Pulsatile Urea Excretion Research Articles

Evidence that Gulf toadfish use pulsatile urea excretion to communicate social status

Gulf toadfish (Opsanus beta), a highly territorial marine teleost species, are believed to communicate through chemicals released across the gill during pulsatile urea excretion. While freshwater teleost and crustacean urinary signals have been shown to relay information about dominance to reduce physical aggression in future encounters, the use of chemical signals to convey social status in marine teleosts is understudied. Behavior and urea excretion patterns were monitored in pairs of male toadfish during an initial agonistic encounter and in a 2nd encounter where a subset of pairs had their nares blocked to determine how olfaction, and thus chemical communication, play a role in establishing dominance. Anosmic toadfish did not experience increases in aggressive behavior, unlike other species previously studied. However, behavior and the pattern of urea excretion were disrupted in anosmic pairs compared to control pairs. Specifically, control subordinate fish had an increase in their dominance index during the 2nd encounter, a response that anosmic subordinate fish did not experience suggesting that without the ability to smell, subordinate fish cannot recognize their opponent and assess their fighting ability and have a reduced chance of winning. These anosmic subordinate fish also had an increase in pulse frequency, perhaps reflecting an increased effort in communication of status. Future research is needed to conclude if peaks in agonistic behavior are coordinated around the time of urea pules. However, the observed changes in behavior and pulsatile urea excretion due to anosmia in the present study provide evidence that toadfish use pulsatile urea excretion to release signals for chemical communication during agonistic encounters.

Do reproductive hormones control Gulf toadfish pulsatile urea excretion?

Gulf toadfish (Opsanus beta) can excrete the majority of their nitrogenous waste as urea in distinct pulses across their gill. Urea pulses are controlled by cortisol and serotonin (5-HT) and are believed to contain chemical signals that may communicate reproductive and/or social status. The objectives of this study were to determine if reproductive hormones are involved in controlling pulsatile urea excretion, and if toadfish respond to prostaglandins as a chemical signal. Specifically, 11-ketotestosterone (11-KT), estradiol (E2), and the teleost pheromone prostaglandin E2 (PGE2) were investigated. Castration during breeding season did not affect pulsatile urea excretion but serial injections of 11-KT outside of breeding season did result in a 48% reduction in urea pulse size in fish of both sexes. Injections of E2 and PGE2, on the other hand, did not alter urea excretion patterns. Toadfish also did not pulse urea in response to waterborne exposure of PGE2 suggesting that this compound does not serve as a toadfish pheromone alone. Toadfish have significantly higher plasma 5-HT during breeding season compared to the months following breeding season. Future research should focus on the composition of the chemical signal in toadfish and the potential importance of seasonal changes in plasma 5-HT in toadfish pulsatile urea excretion and teleost reproduction in general.

Pulsatile urea excretion in Gulf toadfish: the role of circulating serotonin and additional 5-HT receptor subtypes.

The neurochemical serotonin (5-HT) is involved in stimulating pulsatile urea excretion in Gulf toadfish (Opsanus beta) through the 5-HT2A receptor; however, it is not known if (1) the 5-HT signal originates from circulation or if (2) additional 5-HT receptor subtypes are involved. The first objective was to test whether 5-HT may be acting as a hormone in the control of pulsatile urea excretion by measuring potential fluctuations in circulating 5-HT corresponding with a urea pulse, which would suggest circulating 5-HT may be involved with urea pulse activation. We found that plasma 5-HT significantly decreased by 38% 1h after pulse detection when branchial urea excretion was significantly elevated and then returned to baseline. This suggests that 5-HT is removed from the circulation, possibly through clearance or excretion, and may be involved in the termination of pulsatile urea excretion. There appeared to be no pulsatile release of 5-HT from peripheral tissues to trigger a urea pulse. The second objective was to determine if additional 5-HT receptor subtypes, such as an additional 5-HT2 receptor (5-HT2C receptor) or the 5-HT receptors that are linked to cAMP (5-HT4/6/7 receptors), played a role in the stimulation of urea excretion. Intravenous injection of 5-HT2C, 5-HT4, 5-HT6, and 5-HT7 receptor agonists did not result in a urea pulse, suggesting that these receptors, and thus cAMP, are not involved in stimulating urea excretion. The involvement of circulating 5-HT and the 5-HT2A receptor in the regulation of pulsatile urea excretion may provide insight into its adaptive significance.

