The thyroid gland and thyroid hormones in sheepshead minnow (Cyprinodon variegatus) during early development and metamorphosis

The sheepshead minnow is widely used in ecotoxicological studies that recently, begin to focus on potential disruption of the thyroid axis by xenobiotics and endocrine disrupting compounds. However, reference levels of thyroxin (T4) and 3,5,3’-triiodothyronine (T3) and their developmental patterns are unknown. This study set out to describe the ontogeny and morphology of the thyroid gland in sheepshead minnow, and correlate these with whole body concentrations of thyroid hormones during early development and metamorphosis. Couples of three females and two males were placed in breeding chambers designed for this experiment. More than 1000 eggs were collected and maintained in seawater. Embryos were selected under a dissection microscope and placed in incubation dishes (50 per dish) at 25°C. On day 6, embryos hatched and larvae were transferred to 1L beakers. For one week after hatching, larvae were fed on artemia, and from 8 to 30 days post-hatch they were fed on flaked fish food. Embryos were sampled on day 0, 3, 6 post-fertilization and larvae and juveniles were sampled every three days from day 0 to 28 days post-hatch. The pooled samples were taken from several incubation dishes and divided in three replicate batches of 10-30 individuals. T4 and T3 were extracted from whole fish bodies and an enzyme-linked immunoassay was used to measure whole-body hormone levels. At each sampling point 5 individuals were placed in formalin fixative for histology. Length and body mass were measured. Hatching success, gross morphology, thyroid hormone levels and histology data were recorded. The onset of metamorphosis at 12 days post-hatching coincided with surges in whole body T4 and T3 concentrations. Thyroid follicles were first observed in pre-metamorphic larvae at hatching, and were detected exclusively in the subpharyngeal region, surrounding the ventral aorta. Follicle size and epithelial cell heights varied during the developmental phase, indicating fluctuations in thyroid hormone synthesis activity. The increase in the whole body T3/T4 ratio was indicative of an increase in outer ring deiodination. This study establishes a baseline for thyroid hormones in sheepshead minnows, which will be vital for the understanding of thyroid hormone functions and in future studies of thyroid toxicants in this species

Evaluation of the impact of Triclosan on the functioning of the thyroid system in Cyprinodon variegatus L., 1803.

Triclosan (5-chloro-2-[2, 4-dichlorophenoxy] phenol) is an antimicrobial widely used in various industrial products such as textiles, cosmetics and body care products. It is often detected in aquatic environments. The presence of the main biotransformation product, methyl TCS, indicates that this compound is not only degraded, but also persistent and accumulates in aquatic organisms. In this study, the effects of TCS on the thyroid system during embryonic and larval stages in Cyprinodon variegatus were evaluated. In particular, whole body thyroid hormone levels and the activity of deiodinases, enzymes involved in the activation of the thyroid prohormone T4, were measured. Couples of three females and two males were placed in breeding chambers designed for this experiment from which 3832 eggs were obtained by reproduction in the laboratory. Eggs were collected and maintained in seawater. Embryos were selected under a dissection microscope, randomly assigned to each of five treatment groups: Control, DMSO control (vehicle), 20 μg/L TCS, 50 μg/L TCS and 100 μg/L TCS and placed in incubation dishes (50 per dish) at 25°C. On day 6, embryos hatched and larvae were transferred to 1L dishes. The larvae were fed on artemia and on flaked fish food till day 15 post hatching when the fish were analyzed. The fertilization and survival rate, as well as the larval dry weight did not vary significantly between individuals exposed to 20, 50 and 100 μg/L TCS. T3 and T4 concentrations increased significantly in larvae exposed to 50 and 100 μg/L TCS. The study of the activity of enzymes involved in the deiodination of thyroid hormones (ORD) represents a new aspect in the study of endocrine disruption in C. variegatus. Unfortunately, we were not able to detect a net enzymatic T4 deiodination activity, most likely due to the very small amounts of protein and low specific enzyme activity in brain homogenates, carcass and liver

Identification and Functional Characterization of Zebrafish Solute Carrier Slc16a2 (Mct8) as a Thyroid Hormone Membrane Transporter

Most components of the thyroid system in bony fish have been described and characterized, with the notable exception of thyroid hormone membrane transporters. We have cloned, sequenced, and expressed the zebrafish solute carrier Slc16a2 (also named monocarboxylate transporter Mct8) cDNA and established its role as a thyroid hormone transport protein. The cloned cDNA shares 56-57% homology with its mammalian orthologs. The 526-amino-acid sequence contains 12 predicted transmembrane domains. An intracellular N-terminal PEST domain, thought to be involved in proteolytic processing of the protein, is present in the zebrafish sequence. Measured at initial rate and at the body/ rearing temperature of zebrafish (26 C), T(3) uptake by zebrafish Slc16a2 is a saturable process with a calculated Michaelis-Menten constant of 0.8 mu M T(3). The rate of T3 uptake is temperature dependent and Na(+) independent. Interestingly, at 26 C, zebrafish Slc16a2 does not transport T(4). This implies that at a normal body temperature in zebrafish, Slc16a2 protein is predominantly involved in T(3) uptake. When measured at 37 C, zebrafish Slc16a2 transports T(4) in a Na(+)-independent manner. In adult zebrafish, the Slc16a2 gene is highly expressed in brain, gills, pancreas, liver, pituitary, heart, kidney, and gut. Beginning from the midblastula stage, Slc16a2 is also expressed during zebrafish early development, the highest expression levels occurring 48 h after fertilization. This is the first direct evidence for thyroid hormone membrane transporters in fish. We suggest that Slc16a2 plays a key role in the local availability of T(3) in adult tissues as well as during the completion of morphogenesis of primary organ systems. (Endocrinology 152: 5065-5073, 2011

Iron metabolism in mynah birds (Gracula religiosa) resembles human hereditary haemochromatosis

Iron overload is a very frequent finding in several animal species and a genetic predisposition is suggested. In one of the most commonly reported species with susceptibility for iron overload ( mynah bird), it was recently shown that the cause of this pathophysiology is high uptake and retention of dietary iron. Here we compare susceptible ( mynahs) with non-susceptible avian species ( chickens) by evaluating iron uptake at the intestinal absorptive cell level. Enterocytes from mynahs and chickens were isolated and uptake of Fe(II) and Fe(III) was studied in vitro. It was found that Fe( III) uptake is much lower than Fe( II) uptake for both species. Although liver iron, present only in hepatocytes, was at least 10-fold higher in mynahs than chickens, enterocyte Fe( II) uptake was considerably higher in mynahs. Fe( II) uptake showed saturation at the studied concentrations in both species. Kinetic studies revealed a three-fold increase in V-max for mynahs. Calculated values for the uptake kinetics of the probable membrane transporter suggest that mynah bird enterocytes have a significantly higher limiting uptake rate, due to the possible increase in the number of transporters when compared with chicken enterocytes. The susceptibility of this species is due to intestinal iron uptake despite hepatic iron accumulation, implicating a 'mis-sensing' of body iron similarly to human hereditary haemochromatosis

Acclimation of solea senegalensis to different ambient temperatures: Implications for thyroidal status and osmoregulation

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