Cortisol regulates eel (Anguilla anguilla) aquaporin 3 (AQP3) mRNA expression levels in gill.

Previous studies in eel (Anguilla anguilla) gill have shown that the expression of the aquaporin 3 (AQP3) water and small solute channel is dramatically decreased (mRNA abundance decreased by up to 97%) when these euryhaline fish are acclimated from freshwater (FW) to seawater (SW). However, AQP3 mRNA expression levels in the intestine following SW-acclimation do not change. The SW-acclimating corticosteroid hormone, cortisol has previously been shown to regulate the expression of aquaporins (particularly AQP1) in eel osmoregulatory tissues in a tissue-specific and isoform-specific fashion. AQP1 is up-regulated in intestine and oesophagus, but down-regulated in kidney, following SW-acclimation in these fish. This study extends knowledge of the regulation of aquaporin expression by cortisol in the eel and shows that elevated levels of this hormone down-regulate AQP3 mRNA expression in the gill in a similar manner to SW-acclimation. However, the smaller magnitude of the changes in branchial AQP3 expression induced by cortisol-infusion (around a 60% decrease), in comparison to those occurring following SW-acclimation, suggest that other factors must also contribute to AQP3 down-regulation. In a similar fashion to the regulation of AQP1 by cortisol, changes in AQP3 expression following hormone infusion appear to be tissue-specific, as little effect was seen on the level of AQP3 expression in the intestine. Again the apparent lack of change in intestinal AQP3 expression following cortisol-infusion mimicked the invariant level of intestinal AQP3 mRNA abundance following SW-acclimation.

Salinity adaptation and gene profiling analysis in the European eel (Anguilla anguilla) using microarray technology.

The life cycle of the European eel (Anguilla anguilla) includes two long migratory periods, when the newly hatched leptocephali larvae drift on ocean currents from the Sargasso Sea to the shores of Western Europe and then again up to 30 years later when adult eels swim back to their place of birth for reproductive purposes. Prior to the migration from fresh water (FW) to sea water (SW) adult yellow eels undergo various anatomical and physiological adaptations (silvering) which promote sexual development and aid the transition to increased environmental salinities. The aim of this study was to identify and characterise changes in gene expression within the major osmoregulatory tissues of the eel which enable these fish to make the physiological adaptations required for transfer to SW environments. In particular, changes in the expression of the FW-adapting hormone prolactin were correlated with differential expression of known osmoregulatory important genes within the gill, intestine and kidney following the acclimation of eels to SW. Various tissues were sampled from individual fish at selected intervals over a 5-month period following FW/SW transfer and RNA was isolated. Suppressive subtractive hybridization (SSH) was used for enrichment of differentially expressed genes. Microarrays comprising 6144 cDNAs spotted in triplicate, from brain, gill, intestine and kidney libraries (1536 randomly selected clones per tissue library), were hybridized with appropriate targets and analysed. Microarray results were validated using known genes implicated in osmoregulation, such as prolactin, growth hormone, Na, K-ATPase and some unknown genes, the role of which in osmoregulation needs to be elucidated.

Maternal effects due to male attractiveness affect offspring development in the zebra finch.

Maternal effects occur when offspring phenotype is influenced by environmental factors experienced by the mother. Mothers are predicted to invest differentially in offspring in ways that will maximize offspring fitness depending on the environment she expects them to encounter. Here, we test for maternal effects in response to mate attractiveness on offspring developmental traits in the zebra finch Taeniopygia guttata. We controlled for parental genetic quality by manipulating male attractiveness using coloured leg rings and by randomly assigning mating pairs. The potential confounding effect of differential nestling care was controlled for by cross-fostering clutches and by allowing for variance due to foster father attractiveness in general linear models. We found a difference in egg mass investment between attractiveness groups and, importantly, we found that all of the offspring traits we measured varied with the attractiveness of the father. This provides strong evidence for maternal effects in response to mate attractiveness. Furthermore, due to the experiment design, we can conclude that these effects were mediated by differential investment of egg resources and not due to genetic differences or differences in nestling care.