Do Gulf Toadfish Use Pulsatile Urea Excretion to Chemically Communicate Reproductive Status?

Gulf toadfish (Opsanus beta) are exceptionally capable of switching from excreting ammonia as their primary nitrogenous waste to excreting predominantly urea in distinct pulses across the gill. Previous studies suggest that these urea pulses may be used for intraspecific chemical communication. To determine whether pulsatile urea excretion communicates reproductive status, toadfish were sexed using ultrasound and delivered conspecific-conditioned seawater (CC-SW) that previously housed a conspecific of the opposite sex, a conspecific chemical alarm cue (avoidance control), or a prey cue (attraction control). Swim behavior, attraction to or avoidance of the cues, and changes in the pattern of pulsatile urea excretion were monitored during and after delivery. Gulf toadfish did not spend more time in zones that were delivered CC-SW or prey cue. However, male toadfish spent significantly more time swimming after the delivery of female cues than control seawater (SW). In contrast, toadfish did not appear to have an immediate avoidance response to the conspecific alarm cue. Additionally, significantly more toadfish pulsed within 7 h of CC-SW and prey cue delivery compared to control SW, and pulse frequency was 1.6 times greater in response to CC-SW than control SW. These results, in combination with increased urea production and excretion the during breeding season, suggest that toadfish may use pulsatile urea excretion to communicate with conspecifics when exposed to chemosensory cues from the opposite sex.

Extrinsic nerves are not involved in branchial 5-HT dynamics or pulsatile urea excretion in Gulf toadfish, Opsanus beta

Gulf toadfish (Opsanus beta) can switch from continuously excreting ammonia as their primary nitrogenous waste to excreting predominantly urea in distinct pulses. Previous studies have shown that the neurotransmitter serotonin (5-HT) is involved in controlling this process, but it is unknown if 5-HT availability is under central nervous control or if the 5-HT signal originates from a peripheral source. Following up on a previous study, cranial nerves IX (glossopharyngeal) and X (vagus) were sectioned to further characterize their role in controlling pulsatile urea excretion and 5-HT release within the gill. In contrast to an earlier study, nerve sectioning did not result in a change in urea pulse frequency. Total urea excretion, average pulse size, total nitrogen excretion, and percent ureotely were reduced the first day post-surgery in nerve-sectioned fish but recovered by 72h post-surgery. Nerve sectioning also had no effect on toadfish urea transporter (tUT), 5-HT transporter (SERT), or 5-HT2A receptor mRNA expression or 5-HT and 5-hydroxyindoleacetic acid (5-HIAA) abundance in the gill, all of which were found consistently across the three gill arches except 5-HIAA, which was undetectable in the first gill arch. Our findings indicate that the central nervous system does not directly control pulsatile urea excretion or local changes in gill 5-HT and 5-HIAA abundance.

A waterborne chemical cue from Gulf toadfish, Opsanus beta, prompts pulsatile urea excretion in conspecifics

The Gulf toadfish (Opsanus beta) has a fully functional ornithine urea cycle (O-UC) that allows it to excrete nitrogenous waste in the form of urea. Interestingly, urea is excreted in a pulse across the gill that lasts 1–3h and occurs once or twice a day. Both the stress hormone, cortisol, and the neurotransmitter, serotonin (5-HT) are involved in the control of pulsatile urea excretion. This and other evidence suggests that urea pulsing may be linked to toadfish social behavior. The hypothesis of the present study was that toadfish urea pulses can be triggered by waterborne chemical cues from conspecifics. Our findings indicate that exposure to seawater that held a donor conspecific for up to 48h (pre-conditioned seawater; PC-SW) induced a urea pulse within 7h in naïve conspecifics compared to a pulse latency of 20h when exposed to seawater alone. Factors such as PC-SW intensity and donor body mass influenced the pulse latency response of naïve conspecifics. Fractionation and heat treatment of PC-SW to narrow possible signal candidates revealed that the active chemical was both water-soluble and heat-stable. Fish exposed to urea, cortisol or 5-HT in seawater did not have a pulse latency that was significantly different than seawater alone; however, ammonia, perhaps in the form of NH4Cl, was found to be a factor in the pulse latency response of toadfish to PC-SW and could be one component of a multi-component cue used for chemical communication in toadfish. Further studies are needed to fully identify the chemical cue as well as determine its adaptive significance in this marine teleost fish.

