Endocrine disrupting compounds (EDCs), such as Bisphenol A and phthalates, are found ubiquitously in food and water and are of toxicological human health concern especially in developing children and women in their child-bearing age. Exposures to EDCs have recently been suspected to be part of the cause for a significant trend of early puberty onset observed in children over the past few decades in many industrialized countries including the U.S. However the mechanism of how this may occur is not well understood. Increased frequency and amplitude of the pulsatile release of gonadotropin releasing hormone (GnRH) is considered to have a central role in initiating puberty in humans and animals. There has been recent discovery of upstream positive and negative regulators of GnRH, called kisspeptin and gonadotropin inhibitory hormone (GnIH). In this proposed project, early developmental stage zebrafish will be used to test the central hypothesis that endocrine disrupting chemicals affect the pulsatility of GnRH neurons by affecting upstream regulators, kisspeptin and GnIH. It has been technically difficult to monitor the expression, excitation, and synchronization of GnRH neurons in vivo due to its location deep inside the brain. Development of two zebrafish transgenic lines consisting of short-life fluorescence-labeled GnRH will allow such visualization; first one to monitor mRNA expression and the other to monitor excitation, and both will ultimately allow for use in screening for potential endocrine disrupting compounds in the future. These transgenic fish larvae will be treated with Bisphenol A and Bis(2-ethylhexyl) phthalate and time-lapse images will be recorded with a multiphoton fluorescence microscope. Lastly, roles of kisspeptin and GnIH will be investigated with the EDC exposed larvae by morpholino experiments in the transgenic fish lines. Goal of this project is to understand the molecular toxicology mechanism of GnRH disruption by EDCs. Broad and long term goal of this project is to develop tools for high-throughput screening of chemical compounds in order to protect reproductive health in humans. The PD/PI of this project, Dr. Motoko Mukai, DVM, Ph.D. is a board certified toxicologist with clinical diagnostic experiences in animal feed/food safety. She will be mentored by Dr. Joseph Fetcho, a well- established leading neurobiologist, who has been using zebrafish larvae to study neurogenesis, neurodynamics, neural circuits, and regeneration of neurons. Dr. Richard Peterson from University of Wisconsin, Madison, expert in a study of endocrine disruptor on reproduction system will serve as a co-Mentor on this project for molecular toxicology guidance.
Exposures to endocrine disrupting chemicals have recently been suspected to be part of the cause for the significant advancement of puberty onset observed in children in the last few decades. This project is designed to understand the underlying molecular mechanisms using dynamic fluorescently-labeled transgenic zebrafish models. The findings of this research project will provide useful resources and knowledge to develop efficient screening tools for chemical compounds and protect human reproductive health.
