortisol markedly stimulated intestinal expression of AQP-1 in both the seawater transfer control and abrupt 6‰ transfer groups. Vasotocin treatment did not significantly modify AQP-1 expression in all tested organs. Hypothalamic pro-vasotocin expression levels were similar among different treatment groups. / Semi-quantitative RT-PCR analysis was used for studying the effect of salinity and hormones on expression of AQP-1 and pro-vasotocin. In the long-term salinity acclimation experiment, the sea bream were acclimated to six different salinity regimes (0‰, 6‰, 12‰, 33‰, 50‰, 70‰) for four weeks. The abundance of AQP-1 transcript was the highest in intestine of 70‰-acclimated fish among different salinity groups and there was also a statistically significant increase in 12‰-acclimated fish. Branchial AQP-1 expression was significantly upregulated in sea bream acclimated to freshwater. In contrast, the hypothalamic pro-vasotocin expression was significantly downregulated during freshwater acclimation. In addition, the sea bream were also subjected to an abrupt 6%o transfer at different time intervals (2, 6, 12, 72 and 168 hours). RT-PCR analysis revealed there was a transient decrease in branchial AQP-1 expression two hours after abrupt hypo-osmotic exposure and the expression levels subsequently returned to the seawater control levels. The expression levels of hypothalamic pro-vasotocin were not significantly altered by the abrupt exposure treatment. / The present experiments investigated the effects of salinity and hormones on the relative expression of hypothalamic pro-vasotocin, and aquaporin-1 (AQP-1) in intestine, gills and kidney of the silver sea bream Sparus sarba. With the use of immunohistochemical techniques, immunoreactivity of AQP-1 was detected at the basal side of enterocytes and gill chloride cells, and at the apical brush border of kidney tubules whereas AQP-3 was only localized in similar positions in the gills and intestines. AQP-1 was relatively more ubiquitous than AQP-3 and was localized with same cell types as the electrogenic Na+-K+-ATPase in gills and kidney. / The present study had demonstrated the responsiveness of intestinal and branchial AQP-1 expressions of the silver sea bream to environmental salinity perturbations. Further to this, cortisol was observed to upregulate the transcription of AQP-1 in the intestine. Pro-vasotocin expression was altered by long-term salinity adaptation, however, the linkage of this alteration to AQP-1 functioning in different osmoregulatory organs is yet to be elucidated. / Luk, Chun Yin. / Adviser: Norman Y. S. Woo. / Source: Dissertation Abstracts International, Volume: 72-04, Section: B, page: . / Thesis (Ph.D.)--Chinese University of Hong Kong, 2010. / Includes bibliographical references (leaves 200-222). / 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 Company, [200-] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Abstract also in Chinese.
| Identifer | oai:union.ndltd.org:cuhk.edu.hk/oai:cuhk-dr:cuhk_344536 |
| Date | January 2010 |
| Contributors | Luk, Chun Yin., Chinese University of Hong Kong Graduate School. Division of Life Sciences. |
| Source Sets | The Chinese University of Hong Kong |
| Language | English, Chinese |
| Detected Language | English |
| Type | Text, theses |
| Format | electronic resource, microform, microfiche, 1 online resource (xxv, 222 leaves : ill.) |
| Rights | Use of this resource is governed by the terms and conditions of the Creative Commons “Attribution-NonCommercial-NoDerivatives 4.0 International” License (http://creativecommons.org/licenses/by-nc-nd/4.0/) |
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