Osmotic and volaemic effects on drinking rate in elasmobranch fish.

An increase in drinking rate of two species of marine elasmobranch fish, Scyliorhinus canicula and Triakis scyllia, acclimated to 80 % sea water was observed following the introduction of 100 % sea water to experimental tanks. The drinking response in both species was found to be maximal within 6 h, and a significant increase was sustained for up to 24 h in T. scyllia. Plasma osmolality was significantly increased within 6 h following introduction of 100 % sea water, and this increase was principally due to elevated plasma Na(+) and Cl(-) concentrations. Administration of 2 mol l(-1) mannitol, 75 % sucrose and vehicle (elasmobranch Ringer) did not induce a significant increase or decrease in the drinking rate of S. canicula. However, injection of 20 % NaCl was found to decrease drinking rate significantly in S. canicula 60 min after administration. Controlled haemorrhage of approximately 5.7 % of total blood volume in S. canicula induced a rapid 36-fold increase in drinking over basal levels. The present study demonstrates a physiological dipsogenesis in response to hypovolaemia in marine elasmobranch fish as part of their overall iso/hyperosmoregulatory strategy.

The effects of dietary sodium loading on the activity and expression of Na, K-ATPase in the rectal gland of the European dogfish (Scyliorhinus canicula).

cDNA fragments of both the alpha- and beta-subunits of the Na, K-ATPase and a cDNA fragment of the secretory form of Na-K-Cl cotransporter from the European dogfish (Scyliorhinus canicula) were amplified and cloned using degenerate primers in RT-PCR. These clones were used along with a sCFTR cDNA from the related dogfish shark, Squalus acanthias to characterise the expression of mRNAs for these ion transporters in the dogfish rectal gland subsequent to an acute feeding episode. Following a single feeding event where starved dogfish were fed squid portions (20 g squid/kg fish), there was a delayed and transient 40-fold increase in the activity of Na, K-ATPase in crude rectal gland homogenates. Increases in enzyme activity were apparent 3 h after the feeding event and peaked at 9 h before returning to control values within 24 h. These increases in activity were accompanied by small and transient decreases in plasma sodium and chloride concentrations lasting up to 3 days. Significant increases in the expression of mRNAs for alpha- and beta-subunits of the Na, K-ATPase, the Na-K-Cl cotransporter and CFTR chloride channel were detected but not until 1-2 days after the feeding event. It is concluded that the transient increase in Na, K-ATPase activity is not attributable to increases in the abundance of alpha- and beta-subunit mRNAs but must be associated with some, as yet unknown, post-transcriptional activation mechanism.

The control of drinking in elasmobranch fish with special reference to the renin-angiotensin system.

In summary, it is evident that teleost and elasmobranch fish respond to extra-cellular dehydration by increasing drinking rate mediated by an increase in circulating levels of ANG II. However, although the primary stimulus for drinking may be the same, clearly the mechanisms involved in regulating ion and water balance are entirely different. In order to maintain ion and water balance in the face of cellular and extra-cellular dehydration, the integration and hormonal control of renal and extra-renal function in elasmobranchs has developed in a very different manner to that described for teleost fish.

The dipsogenic effect of the renin-angiotensin system in elasmobranch fish.

This study investigated the control of drinking in elasmobranch fish through manipulation of the homologous renin-angiotensin system (RAS). The smooth muscle relaxant papaverine was found to increase basal drinking levels in the European lesser-spotted dogfish, Scyliorhinus canicula, almost 20-fold. However, this response was significantly reduced with the coadministration of the angiotensin-converting enzyme inhibitor captopril which had no effect when administered alone. Captopril was also found to block a 7-fold increase in drinking rate following administration of homologous angiotensin I in S. canicula. Finally, administration of homologous angiotensin II produced a dose-dependent response in drinking rate in two species of elasmobranchs, S. canicula and the Japanese dogfish, Triakis scyllia. These results demonstrate a central role of the RAS in the control of drinking in elasmobranch fish.

Angiotensin II binding sites in the heart of Scyliorhinus canicula: an autoradiographic study.