The Effect of Stress on Gill Basolateral Membrane Binding Kinetics of 5‐HT2 Receptor Ligands: Potential Implications for Urea Excretion Mechanisms

The goal of this study was to determine the relationship between cortisol and the toadfish serotonin 2A (5-HT2A ) receptor, which is believed to be responsible for the activation of the toadfish urea transporter, tUT. We hypothesize that elevations in cortisol would play a role in the regulation of the 5-HT2A receptor at the level of mRNA expression, ligand binding, and/or function. To test this idea, cortisol levels were manipulated by either crowding or through treatment with the cortisol synthesis blocker, metyrapone. Crowded fish had significantly higher circulating cortisol levels compared to uncrowded fish and cortisol levels in metyrapone-treated fish were significantly lower than saline-treated controls. No significant difference was measured in gill 5-HT2A mRNA expression levels between uncrowded and crowded, control- or metyrapone-treated fish. Furthermore, no significant difference was measured in [(3) H]-5-HT binding kinetics or in the competitive binding of the 5-HT2 agonist, α-methyl 5-HT, to isolated gill basolateral membranes of uncrowded or crowded toadfish. However, the binding maximum (Bmax ) of the 5-HT2A receptor antagonist, [(3) H]-ketanserin, was significantly different between all four groups of fish (metyrapone > control > crowded > uncrowded). Furthermore, metyrapone-treated fish excreted approximately twofold more urea compared to controls when injected with α-methyl 5-HT, a 5-HT2 receptor agonist shown to stimulate urea excretion. Our results suggest that cortisol may have differential effects on 5-HT receptor binding, which could have potential implications on the control of pulsatile urea excretion in toadfish.

The toadfish serotonin 2A (5-HT2A) receptor: molecular characterization and its potential role in urea excretion

Based on early pharmacological work, the serotonin 2A (5-HT2A) receptor subtype is believed to be involved in the regulation of toadfish pulsatile urea excretion. The goal of the following study was to characterize the toadfish 5-HT2A receptor at a molecular level, to determine the tissues in which this receptor is predominantly expressed and to further investigate the pharmacological specificity of toadfish pulsatile urea excretion by examining the effect of ketanserin, a 5-HT2A receptor antagonist, on resting rates of pulsatile urea excretion. The full-length toadfish 5-HT2A receptor encodes a 496 amino acid sequence and shares 57–80% sequence identity to 5-HT2A receptors of other organisms, with 100% conservation among important ligand-binding residues. Toadfish 5-HT2A receptor mRNA expression was highest in the swim bladder and gonad, followed by the whole brain. All other tissues tested (esophagus, stomach, anterior intestine, posterior intestine, rectum, liver, kidney, heart, muscle and gill) had mRNA expression levels that were significantly less than whole brain. Toadfish 5-HT2A receptor mRNA expression within the brain was highest in the hindbrain, telencephalon and midbrain/diencephalon regions. Treatment with the 5-HT2A receptor antagonist, ketanserin, resulted in a significant decrease in the pulsatile component of spontaneous urea excretion due to a reduction in urea pulse size with no significant change in pulse frequency. These results lend further support for the 5-HT2A receptor in the regulation of pulsatile urea excretion in toadfish.

