Development of transgenic lines to study pituitary cell development and plasticity in medaka (Oryzias latipes)

Abstract

The reproductive system is an important process highly regulated by hormones produced by the pituitary gland, including the thyroid-stimulating hormone (Tsh) and the folliclestimulating hormone (Fsh). The pituitary is a highly plastic organ and adapts hormone production according to their demand. The mechanisms involved in this plasticity are poorly known, same as their regulation. To further study the development and plasticity of these endocrine cell populations, this project developed two new transgenic lines in the model fish Japanese medaka (Oryzias latipes). First, a transgenic line named tg(tshbb:tdTomato) was produced, in which the gene encoding the fluorescent protein tdTomato was under the control of the tshbb promoter. This line allowed us to visualize the cells expressing the tshbb gene of which the function and location remain mostly unknown to this day. A fluorescent cell population was observed between the eyes in developing transgenic medaka larvae. This could hopefully indicate that Tshbb expression has successfully been visualized for the first time in this species, whereas further studies are needed to characterize this endocrine cell population. Second, a transgenic line named tg(fshb:NTR-CFP/fshb:DsRed2) carried the gene encoding the bacteria-derived protein nitroreductase and the fluorescence proteins CFP and DsRed2 under the control of the fshb promoter. Nitroreductase expression is known to induce cellspecific apoptosis when treated with ronidazole. With these properties, the transgenic line could be useful for investigating regeneration of removed tissue. Dissected brain- and pituitary tissue was kept in specific culture conditions which were shown to minimize tissue degradation. In these conditions, high-concentration treatments of ronidazole demonstrated to ablate Fsh cells. Regeneration of these cells was unfortunately not observed in this project, so more studies are needed to characterize their role in pituitary plasticity. Further development of the transgenic lines developed in this study will allow deeper investigation of these two endocrine cell populations of the pituitary gland. The unknown function of Tshbb may finally be discovered, and the regeneration of Fsh cells may be explored deeper in the medaka.