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Prolactin (PRL) in fish is considered to be an osmoregulatory hormone, although some studies suggest that it may influence the production of steroid hormones in the gonads. The objective of the present study was to establish if PRL is involved in reproduction of the gilthead seabream—a protandrous hermaphrodite. Adult and juvenile gilthead seabream received implants of estradiol-17b (E2) for 1 wk during the breeding season, and the mRNA expressions of PRL and PRL receptor (sbPRLR) were determined. Northern blot analysis revealed a single pituitary PRL transcript, the expression of which was significantly reduced by E2 treatment in adults but significantly increased in juvenile fish. In adult gonads, four sbPRLR transcripts of 1.1, 1.3, 1.9, and 2.8 kilobases were observed. A competitive reverse transcription-polymerase chain reaction was developed and used to determine how E2 treatment alters expression of the gonadal sbPRLR gene. Seabream PRLR was detectable in all samples analyzed by this assay. Levels of sbPRLR mRNA increased significantly (50-fold) after E2 treatment in adults, but a 24-fold decrease was measured in juveniles. Immunohistochemistry using specific polyclonal antibodies raised against an oligopeptide from the extracellular domain of sbPRLR detected the receptor in spermatogonia and oocytes. Taken together, the preceding results suggest that in the seabream, PRL may act on both testis and ovary via its receptor and that the stage of maturity influences this process. The full characterization and relative importance of the different transcripts of sbPRLR in eliciting the action of PRL in the gonads remain to be elucidated.

Olfactory sensitivity to changes in environmental Ca2+ has been demonstrated in two teleost species; a salmonid (Oncorhynchus nerka) and a marine/estuarine perciform (Sparus aurata). To assess whether this phenomenon is restricted to species that normally experience large fluctuations in external ion concentrations (e.g. moving from sea water to fresh water) or is present in a much wider range of species, we investigated olfactory Ca2+ sensitivity in the goldfish (Carassius auratus), which is a stenohaline, non-migratory freshwater cyprinid. Extracellular recording from the olfactory bulb in vivo by electroencephalogram (EEG) demonstrated that the olfactory system is acutely sensitive to changes in external Ca2+ within the range that this species is likely to encounter in the wild (0.05–3 mmol l–1). The olfactory system responded to increases in external calcium with increasing bulbar activity in a manner that fitted a conventional Hill plot with an apparent EC50 of 0.9±0.3 mmol l–1 (close to both ambient and plasma free [Ca2+]) and an apparent Hill coefficient of 1.1±0.3 (means ± S.E.M., N=6). Thresholds of detection were below 50 mmol l–1. Some olfactory sensitivity to changes in external [Na+] was also recorded, but with a much higher threshold of detection (3.7 mmol l–1). The olfactory system of goldfish was much less sensitive to changes in [Mg2+] and [K+]. Preliminary data suggest that Ca2+ and Mg2+ are detected by the same mechanism, although with a much higher affinity for Ca2+. Olfactory sensitivity to Na+ may warn freshwater fish that they are reaching the limit of their osmotic tolerance when in an estuarine environment. Olfaction of serine, a potent odorant in fish, was not dependent on the presence of external Ca2+ or Na+. Finally, the teleost Ca2+-sensing receptor (Ca-SR) was shown to be highly expressed in a subpopulation of olfactory receptor neurones by both immunocytochemistry and in situ hybridisation. The olfactory sensitivity to Ca2+ (and Mg2+) is therefore likely to be mediated by the Ca-SR. We suggest that olfactory Ca2+ sensitivity is a widespread phenomenon in teleosts and may have an input into the physiological mechanisms regulating internal calcium homeostasis.

The cDNA for the calcium-sensing receptor (CaSR) gene has been cloned from the marine teleost Sparus aurata, the sea bream. The isolated clones were 3.3 kb long with an open reading frame of 2820 bp, a 50 UTR of 240 bp, and 30 UTR of 248 bp. The gene codes for a mature peptide of 940 amino acids which has three principal domains; the extracellular region is more than half the total protein, there is a seven-transmembrane domain, and there is a short intracellular domain. There is considerable sequence identity, 91%, shared between the CaSR of sea bream and puffer fish but overall similarities with mammalian CaSR peptides vary between 44% for rat and mouse and 48% with human CaSR. Nevertheless, the 18 cysteine residues of the extracellular domain are present in all sequences so far analysed of which 9 form a cysteine-rich region in sea bream similar to mammalian CaSR. The distribution of CaSR in sea bream tissues detected by in situ hybridisation showed gene expression in epithelia associated with ion transport or ion regulation including the hind gut, chloride cells of the gills, operculum, gall bladder, pituitary adenohypophysis, and coronet cells of the saccus vasculosus; this distribution was confirmed by RT-PCR. By in situ hybridisation, CaSR gene expression was also present in olfactory nerves and leucocytes.