5-Hydroxytryptamine initiates pulsatile urea excretion from perfused gills of the gulf toadfish (Opsanus beta)

When stressed, toadfish become ureotelic and excrete almost all of their nitrogenous waste in 1–3 daily pulses of urea-N across the gills. Intravascular injections of 5-hydroxytyptamine (5-HT; serotonin) and analogues also elicit marked excretory pulses of urea-N from toadfish in vivo, suggesting that 5-HT release is the proximate trigger for spontaneous pulses. However it is unclear whether 5-HT is acting on the gills directly or elsewhere to cause the effect indirectly. A perfused whole gill preparation which maintained normal pressure relationships and stable vascular resistance was employed to address this question. Bolus injections into the ventral aortic perfusate of either 5-HT (1, 10μmolkg−1) or the specific 5-HT2 receptor agonist α-methyl 5-HT (1, 10μmolkg−1) elicited rapid urea-N pulses from perfused toadfish gills. The effective doses, the post-injection delays (5.5±1.3min, range=2–22), the percent occurrences (57–85%), and the magnitude of the induced urea-N pulses (615.4±131.3μmol-Nkg−1, range 66.0–2634.0), were all similar to those previously reported when these agents were injected in vivo. Bolus injections of 5-HT and α-methyl 5-HT also elicited a biphasic response in ventral aortic pressure, reflecting an initial rapid short-lived vasodilation and a subsequent longer-lasting vasoconstriction. These events were similar to those which have been recorded to occur at a greater frequency during spontaneous urea-N pulsing in vivo. Neither the urea-N pulsing nor the cardiovascular responses to 5-HT were inhibited by the 5-HT2A receptor subtype blocker, ketanserin (pre-injection with 10μmolkg−1 plus 33μmolL−1 in the perfusate). Overall, these results provide strong support for the idea that the proximate stimulus for natural urea pulsing in vivo is 5-HT mobilization, acting directly in the gills.

Evidence for transcriptional regulation of the urea transporter in the gill of the Gulf toadfish, Opsanus beta

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 18 h following a urea pulse event with no change in GR mRNA. In silico analysis of an isolated 1.2 kb 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 48 h crowding stress) and high (via infusion for 48 h) 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.

Cardiovascular and respiratory reflexes of the gulf toadfish (Opsanus beta) during acute hypoxia

The acute cardiovascular and respiratory responses of the gulf toadfish, Opsanus beta, to acute hypoxia or exposure to the O(2) chemoreceptor stimulant, sodium cyanide (NaCN) were characterized and the role of serotonin type 2 (5-HT(2)) receptors in mediating these responses was investigated. Toadfish responded to hypoxia or NaCN exposure with a decrease in heart rate (fH) and an increase in breathing amplitude (V(AMP)) but no change in breathing frequency (fR). The bradycardia appeared to be mediated to some extent by 5-HT(2) receptors, as methysergide, a non-selective 5-HT(1/2) receptor antagonist, and ketanserin, a 5-HT(2) receptor antagonist, attenuated the response. Injection of alpha-methyl-5-HT, a 5-HT(2) agonist, also resulted in bradycardia that was inhibited by ketanserin, lending further support for 5-HT(2) receptor involvement, possibly 5-HT(2A) or 5-HT(2C), in the regulation of fH. External NaCN exposure resulted in a significant decrease in caudal arterial blood pressure (P(CA)) that was attenuated by methysergide. In contrast, injection with alpha-methyl-5-HT resulted in a substantial increase in P(CA) that was not affected by ketanserin, suggesting the possible involvement of 5-HT(2B) or 5-HT(2C) receptors. These data are the first to suggest a unique distribution of 5-HT(2B/2C) receptors may be involved in mediating vasoconstriction of the systemic vasculature of toadfish. These data also provide mechanistic support for why pulsatile urea excretion, believed to be regulated by 5-HT via the toadfish 5-HT(2A) receptor, is not triggered by hypoxia or external chemoreceptor activation.

Is the 5-HT2A receptor involved in mediating pulsatile urea excretion in gulf toadfish, Opsanus beta?

Is the 5-HT2A receptor involved in mediating pulsatile urea excretion in gulf toadfish, Opsanus beta?

The regulatory role of glucocorticoid and mineralocorticoid receptors in pulsatile urea excretion of the gulf toadfish,Opsanus beta

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 3 h. 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 (MR). 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 ( approximately 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.

