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

Abstract

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