Dogfish (125)I [Asn(1), Pro(3), Ile(5)] angiotensin II ((125)I dfANG II) was used to establish the specific binding patterns of the different cardiac regions of the elasmobranch Scyliorhinus canicula by in vitro autoradiography. In the ventricular myocardium Scatchard analysis of saturation and displacement binding data revealed two classes of high- and low-affinity dfANG II binding sites (K(d) = 53 +/- 10 and 1300 +/- 900 pM). Two classes of dfANG II binding sites were also detected in the atrium (K(d) = 47 +/- 13 and 4690 +/- 930 pM) and in the outer layer of the conus arteriosus (K(d) = 16 +/- 9 and 398 +/- 83 pM). Conversely, the ventricular endocardium and the inner conal layer were characterized by a single class of dfANG II binding sites with affinity values of 48 +/- 11 and 106 +/- 3.3 pM, respectively. Competition experiments with either cold dfANG II or CV11974 or CGP42112 (specific ligands for mammalian AT(1) and AT(2) receptors, respectively) demonstrated a prevalence of CGP42112-selective dfANG II binding sites in both the inner and the outer conal layers. In the atrium, the ventricular myocardium, and the outer conal layer, dfANG II high-affinity binding sites poorly discriminated among the cold ligands. These results suggest that the dogfish heart may be a target organ of ANG II with distinct ANG II receptor subtype distributions.

Angiotensin and angiotensin receptors in cartilaginous fishes.

In mammals, a principal bioactive component of the renin-angiotensin system (RAS), angiotensin II (ANG II), is known to be vasopressor, dipsogenic, a stimulant of adrenocortical secretion and to control glomerular and renal tubular function. Historically, a RAS analogous to that found in mammals was thought to have first evolved in the bony fishes. Recent research has identified the unusually structured elasmobranch [Asp(1)-Pro(3)-Ile(5)] ANG II. Physiological studies have demonstrated that ANG II in elasmobranchs is vasopressor, and stimulates interrenal gland production of the elasmobranch corticosteroid 1alpha-hydroxycorticosterone. The specific binding of ANG II in elasmobranchs has been reported in gills, heart, interrenal gland, gut and rectal gland. The precise osmoregulatory role ANG II plays in cartilaginous fishes is not yet known; however, putative evidence is emerging for a role in the control of drinking rate, rectal gland secretion, and kidney function.

Expression of a duplicate Na,K-ATPase beta(1)-isoform in the European eel (Anguilla anguilla).

Recent studies on teleost fish have suggested that their genomes have undergone ancient polyploidization events resulting in the duplication of the genome. A duplicate copy of the Na,K-ATPase beta(1)-isoform (called beta(233)) has been identified in the European eel (Anguilla anguilla). The beta(233)-isoform shares high levels of nucleotide (74.8%) and amino acid (69.9%) homology with the eel beta(1)-subunit as well as other vertebrate beta(1)-sequences. Compared with the widely expressed beta(1)-isoform, expression of beta(233)-mRNA is mainly restricted to epithelial tissues. Seawater acclimation induced increases in beta(233)-mRNA levels in kidney, gill, and intestine of migratory "silver" but not the nonmigratory "yellow" adult eels, suggesting that the factors responsible for this upregulation are themselves developmentally regulated. Expression of a variably glycosylated 40- to 52-kDa beta(233)-protein in both gill "chloride" and intestinal epithelial cells suggests that the beta(233)-isoform of Na,K-ATPase may play an important functional role in the major osmoregulatory tissues of euryhaline fish such as the eel.

Male attractiveness and differential testosterone investment in zebra finch eggs.

Good-genes hypotheses of sexual selection predict that offspring fathered by preferred males should have increased viability resulting from superior genetic quality. Several studies of birds have reported findings consistent with this prediction, but maternal effects are an important confounding variable. Those studies that have attempted to control for maternal effects have only considered differential maternal investment after egg laying. However, female birds differentially deposit testosterone in the eggs, and this influences the development of the chick. This study shows that female birds deposit higher amounts of testosterone and 5alpha-dihydrotestosterone in their eggs when mated to more attractive males.