Cortisol-sensitive urea transport across the gill basolateral membrane of the gulf toadfish (Opsanus beta)

Gulf toadfish (Opsanus beta) use a unique pulsatile urea excretion mechanism that allows urea to be voided in large pulses via the periodic insertion or activation of a branchial urea transporter. The precise cellular and subcellular location of the facilitated diffusion mechanism(s) remains unclear. An in vitro basolateral membrane vesicle (BLMV) preparation was used to test the hypothesis that urea movement across the gill basolateral membrane occurs through a cortisol-sensitive carrier-mediated mechanism. Toadfish BLMVs demonstrated two components of urea uptake: a linear element at high external urea concentrations, and a phloretin-sensitive saturable constituent (K(m) = 0.24 mmol/l; V(max) = 6.95 micromol x mg protein(-1) x h(-1)) at low urea concentrations (<1 mmol/l). BLMV urea transport in toadfish was unaffected by in vitro treatment with ouabain, N-ethylmaleimide, or the absence of sodium, conditions that are known to inhibit sodium-coupled and proton-coupled urea transport in vertebrates. Transport kinetics were temperature sensitive with a Q(10) > 2, further suggestive of carrier-mediated processes. Our data provide evidence that a basolateral urea facilitated transporter accelerates the movement of urea between the plasma and gills to enable the pulsatile excretion of urea. Furthermore, in vivo infusion of cortisol caused a significant 4.3-fold reduction in BLMV urea transport capacity in lab-crowded fish, suggesting that cortisol inhibits the recruitment of urea transporters to the basolateral membrane, which may ultimately affect the size of the urea pulse event in gulf toadfish.

11.2. The impact of the selective serotonin reuptake inhibitor, fluoxetine (Prozac), on pulsatile urea excretion in gulf toadfish, Opsanus beta

11.2. The impact of the selective serotonin reuptake inhibitor, fluoxetine (Prozac), on pulsatile urea excretion in gulf toadfish, Opsanus beta

Is urea pulsing in toadfish related to environmental O2 or CO2 levels?

The neurochemical, serotonin (5-hydroxytryptamine; 5-HT) is involved in the regulation of toadfish pulsatile urea excretion as well as the teleost hypoxia response. Thus, the goal of this study was to determine whether environmental conditions that activate branchial chemoreceptors also trigger pulsatile urea excretion in toadfish, since environmental dissolved oxygen levels in a typical toadfish habitat show significant diel fluctuations, often reaching hypoxic conditions at dawn. Toadfish were fitted with arterial, venous and/or buccal catheters and were exposed to various environmental conditions, and/or injected with the O(2) chemoreceptor agonist NaCN or the 5-HT(2) receptor agonist alpha-methyl-5HT. Arterial PO(2), as well as ammonia and urea excretion were monitored. Natural fluctuations in arterial PO(2) levels in toadfish did not correlate with the occurrence of a urea pulse. Chronic exposure (24 h) of toadfish to hyperoxia was without effect on nitrogen excretion, however, exposure to hypoxia caused a significant reduction in the frequency of urea pulses, and exposure to hypercapnia resulted in a reduction in the percentage of nitrogen waste excreted as urea. Of toadfish exposed acutely to hypoxia, 20% pulsed within 1 h, whereas none pulsed after normoxic or hypercapnic treatments. Furthermore, 20% of fish injected intravenously with NaCN pulsed within 1 h of injection, but no fish pulsed after injection of NaCN into the buccal cavity. To test whether environmental conditions affected 5-HT(2) receptors, toadfish were injected with alpha-methyl-5HT, which elicits urea pulses in toadfish. No significant differences in pulse size occurred among the various environmental treatments. Our findings suggest that neither the environmental conditions of hypoxia, hyperoxia or hypercapnia, nor direct branchial chemoreceptor activation by NaCN play a major role in the regulation of pulsatile urea excretion in toadfish.

Does Pulsatile Urea Excretion Serve as a Social Signal in the Gulf ToadfishOpsanus beta?

