In Vitro Culture Conditions for Reproductive Endocrine Tissues of Fathead Minnow (Pimephales promelas)

Abstract

Studies examined the effects of different culture conditions for the brain, gonads (testis and ovary), and liver of adult male and Female Fathead Minnow (Pimephales promelas) (FHM) during a 28 day assessment of viability and functionality. Five parameters were tested including media pH, incubation temperature, media composition, media exchange rate, and substrate. Within the ranges tested, incubation temperature was the most influential on tissue viability with dramatic improvements using cooler incubation temperatures (<18°C) over the entire 28 day test period. Tissues remained equally viable in five different common culture media, although highest viability with each medium was observed with a weekly media exchange over more frequent exchanges, alginate over polystyrene, and a media pH near physiological values. Use of these combined conditions resulted in continual function of testis and ovarian tissue over the entire 28 days, as indicated by continued production of 11-ketotestosterone (11-KT) by testes and estradiol (E2) by ovaries. Liver function was maintained through seven days, although vitellogenin production in response to added E2 eventually decreased over time. Consistent tissue viability over time periods commonly used for in vivo studies will enhance the link of in vitro tissue toxicology studies to whole-fish and population level impacts.