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The present study investigated the potential role of hypoxia-inducible factor (HIF) in calcium homeostasis in developing zebrafish (Danio rerio). It was demonstrated that zebrafish raised in hypoxic water (30 mmHg; control, 155 mmHg PO2) until 4 days post-fertilization exhibited a substantial reduction in whole-body Ca2+ levels and Ca2+ uptake. Ca2+ uptake in hypoxia-treated fish did not return to prehypoxia (control) levels within 2 h of transfer back to normoxic water. Results from real-time PCR showed that hypoxia decreased the whole-body mRNA expression levels of the epithelial Ca2+ channel (ecac), but not plasma membrane Ca2+-ATPase (pmca2) or Na+/ Ca2+-exchanger (ncx1b). Whole-mount in situ hybridization revealed that the number of ecac-expressing ionocytes was reduced in fish raised in hypoxic water. These findings suggested that hypoxic treatment suppressed the expression of ecac, thereby reducing Ca2+ influx. To further evaluate the potential mechanisms for the effects of hypoxia on Ca2+ regulation, a functional gene knockdown approach was employed to prevent the expression of HIF-1αb during hypoxic treatment. Consistent with a role for HIF-1αb in regulating Ca2+ balance during hypoxia, the results demonstrated that the reduction of Ca2+ uptake associated with hypoxic exposurewas not observed in fish experiencing HIF-1αb knockdown. Additionally, the effects of hypoxia on reducing the number of ecac-expressing ionocytes was less pronounced in HIF-1αb-deficient fish. Overall, the current study revealed that hypoxic exposure inhibited Ca2+ uptake in developing zebrafish, probably owing to HIF-1αb-mediated suppression of ecac expression. [ABSTRACT FROM AUTHOR]

The present study tested the hypothesis that zebrafish (Danio rerio) aquaporin-1a1 (AQP1a1) serves as a multi-functional channel for the transfer of the small gaseous molecules, CO2 and ammonia, as well as water, across biological membranes. Zebrafish embryos were microinjected with a translation-blocking morpholino oligonucleotide targeted to AQP1a1. Knockdown of AQP1a1 significantly reduced rates of CO2 and ammonia excretion, as well as water fluxes, in larvae at 4 days post fertilization (dpf). Because AQP1a1 is expressed both in ionocytes present on the body surface and in red blood cells, the haemolytic agent phenylhydrazine was used to distinguish between the contributions of AQP1a1 to gas transfer in these two locations. Phenylhydrazine treatment had no effect on AQP1a1-linked excretion of CO2 or ammonia, providing evidence that AQP1a1 localized to the yolk sac epithelium, rather than red blood cell AQP1a1, is the major site of CO2 and ammonia movements. The possibility that AQP1a1 and the rhesus glycoprotein Rhcg1, which also serves as a dual CO2 and ammonia channel, act in concert to facilitate CO2 and ammonia excretion was explored. Although knockdown of each protein did not affect the abundance of mRNA and protein of the other protein under control conditions, impairment of ammonia excretion by chronic exposure to high external ammonia triggered a significant increase in the abundance of AQP1a1 mRNA and protein in 4 dpf larvae experiencing Rhcg1 knockdown. Collectively, these results suggest that AQP1a1 in zebrafish larvae facilitates the movement of CO2 and ammonia, as well as water, in a physiologically relevant fashion. [ABSTRACT FROM AUTHOR]