Submitted by Deborah Power (dpower@ualg.pt) on 2014-10-21T09:02:42Z No. of bitstreams: 1 Cloning, characterization, and tissue distribution of prolactin receptor in the sea bream (Sparus aurata).pdf: 1000052 bytes, checksum: 6d76e09bec31ed696b04740fafcad5fd (MD5) Approved for entry into archive by Carla Janeira (cjaneira@ualg.pt) on 2014-10-21T09:34:43Z (GMT) No. of bitstreams: 1 Cloning, characterization, and tissue distribution of prolactin receptor in the sea bream (Sparus aurata).pdf: 1000052 bytes, checksum: 6d76e09bec31ed696b04740fafcad5fd (MD5) Made available in DSpace on 2014-10-21T09:34:43Z (GMT). No. of bitstreams: 1 Cloning, characterization, and tissue distribution of prolactin receptor in the sea bream (Sparus aurata).pdf: 1000052 bytes, checksum: 6d76e09bec31ed696b04740fafcad5fd (MD5) Previous issue date: 2001

In this study we describe the isolation and characterisation of the parathyroid hormone-related protein (PTHrP) gene from the teleost Fugu rubripes. The gene has a relatively simple structure, compared with tetrapod PTHrP genes, composed of three exons and two introns, encompassing 2.25 kb of genomic DNA. The gene encodes a protein of 163 amino acids, with a putative signal peptide of 37 amino acids and a mature peptide of 126 amino acids. The overall homology with known tetrapod PTHrP proteins is low (36%), with a novel sequence inserted between positions 38 and 65, the absence of the conserved pentapeptide (TRSAW) and shortened C-terminal domain. The N-terminus shows greater conservation (62%), suggesting that it may have a hypercalcaemic function similar to that of tetrapod PTHrP. In situ localisation and RT–PCR have demonstrated the presence of PTHrP in a wide range of tissues with varying levels of expression. Sequence scanning of overlapping cosmids has identified three additional genes, TMPO, LDHB and KCNA1, which map to human chromosome 12, with the latter two mapping to 12p12-11.2. PTHrP in human also maps to this chromosome 12 sub-region, thus demonstrating conservation of synteny between human and Fugu.

A major action of prolactin (PRL) in teleost fish is the maintenance of hydromineral balance in euryhaline species in fresh water. The function of PRL in marine teleosts is less certain and unlike euryhaline teleosts, such as tilapia and salmon, there is relatively little information about protein or gene structure. Associated with studies to determine potential functions of PRL, pituitary prolactin cDNA has been cloned and sequenced from sea bream (Sparus aurata), a marine teleost. The sequence obtained spanned 1349 bp and contained an open reading frame encoding a protein of 212 amino acids composed of a putative signal peptide of 24 residues and a mature protein of 188 amino acids. N-terminal sequencing of the native protein confirmed unambiguously the cleavage site, Ala24, Val25, predicted from alignments of the sea bream PRL cDNA with that of other teleosts. The presence of only one form of PRL in sea bream was supported by identification using Northern blots of only a single transcript of 1.35 kb. Reverse transcription and polymerase chain reaction techniques coupled with Southern blot analysis resulted in the detection of PRL in the pituitary but also in the intestine, liver, ovary, and testes. This work was supported by a grant from the Portuguese Fundacao para Ciencia e Tecnologia (2/2.1/BIA/211/94). Cecilia R. A. Santos was in receipt of a JNICT grant (BD/2632/93/IG). We are grateful toMs Nair Enxerto for assistance in the study. Note added in proof. In a recent publication Bole-Feysot et al., 1998, refer to over 300 separate functions of PRL in vertebrates. (Bole- Feysot, C., Goffin, V., Edery, M., Binart, N., and Kelly, P. A. (1998). Prolactin (PRL) and its receptor: Actions, signal transduction pathways and phenotypes observed in PRL receptor knockout mice. Endocr. Rev. 19, 225–268).

