Lea R. Medeiros

Fluoxetine treatment affects nitrogen waste excretion and osmoregulation in a marine teleost fish.

Measurable quantities of the selective serotonin reuptake inhibitor (SSRI), fluoxetine, have been found in surface waters and more recently in the tissues of fish. This highly prescribed pharmaceutical inhibits the reuptake of the monoamine, serotonin (5-HT; 5-hydroxytryptamine), causing a local amplification of 5-HT concentrations. Serotonin is involved in the regulation of many physiological processes in teleost fish including branchial nitrogen excretion and intestinal osmoregulation. Since the gill and intestine are directly exposed to the environment, environmental exposure to fluoxetine has the potential of affecting both these mechanisms. In the present study, we test the potential sensitivity of these processes to fluoxetine by implanting gulf toadfish, Opsanus beta, intraperitoneally with different concentrations of fluoxetine (0 (control), 25, 50, 75 and 100 microgg(-1)). Fluoxetine treatments of 25 and 50 microgg(-1) were sub-lethal and were used in subsequent experiments. Fish treated with both 25 and 50 microgg(-1) fluoxetine had significantly higher circulating levels of 5-HT than control fish, suggesting that any 5-HT sensitive physiological process could potentially be affected by these two fluoxetine doses. However, only fish treated with 25 microgg(-1) fluoxetine showed a significant increase in urea excretion. A similar increase was not measured in fish treated with 50 microgg(-1) fluoxetine, likely because of their high circulating levels of cortisol which inhibits urea excretion in toadfish. Intestinal fluid absorption appeared to be stimulated in fish treated with 25 microgg(-1) fluoxetine but inhibited in 50 microgg(-1) treated fish. Despite these differing responses, both doses of fluoxetine resulted in lowered plasma osmolality values, which was expected based on the stimulation of fluid absorption in the 25 microgg(-1) fluoxetine-treated fish but is surprising with the 50 microgg(-1) treated fish. In the case of the latter, the corresponding stress response invoked by this level of fluoxetine may have resulted in an additional osmoregulatory response which accounts for the lowered plasma osmolality. Our findings suggest that branchial urea excretion and intestinal osmoregulation are responsive to the SSRI, fluoxetine, and further investigation is needed to determine the sensitivity of these processes to chronic waterborne fluoxetine contamination.

The serotonin subtype 1A receptor regulates cortisol secretion in the Gulf toadfish, Opsanus beta.

It is well established that serotonin (5-HT; 5-hydroxytryptamine) plays a role in mammalian regulation of the hypothalamic-pituitary-adrenal (HPA) axis via the 5-HT receptor subtype 1A (5-HT(1A)). To date, there has not been a comprehensive investigation of the molecular, pharmacological and physiological aspects of the 5-HT(1A) receptor and its role in the activation of the hypothalamic-pituitary-interrenal (HPI) axis in teleost fish. The 5-HT(1A) receptor of the Gulf toadfish (Opsanus beta) was cloned and sequenced, showing 67.5% amino acid similarity to the human homologue. The 5-HT(1A) receptor was distributed throughout the brain, with the whole brain containing significantly higher levels of 5-HT(1A) mRNA compared to all other tissues and the midbrain/diencephalon region containing significantly higher levels of transcript than any other brain region. Substantial levels of transcript were also found in the pituitary, while very low levels were in the kidney that contains the interrenal cells. Xenopus oocytes injected with toadfish 5-HT(1A) receptor cRNA displayed significantly higher binding of [(3)H]5-HT that was abolished by the mammalian 5-HT(1A) receptor agonist, 8-OH-DPAT, indicating a conserved binding site of the toadfish 5-HT(1A) receptor and a high specificity for the agonist. Supporting this, binding of [(3)H]5-HT was not affected by the mammalian 5-HT(1B) receptor agonist, 5-nonyloxytryptamine, the 5-HT(7) receptor antagonist, SB269970, or the 5-HT(2) receptor agonist, alpha-methylserotonin. Confirming these molecular and pharmacological findings, intravenous injection of 8-OH-DPAT stimulated the HPI axis to cause a 2-fold increase in circulating levels of cortisol. The present study of the 5-HT(1A) receptor in a single teleost species illustrates the high conservation of this 5-HT receptor amongst vertebrates.