Renin-angiotensin system in elasmobranch fish: A review.

The renin-angiotensin system (RAS) has been identified recently in elasmobranch fish, and the structure of angiotensin II (ANG II) is unusual ([Asp(1),Pro(3),Ile(5)]-ANG II) compared to other vertebrates. Receptors for ANG II have been identified in blood vessels and in a variety of osmoregulatory tissues including the gill, kidney and rectal gland. In addition, there is considerable binding to the interrenal gland and the stimulation of 1alpha-hydroxycorticosterone production in vitro suggests a physiological role in corticosteroidogenesis. ANG II is a potent vasoconstrictor and this effect does not appear to be mediated by sympathetic activation or catecholamine release. Although the RAS may not be involved in maintaining basal blood pressure, it may be important in situations in which blood pressure is reduced. Understanding of the role of ANG II as an osmoregulatory hormone is only just emerging with putative roles in the control of gill, rectal gland and perhaps, drinking. In addition, the stimulation of corticosteroid secretion may provide another means of controlling osmoregulation. J. Exp. Zool. 284:526-534, 1999.

Direct vasoconstrictor action of homologous angiotensin II on isolated arterial ring preparations in an elasmobranch fish.

Arterial rings were prepared from the branchial artery, coeliac artery and ventral aorta of the Japanese dogfish Triakis scyllia and used to determine arterial contraction in a myograph. Noradrenaline caused a dose-dependent contraction (10(-9)-3 x 10(-6) M) that was completely inhibited by pre-treatment with 10(-7) M phentolamine. Homologous dogfish angiotensin II (ANG II) ([Asn1, Pro3, Ile5]-ANG II) also caused dose-dependent contraction (10(-9)-3 x 10(-6) M), but phentolamine had no effect on this response. Administration of dogfish angiotensin I (ANG-I) ([Asn1, Pro3, Ile5, Gln9]-ANG I) resulted in a contraction similar to that produced by ANG II and the effect could be blocked with 10(-7) M captopril. The mammalian ANG II receptor antagonists [Sar1, Ile8]-ANG II and [Sar1, Ala8]-ANG II caused dose-dependent contractions of coeliac artery rings, but were less potent than homologous ANG I and ANG II. These results show that the contractile effect of [Asn1, Pro3, Ile5]-ANG II is not mediated by the alpha-adrenergic system and contractions of arterial rings by noradrenaline and elasmobranch ANG II are mediated by separate vascular receptors. The elasmobranch ANG II vascular receptor may have co-evolved with the unusual structure of this peptide.

A radioimmunoassay for the determination of angiotensin II in elasmobranch fish.

A homologous radioimmunoassay was developed to determine the concentration of angiotensin II (Asn1, Pro3, Ile5)-Ang II) in elasmobranchs. Cross-reactivity with elasmobranch angiotensin I and other heterologous angiotensins was high and therefore all potentially cross-reacting angiotensins were separated by high performance liquid chromatography after prior extraction with Sep-Pak C18 cartridges. The validity of the assay for the determination of elasmobranch Ang II was demonstrated by parallelism with a series of Ang II standards with serially diluted elasmobranch plasma extracts. Overall recovery of elasmobranch Ang II added to a plasma pool was 75.1 +/- 5.2%. Plasma Ang II concentrations measured by our RIA were similar in fish adapted to 70, 100, or 120% SW at 139 +/- 20.1, 109 +/- 15.3, and 119 +/- 16.3 fmol . ml-1, respectively.

Cardiovascular actions of dogfish urotensin I in the dogfish, Scyliorhinus canicula.