This study evaluated the hypothesis that the pulsatile excretion of urea by toadfish could serve as a social signal. In the first experiment, physiological parameters were measured in pairs of dominant and subordinate toadfish. Subordinate toadfish had elevated concentrations of circulating plasma cortisol, an effect maintained even after cannulation. In the second experiment, one fish of a pair was injected with 14C-urea, and the occurrence of urea pulses during social encounters was documented. Social status did not influence the order of pulsing, that is, whether a dominant or subordinate fish pulsed first during a social encounter. However, in seven out of eight pairs, both toadfish pulsed within 2 h of each other, indicating some form of communication between fish. In the third and final experiment, the response of toadfish to urea (natural or synthetic) was observed. There was a tendency for toadfish to avoid synthetic urea but there was no apparent behavioural response to water containing toadfish urea. Pulsing events do not appear to play an integral role during social encounters as previously hypothesised, but the close timing of pulses in toadfish pairs suggests some transfer of information.

Glucocorticoid receptors are involved in the regulation of pulsatile urea excretion in toadfish

The objectives of this study were to characterize the pattern of pulsatile urea excretion in the gulf toadfish in the wake of exogenous cortisol loading and to determine the receptors involved in the regulation of this mechanism. Toadfish were fitted with indwelling arterial catheters and were infused with isosmotic NaCl for 48 h after which fish were treated with cortisol alone, cortisol + peanut oil, cortisol + RU486 (a glucocorticoid receptor antagonist) or cortisol + spironolactone (a mineralocorticoid receptor antagonist). Upon cortisol loading, fish treated with cortisol alone, cortisol + oil or cortisol + spironolactone experienced a two- to threefold reduction in pulsatile urea excretion. This reduction was due to a decrease in urea pulse size with no effect on pulse frequency compared to values measured during the control NaCl infusion period. In addition, these fish showed an increase in plasma urea concentrations upon treatment. These apparent effects of cortisol treatment were abolished in fish treated with cortisol + RU486. In contrast, these fish showed an increase in pulsatile urea excretion mediated by a twofold increase in pulse size with no change in frequency. Likewise, fish treated with cortisol + RU486 showed a significant decrease in plasma urea concentrations over the course of the experiment. The findings of this study indicate that high levels of cortisol reduce pulsatile urea excretion by decreasing pulse size. In addition, it appears that glucocorticoid receptors and not mineralocorticoid receptors are involved in the regulation of the toadfish pulsatile urea excretion mechanism.

SPECIAL COLLECTION: DOGMAS AND CONTROVERSIES IN THE HANDLING OF NITROGENOUS WASTES

Science is full of dogmas. But during the process of discovery, some dogmas face challenges, and the dogmas in nitrogen waste management are no different. As all biology students know, the dogma holds that waste proteins are broken down and excreted from the body in a variety of forms, such as

Do circulating plasma AVT and/or cortisol levels control pulsatile urea excretion in the gulf toadfish ( Opsanus beta)?

Previous work has shown that pulsatile urea excretion at the gills of the gulf toadfish is due to periodic activation of a facilitated diffusion transport system with molecular and pharmacological similarity to the UT-A transport system of the mammalian kidney. In mammals, AVP and glucocorticoids are two important endocrine regulators of this system. The present study focused on the potential role of circulating AVT (the teleost homologue of AVP) and cortisol levels as possible triggers for urea pulses. Long-term (34–84 h) monitoring of plasma levels by repetitive sampling at 2-h intervals from chronic cannulae in individual toadfish demonstrated that circulating AVT concentrations are low (10 −12–10 −11 M), and show no relationship to the occurrence of natural urea pulses. In contrast, plasma cortisol levels decline greatly prior to natural pulses and rise rapidly thereafter. AVT injections into the caudal artery or ventral aorta elicited pulse events, but these were extremely small (1–10%) relative to natural pulses, and occurred only at unphysiological dose levels (10 −9 M in the plasma). AVP was a partial agonist, but isotocin, insulin-like growth factor-1, and atrial natriuretic peptide were without effect at the same concentration. Artificially raising plasma cortisol levels by cortisol injection tended to reduce responsiveness to AVT. Pharmacological reduction of plasma cortisol levels by metyrapone injection elicited small pulses similar to those caused by AVT. Following such pulse events, AVT was ineffective in inducing pulses. We conclude that decreases in circulating cortisol play an important permissive role in urea pulsing, but that circulating AVT levels are not involved.