Cortisol-mediated downregulation of the serotonin 1A receptor subtype in the Gulf toadfish, Opsanus beta

In both mammals and teleost fish, serotonin stimulates cortisol secretion via the 5-HT1A receptor. Additionally, a negative feedback loop exists in mammals whereby increased circulating levels of cortisol inhibit 5-HT1A receptor activity. To investigate the possibility of such a feedback mechanism in teleosts, plasma cortisol levels and signaling in Gulf toadfish (Opsanus beta) were manipulated and the role of cortisol in the control of 5-HT1A evaluated. Despite a significant 4-fold increase in plasma [cortisol], crowded toadfish expressed similar amounts of 5-HT1A mRNA transcript as uncrowded toadfish; whereas, cortisol-implanted fish possessed 41.8% less 5-HT1A mRNA transcript compared to vehicle-implanted controls. This cortisol effect appeared to be reversed in RU486-injected fish, which blocks glucocorticoid receptors, as these fish expressed nearly twice as much 5-HT1A receptor transcript as the vehicle-injected fish despite significantly elevated cortisol levels. The binding affinity for the 5-HT1A receptor in the brain did not vary between any groups; however, maximum binding was significantly higher in uncrowded toadfish compared to crowded, and the same significant difference was observed between the maximum binding of vehicle and cortisol-implanted fish. The opposite trend was seen in RU486-injected and vehicle-injected fish, with RU486-injected fish having significantly higher maximal binding compared to vehicle-injected controls. Injection with the 5-HT1A receptor agonist 8-hydroxy-2-(di-n-propylamino)tetralin revealed an inhibition of cortisol secretion that was independent of 5-HT1A transcript and protein binding. These results suggest that cortisol plays a role in regulating the 5-HT1A receptor via GR-mediated pathways; however, further study is necessary to elucidate how and where this inhibition is mediated.

Elevated cortisol inhibits adrenocorticotropic hormone- and serotonin-stimulated cortisol secretion from the interrenal cells of the Gulf toadfish (Opsanus beta)

Stimulation of the toadfish 5-HT(1A) receptor by serotonin (5-hydroxytryptamine; 5-HT) or 8-OH-DPAT, a 5-HT(1A) receptor agonist, results in a significant elevation in plasma cortisol. Conversely, chronic elevation of plasma cortisol has been shown to decrease brain 5-HT(1A) receptor mRNA and protein levels via the glucocorticoid receptor (GR); however, there appears to be a disconnect between brain levels of the receptor and cortisol release. We hypothesized that elevated plasma cortisol would inhibit both adrenocorticotropic hormone (ACTH)- and 5-HT-stimulated cortisol release from the interrenal cells of Gulf toadfish, that ACTH sensitivity would not be GR-mediated and 5-HT-stimulated cortisol release would not be via the 5-HT(1A) receptor. To test these hypotheses, interrenal cells from uncrowded, crowded, vehicle-, and cortisol-implanted toadfish were incubated with either ACTH, 5-HT or 5-HT receptor agonists, and cortisol secretion was measured. Incubation with ACTH or 5-HT resulted in a stimulation of cortisol secretion in uncrowded toadfish. Cortisol secretion in response to ACTH was not affected in crowded fish; however, interrenal cells from cortisol-implanted toadfish secreted significantly less cortisol than controls, a response that was not reversed upon treatment with the GR antagonist RU486. 5-HT-stimulated cortisol release was significantly lower from both crowded and cortisol-implanted toadfish interrenal cells compared to controls. Incubation with either a 5-HT(4) or a 5-HT(2) receptor agonist significantly stimulated cortisol secretion; however, incubation with 8-OH-DPAT did not, suggesting that the 5-HT(1A) receptor is not a mediator of cortisol release at the level of the interrenal cells. Combined, these results explain in part the disconnect between brain 5-HT(1A) levels and cortisol secretion.

