Search

In their native environment, gulf toadfish excrete equal quantities of ammonia and urea. However, upon exposure to stressful conditions in the laboratory (i.e. crowding, confinement or air exposure), toadfish decrease branchial ammonia excretion and become ureotelic. The objective of this study was to determine the influences of cortisol and ammonia on ammonia excretion relative to expression of Rhesus (Rh) glycoproteins and the ammonia-fixing enzyme, glutamine synthetase (GS). In vivo infusions and/or injections were used to manipulate corticosteroid activity and plasma ammonia concentrations in ureotelic toadfish. Metyrapone treatment to lower circulating cortisol levels resulted in a 3.5-fold elevation of ammonia excretion rates, enhanced mRNA expression of two of the toadfish Rh isoforms (Rhcg1 and Rhcg2), and decreased branchial and hepatic GS activity. Correspondingly, cortisol infusion decreased ammonia excretion 2.5-fold, a change that was accompanied by reduced branchial expression of all toadfish Rh isoforms (Rhag, Rhbg, Rhcg1 and Rhcg2) and a twofold increase in hepatic GS activity. In contrast, maintenance of high circulating ammonia levels by ammonia infusion enhanced ammonia excretion and Rh expression (Rhag, Rhbg and Rhcg2). Toadfish treated with cortisol showed an attenuated response to ammonia infusion with no change in Rh mRNA expression or GS activity. In summary, the evidence suggests that ammonia excretion in toadfish is modulated by cortisol-induced changes in both Rh glycoprotein expression and GS activity. [ABSTRACT FROM AUTHOR]

Abstract: Ureotelic Gulf toadfish (Opsanus beta) do not excrete urea continuously; instead, urea is accumulated internally until a branchial urea transport mechanism is activated to facilitate the excretion of urea in distinct pulses. This unusual pulsatile urea excretion pattern is regulated, in part, by permissive declines in circulating cortisol concentrations. The current study examined toadfish urea transporter (tUT) and glucocorticoid receptor (GR) transcript levels in toadfish gill following chronic (days) and acute (hours) changes in corticosteroid activity. Experimentally lowering circulating cortisol did not significantly alter tUT mRNA abundance but increased GR mRNA. On an acute timescale, a 6.2-fold upregulation of tUT mRNA occurred 12 to 18h following a urea pulse event with no change in GR mRNA. In silico analysis of an isolated 1.2kb fragment, upstream promoter region of the tUT gene, revealed 6 putative glucocorticoid response element (GRE) half sites. In vivo reporter assays of the tUT promoter fragment demonstrated relative luciferase activity was enhanced 3.4- and 9.8-fold following exposure to moderate (via a 48h crowding stress) and high (via infusion for 48h) cortisol. We conclude that a GRE-mediated upregulation of mRNA may be required to maintain tUT activity by offsetting post-transcriptional and/or post-translational changes that may be associated with chronically elevated plasma cortisol. [Copyright &y& Elsevier]

Gulf toadfish, Opsanus beta, are one among a group of unusual teleosts that excrete urea as their predominant nitrogen end product in response to stressful conditions. Under conditions of crowding or confinement, fasted toadfish excrete the majority of their nitrogen waste in large pulses of urea (>90% of total nitrogen) lasting up to 3h. An earlier study demonstrated that cortisol has an inhibitory influence on urea pulse size. The present study tested the hypothesis that cortisol mediates changes in urea pulse size in ureotelic toadfish through the glucocorticoid receptor (GR) and not the mineralocorticoid receptor (MA). In vivo pharmacological investigations were used to manipulate the corticosteroid system in crowded toadfish, including experimentally lowering plasma cortisol levels by the injection of metyrapone, blocking cortisol receptors through exposure to either RU-486 (GR antagonist) and spironolactone (MR antagonist), or through exogenous infusion of the tetrapod mineralocorticoid aldosterone (tetrapod MR agonist). The data demonstrate that lowering the activity of cortisol, either by inhibiting its synthesis or by blocking its receptor, resulted in a two- to threefold increase in pulse size with no accompanying change in pulse frequency. Treatment with spironolactone elicited a minor (-1.5-fold) reduction in pulse size, as did aldosterone treatment, suggesting that the anti- mineralocorticoid spironolactone has an agonistic effect in a piscine system. In summary, the evidence suggests that urea transport mechanisms in pulsing toadfish are upregulated in response to low cortisol, mediated primarily by GRs, and to a lesser extent MRs. [ABSTRACT FROM AUTHOR]