Molly H. B. Amador

Molecular and functional characterization of the Gulf toadfish serotonin transporter SLC6A4

ABSTRACT The serotonin transporter (SERT) functions in the uptake of the neurotransmitter serotonin (5-HT) from the extracellular milieu and is the molecular target of the selective serotonin re-uptake inhibitors (SSRIs), a common group of anti-depressants. The current study comprehensively assesses the sequence, tissue distribution, transport kinetics and physiological function of a teleost SERT. The 2022 bp toadfish SERT sequence encodes a protein of 673 amino acids, which shows 83% similarity to zebrafish SERT and groups with SERT of other teleosts in phylogenetic analysis. SERT mRNA is ubiquitous in tissues and is expressed at high levels in the heart and, within the brain, in the cerebellum. SERT cRNA expressed in Xenopus laevis oocytes demonstrates a Km value of 2.08±0.45 μmol l–1, similar to previously reported Km values for zebrafish and human SERT. Acute systemic blockade of SERT by intraperitoneal administration of the SSRI fluoxetine (FLX) produces a dose-dependent increase in plasma 5-HT, indicating effective inhibition of 5-HT uptake from the circulation. As teleosts lack platelets, which are important 5-HT sequestration sites in mammals, the FLX-induced increase in plasma 5-HT suggests that toadfish tissues may normally be responsible for maintaining low 5-HT concentrations in the bloodstream. Summary: The toadfish serotonin transporter is highly conserved with that of other teleosts and is ubiquitously expressed in toadfish tissues. Systemic inhibition of the transporter produces dose-dependent elevations in plasma serotonin.

Does fluoxetine exposure affect hypoxia tolerance in the Gulf toadfish, Opsanus beta?

Due to ineffective wastewater treatment technologies, pharmaceuticals such as the selective serotonin reuptake inhibitors (SSRIs)-a common class of antidepressants which inhibit the serotonin transporter (SERT)-can be found in surface waters and marine receiving waters near wastewater effluents. Understanding how exposure to these chemicals might impact non-target organisms, especially combined with other environmental stressors like hypoxia, is essential in order to thoroughly evaluate environmental risk. It was hypothesized that both acute and chronic exposure to the SSRI fluoxetine (FLX) would interfere with the metabolic hypoxia response of the Gulf toadfish, Opsanus beta. Here we demonstrate that acute intraperitoneal treatment with 50 μg g-1 FLX significantly reduces the regulation index, or degree of metabolic regulation, in toadfish. Acute FLX exposure significantly reduced SERT mRNA expression in the first and third gill arches, but mRNA expression was not affected in heart tissues or in the second gill arch. In contrast, the regulation index was unaffected by 14-17 day waterborne FLX exposure to environmentally relevant (0.01 μg L-1) and approximately 1000-fold higher (8.5 μg L-1) concentrations. However, the higher concentration was sufficient to induce a systemic elevation in plasma serotonin concentrations. Chronic FLX exposure did not alter SERT mRNA expression in heart or gill tissues. The results of this study implicate the involvement of 5-HT pathways in hypoxia tolerance but demonstrate that current environmental levels of FLX are insufficient to impair the metabolic hypoxia response in marine fish.

In rough waters, sea bass should stick together

![Graphic][1] Schooling behaviour in fishes reduces the energetic costs of swimming for the schools members by essentially allowing each fish to ‘draft’ off of others in the group. This phenomenon has been thoroughly studied in laboratory settings under conditions of laminar flow –

Fly breath sets mites up for success

![Graphic][1] For parasites to thrive, they must first locate and then derive resources from a host. Some parasites can use chemical cues – such as CO2 released through respiration – to find hosts, and when multiple hosts live in the same area, parasites often preferentially infest

Sea bass (can't) smell trouble under elevated CO2

![Graphic][1] Elevated levels of dissolved carbon dioxide (CO2), such as those expected by the end of this century, have been shown to harm marine fish by affecting how they sense and interact with their environment, ultimately increasing their mortality. The sensory and behavioural changes

The serotonin transporter (SERT) and non-selective transporters are involved in peripheral serotonin uptake in the Gulf toadfish, Opsanus beta

In mammals, circulating serotonin [5-hydroxytryptamine (5-HT)] is sequestered by platelets via the 5-HT transporter (SERT) to prevent unintended signaling by this potent signaling molecule. Teleost fish appear to lack a similar circulating storage pool, although the diverse effects of 5-HT in teleosts likely necessitate an alternative method of tight regulation, such as uptake by peripheral tissues. Here, a 5-HT radiotracer was used to explore the 5-HT uptake capacity of peripheral tissues in the Gulf toadfish, Opsanus beta, and to elucidate the primary excretion routes of 5-HT and its metabolites. Pharmacological inhibition of SERT and other transporters enabled assessment of the SERT dependence of peripheral 5-HT uptake and excretion. The results indicated a rapid and substantial uptake of 5-HT by the heart atrium, heart ventricle, and gill that was at least partly SERT dependent. The results also supported the presence of a partial blood-brain barrier that prevented rapid changes in brain 5-HT content despite fluctuating plasma 5-HT concentrations. The renal pathway appeared to be the dominant excretory route for 5-HT and its metabolites over shorter time frames (up to ~30 min), but hepatic excretion was substantial over several hours. SERT inhibition ultimately reduced the excretion of 5-HT and its metabolites by urinary, biliary, and/or intestinal pathways. In addition, branchial excretion of 5-HT and its metabolites could not be ruled out. In summary, this study reveals that the toadfish heart and gill play active roles in regulating circulating 5-HT and yields important insights into the control of peripheral 5-HT in this teleost fish.

Unearthing HIFs in a hardy marsh fish

![Graphic][1] When it comes to surviving hypoxia, or low oxygen, animals get by with a little help from their factors – hypoxia-inducible factors (HIFs), that is. HIFs are useful molecules that, when activated by hypoxia, help form switches to crank gene expression up or down as needed,

Honeybees remember that size matters

![Graphic][1] Honeybees likely use relational information to decide which flowers are optimal for foraging nectar. For instance, laboratory research has shown that the bees are able to discriminate between sizes of stimuli as well as their relative positions and can apply learned rules to

[Cardiac] Pressure's on for thirsty trout

![Graphic][1] Because it is much saltier than their body fluids, the ocean is a very dehydrating environment for fishes. Species that can survive in both freshwater and saltwater, such as trout and salmon, must therefore undergo significant physiological changes when transitioning to seawater

Hagfish pump some iron

![Graphic][1] Along with other nutrients, animals need trace metals to function. Iron, the oxygen-binding component in the blood pigment hemoglobin, is a critical trace metal and is usually absorbed through the gut. Mammals can alter iron metabolism in order to meet oxygen demand during