The goal of the present study was to investigate the role of circulating cortisol and urea in the transcriptional regulation of branchial glutamine synthetase (GS), which incorporates NH3 into glutamate to form glutamine, and the toadfish urea transporter, tUT, which is involved in urea excretion across the gill of the gulf toadfish. GS (of which there are two isoforms, LGS and GGS) and tUT mRNA expression and activity were measured in toadfish exposed to treatments that would induce variable stress responses. In addition, the role of circulating urea in tUT regulation was investigated by infusing toadfish with urea alone or in combination with intraperitoneal injection of RU486, a corticosteroid type II receptor antagonist. There was a 4.8-fold upregulation in the mRNA expression of the gill-specific GS isoform (GGS) in response to cortisol infusion and a similar upregulation in the more ubiquitous isoform (LGS). Furthermore, there was a significant 1.9-fold and 3.3-fold upregulation in the mRNA expression of the toadfish urea transporter, tUT, in response to stress through crowding or exogenous cortisol loading through infusion, respectively. In addition, tUT was found to have a urea-sensitive component to transcriptional regulation that was independent of circulating cortisol concentrations. However, the changes measured in mRNA expression of GGS, LGS and tUT did not correspond with changes in protein activity. To determine the cell type(s) involved in glutamine production and urea excretion, we attempted to localize GGS, LGS and tUT using in situ hybridization. This study is the first to show that GGS and tUT expression appear to occur in gill mitochondria-rich cells of toadfish, suggesting that these cells play a combined glutamine production and urea excretion role, which may have implications for predator avoidance. [ABSTRACT FROM AUTHOR]

Experiments were performed to test the hypothesis that the marked ventilatory and cardiovascular responses to hypercarbia in rainbow trout Oncorhynchus mykiss arise from specific stimulation of chemoreceptors localised to the first gill arch. This was accomplished by measuring cardiorespiratory variables during acute hypercarbia (20 min at P[sub CO[sub 2]]=8mmHg; 1mmHg=0.133kPa) in fish subjected to selective bilateral extirpation of the first gill arch. The cardiovascular responses to hypercarbia in the intact fish included a significant bradycardia (from 75.0±1.6 to 69.0±2.0 beats min[sup -1]; means ± S.E.M.; N=16), an increase in dorsal aortic blood pressure (from 30.8±1.5 to 41.9±2.5 mmHg; N=16) and a rise in systemic vascular resistance (from 1.1±0.1 to 1.4±0.1 mmHg ml[sup -1] kg[sup -1] min[sup -1]; N=16). Removal of the first gill arch or pre-treatment with the muscarinic receptor antagonist atropine prevented the hypercarbic bradycardia without affecting the pressure or resistance responses. Correlation analysis, however, revealed shallow but significant inverse relationships between water P[sub CO[sub 2]] and cardiac frequency in both atropinised (r²=0.75) and gill-extirpated (r²=0.90) fish, suggesting a direct mild effect of CO[sub 2] on cardiac function. The ventilatory response to hypercarbia in the intact fish consisted of an increase in ventilation amplitude (from 0.62±0.06 to 1.0±0.13cm; N=16) with no change in breathing frequency. Removal of the first gill arch lowered resting breathing frequency and prevented the statistically significant elevation of breathing amplitude. Gill extirpation, however, did not totally abolish the positive correlation between water P[sub CO[sub 2]] and ventilation amplitude (r²=0.84), suggesting the presence of additional (although less important) chemoreceptive sites that are not confined to the first gill arch. Plasma catecholamine... [ABSTRACT FROM AUTHOR]

Focuses on a study which assessed the potential direct and modulating effects of acute hypoxia on catecholamine secretion in rainbow trout, Oncorhynchus mykiss using a perfused cardinal vein preparation and chronically cannulated fish. Materials and methods; Effects of hypoxic perfusate on in situ basal catecholamine secretion rates; Influence of acute moderate hypoxia on nicotine-evoked catecholamine release in vivo.

Deals with a study which cloned and characterized the B subunit of the trout H...V-ATPase to examine by northern analysis the expression and tissue distribution of the B subunit mRNA during external hypercapnia. Materials and methods; Results and discussion.

Discusses a study which assessed the relative involvement of nicotinic and muscarinic cholinergic receptors in the neuronal control of catecholamine secretion from the chromaffin tissue of rainbow trout. Background information on the mechanism of catecholamine secretion in teleosts; Materials and methods; Results; Discussion.