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-HT(2A)) 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-HT(2A) 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-HT(2A) receptor antagonist, on resting rates of pulsatile urea excretion. The full-length toadfish 5-HT(2A) receptor encodes a 496 amino acid sequence and shares 57-80% sequence identity to 5-HT(2A) receptors of other organisms, with 100% conservation among important ligand-binding residues. Toadfish 5-HT(2A) 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-HT(2A) receptor mRNA expression within the brain was highest in the hindbrain, telencephalon and midbrain/diencephalon regions. Treatment with the 5-HT(2A) 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-HT(2A) receptor in the regulation of pulsatile urea excretion in toadfish.

Demographic Parameters of Yellowfin Croaker, Umbrina roncador (Perciformes: Sciaenidae), from the Southern California Bight.

ABSTRACT The yellowfin croaker, Umbrina roncador Jordan & Gilbert, 1882, is a common nearshore and surf-zone species in the southern California bight. Age was determined for individuals (n = 1,209) using annual increments in otoliths, and size at age was modeled using the von Bertalanffy growth curve (L∞ = 307.754 mm, k = 0.278 yr−1, t0 = −0.995 yr; maximum age = 15 yr). Females (L∞= 313.173 mm, k = 0.307 yr−1, t0 = −0.771 yr) grew significantly faster and larger than males (L∞= 298.886, k = 0.269 yr−1, t0 = −1.072 yr). Age and growth modeling based upon otolith length (OL) and width (OW) measurements were assessed and were consistent with body measurements. Males and females were found in all size classes and in an overall 51.49 ratio that was not significantly different from a 50% sex ratio, suggesting that these fish are gonochores. Fish were reproductive during summer months, with gonadosomatic indices (females, 5.65%; males, 5.51%) consistent with group-spawning fishes. Data from two separate monitoring programs indicated that yellowfin croaker catch-per-unit-effort (CPUE) fluctuated appreciably from 1992 to 2006 on both spatial and temporal scales. CPUE also declined significantly in the latter years of these programs. Based on samples collected between 2003 and 2004, an estimate of overall annual total mortality was A = 0.4492, and instantaneous coefficient of total mortality was estimated at Z = 0.5964. Recruitment year classes were back calculated using annual survivorship. Year class strength was variable and declined significantly by the end of this study. Considering the high temporal and spatial variation in estimates of abundance and recruitment, coupled with the likelihood that these fish employ a probable group-spawning reproductive behavior, we recommend a cautious approach for the future management of this species.

Patterns of Life History and Habitat Use of an Important Recreational Fishery Species, Spotfin Croaker, and Their Potential Fishery Implications

Abstract Spotfin croakers Roncador stearnsii, a prized recreational catch, were collected throughout the Southern California Bight, primarily as bycatch from a long-term, scientific gill-net collection effort. The maximum otolith-based age in the present study was 24 years—14 years greater than in a previous scale-based aging study. Multiple models were used to estimate mean length at age, including models that utilize larvae as well as juveniles and adults, and the model selection results suggest sexual dimorphism in growth patterns. The juvenile and adult catch per unit effort reflected a clear pattern of habitat selectivity, with fish strongly preferring soft-bottom habitats. Catches in rocky-reef areas were limited but tended to increase with water temperature. The data also suggest that spotfin croakers segregate themselves sexually during the spawning season, when recreational fishing from jetties will target males and fish caught in bays and estuaries are more likely to be spawning females. These results provide further evidence for the importance of protection and restoration efforts for estuaries and bays along this well-developed coastline. The growth of larvae captured in plankton tows in July and September 2004 was substantially faster than that of larvae sampled in May, which coincided with warmer sea surface temperatures, highlighting a potentially important relationship relating temperature (and therefore geography) and settlement success.