A synthetic replicate of dogfish urotensin 1 (U-I), a 41-amino-acid residue peptide isolated from an extract of the caudal spinal cord region of the European spotted dogfish Scyliorhinus canicula was prepared in order to study its cardiovascular actions in the species of origin. Bolus intraarterial injections of dogfish U-I (0.3-30 nmol/kg body wt) into the celiac artery of unanesthetized dogfish produced a transient fall in arterial blood pressure (P < 0.05 in the dose range 1-3 nmol/kg) followed by a sustained and dose-dependent rise in pressure (P < 0.05 in the dose range 1-30 nmol/kg). The maximum depressor response (to 3 nmol/kg) was 0.25 +/- 0.08 kPa and the maximum pressor response (to 30 nmol/kg) was 1.08 +/- 0.09 kPa. There was no significant effect on heart rate at any dose tested. Pretreatment of the animals with the alpha-adrenergic receptor antagonist phentolamine significantly (P < 0.05) attenuated the pressor response to injections of dogfish U-I (1 nmol/kg and 10 mol/kg), demonstrating that the effects of the peptide are mediated, at least in part, through release of catecholamines. The data suggest that U-I, released together with potent pressor peptide urotensin II from the caudal neurosecretory system, may play a physiological role in cardiovascular regulation in elasmobranchs.

The presence of angiotensin II receptors in elasmobranchs.

The presence of specific Ang II receptors in membrane fractions was investigated using 125I-labeled homologous Ang II ([Asn1, Pro3, Ile5]Ang II; df Ang II) in Triakis scyllia. Specific binding sites occurred in a variety of tissues, with highest binding in interrenal tissue (17.11 +/- 2.45 fmol Ang II/mg protein) and gill (6.26 +/- 0. 69 fmol Ang II/mg protein) and possible Ang II receptors in rectal gland and other tissues. 125I-[Asn1, Pro3, Ile5]Ang II (10(-10)M) binding to branchial cell membrane fraction (25 microg protein) in 5 mM MgCl2, 125 mM NaCl, 50 mM Tris-HCl, 0.2% bovine serum albumin at 28 degrees (1) is rapid and saturable; (2) increases as a function of membrane concentration and time; and (3) optimally fits to a two-site (high-and low-affinity) model. The equilibrium dissociation constant (0.11 +/- 0.01 nM) and binding site concentration (35.00 +/- 1.16 fmol/mg protein) are similar to those of mammalian and avian vascular Ang II receptors. Bound labeled ligand was not competitively displaced by dogfish Ang I, dogfish C-type natriuretic peptide, bradykinin, or the AT1 receptor antagonist, CV 11974. The AT2 receptor antagonist, CGP 42112, was much less potent at displacing the labeled ligand compared to the unlabeled ligand.

The role of the renin-angiotensin system in the control of blood pressure and drinking in the European eel, Anguilla anguilla.

The role of the renin-angiotensin system (RAS) in the control of blood pressure and drinking was investigated in fresh water (FW)- and seawater (SW)-adapted eels, Anguilla anguilla, by comparing the effects of pharmacological manipulation through the use of papaverine (stimulator) and captopril (inhibitor) on the endogenous system. In SW eels basal blood pressure levels were lower (23.3 +/- 0.8 mm Hg) with correspondingly higher basal drinking rates (0.51 +/- 0.07 ml/kg/hr) and plasma AII concentrations (32.89 +/- 4.19 fmol/ml) compared to FW eels (33.8 +/- 1.3 mm Hg, 0.06 +/- 0.02 ml/kg/hr, 9.72 +/- 0.60 fmol/ml, respectively). In FW eels papaverine caused immediate hypotension with full recovery, decrease in plasma osmolality, and increase in drinking rate and plasma AII concentration, but in SW eels, hypotension with full recovery and an increase in plasma osmolality, drinking rate, and plasma AII concentration occurred. In FW eels captopril had no effect on the parameters measured, but in SW eels it caused a sustained decrease in blood pressure and a decline in the basal drinking rate and plasma AII concentration. Papaverine was also administered 15 min after captopril. In FW eels this manipulation caused hypotension only after the papaverine injection, followed by a partial recovery. Osmolality was unaffected, the previously observed papaverine-induced dipsogenic response was blocked, and the rise in plasma AII concentrations was smaller than with papaverine only. In SW eels there was an immediate hypotension after captopril administration with full recovery.(ABSTRACT TRUNCATED AT 250 WORDS)