Submitted by Adelino Canario (acanario@ualg.pt) on 2014-05-09T10:33:26Z No. of bitstreams: 1 The effect of stocking density on growth in the gilthead sea-bream, Sparus aurata (L.).pdf: 392980 bytes, checksum: 14a35867ac84b042ee3506a56a79e491 (MD5) Approved for entry into archive by Merja Muzavor (mmuzavor@ualg.pt) on 2014-05-15T14:33:24Z (GMT) No. of bitstreams: 2 7223591071001921.zip: 374383 bytes, checksum: 4ce2e77abb926c194af706812efeb7b1 (MD5) The effect of stocking density on growth in the gilthead sea-bream, Sparus aurata (L.).pdf: 392980 bytes, checksum: 14a35867ac84b042ee3506a56a79e491 (MD5) Made available in DSpace on 2014-05-15T14:33:24Z (GMT). No. of bitstreams: 2 7223591071001921.zip: 374383 bytes, checksum: 4ce2e77abb926c194af706812efeb7b1 (MD5) The effect of stocking density on growth in the gilthead sea-bream, Sparus aurata (L.).pdf: 392980 bytes, checksum: 14a35867ac84b042ee3506a56a79e491 (MD5) Previous issue date: 1998

Submitted by Deborah Power (dpower@ualg.pt) on 2014-10-23T08:53:11Z No. of bitstreams: 1 Parathyroid hormone-related protein and somatolactin in sea bream (Sparus aurata) brain and pituitary.pdf: 434772 bytes, checksum: 5005ffce0fd82896707cb70657125341 (MD5) Approved for entry into archive by Carla Janeira (cjaneira@ualg.pt) on 2014-10-24T09:44:37Z (GMT) No. of bitstreams: 1 Parathyroid hormone-related protein and somatolactin in sea bream (Sparus aurata) brain and pituitary.pdf: 434772 bytes, checksum: 5005ffce0fd82896707cb70657125341 (MD5) Made available in DSpace on 2014-10-24T09:44:37Z (GMT). No. of bitstreams: 1 Parathyroid hormone-related protein and somatolactin in sea bream (Sparus aurata) brain and pituitary.pdf: 434772 bytes, checksum: 5005ffce0fd82896707cb70657125341 (MD5) Previous issue date: 1997

1. Parathyroid hormone-related protein (PTHrP) is an important mediator of humoral hypercalcaemia of malignancy in humans. Normal human subjects have very low levels of PTHrP in their circulation. 2. Parathyroid hormone-related protein has recently been demonstrated in high levels in the circulation and tissues of the sea bream and the dogfish, leading to the hypothesis that PTHrP may be a 'classical' hormone in fish. 3. Immunohistochemistry and in situ hybridization were performed to investigate the evolutionary history of PTHrP. Tissues were examined from a number of lower vertebrates, including lungfish, lamprey and several species of bony and cartilaginous fish. Parathyroid hormone-related protein was localized to the skin and to kidney tubules in all animals studied. In the developing lungfish, PTHrP was observed in the notochord, developing brain and skeletal muscle layers. These results suggest that PTHrP is of ancient origin and has a basic and fundamental function in vertebrates.

Texto integral disponível através do editor The effects of an N-terminal peptide (amino acids 1-38) of Fugu parathyroid hormone-related protein (PTHrP 1-38) on calcium regulation of larval sea bream were investigated in seawater (36 per thousand) and after transfer to dilute seawater (12 per thousand). Exposure to PTHrP 1-38 evoked a 1.5-fold increase in calcium influx in both full-strength and dilute seawater. Calcium influx in dilute seawater-adapted larvae was roughly one-half that observed in full-strength seawater controls. PTHrP 1-38 also reduced drinking of fish in seawater but, at all concentrations tested, was without effect in dilute seawater. The amount of water imbibed was 55% lower in dilute seawater than in seawater. PTHrP 1-38 exposure affected the calcium influx route: the main contribution of calcium uptake shifted from intestinal absorption to extraintestinal uptake, probably by the induction of a dose-dependent increase in branchial (active) transport. Moreover, seawater-adapted fish exposed to 1 nM and 10 mM PTHrP 1-38 experienced a 2.5-fold reduction in overall calcium efflux. Overall, the calciotropic action of PTHrP 1-38 resulted in a dose-dependent increase in net calcium balance.