BOVINE AND BABOON OOCYTE DEVELOPMENT IN VITRO

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In female mammals most oocytes reside in primordial follicles in a resting stage. Virtually nothing is known about the mechanisms that regulate the movement of these follicles into the pool of growing follicles. The pool of resting primordial follicles is a resource, as yet untapped, that could be exploited as source of material for in vitro maturation (IVM) and in vitro fertilization (IVF) to hasten genetic improvement of domestic species, preserve endangered species, and provide alternative methods for alleviating infertility in women. We have developed a culture system that supports activation of primordial follicles in organ cultures of bovine and baboon ovarian cortex. Cortical explants survive in culture in serum-free conditions for at least 7-10 days and a high percentage of primordial follicles leave the resting pool and progress to the primary follicle stage, but only a few follicles develop to the multilayered secondary stage. We now propose to build on these striking findings and initiate a new project to determine culture conditions that will support further development of follicles in vitro. Although cattle will be the primary animal model for these studies, we will pursue parallel studies with a primate model, the baboon, because we have unique access to fetal baboon ovaries at little cost. The overall hypothesis is that initiating growth of primordial follicles to the secondary follicle stage in organ cultures of ovarian cortex, followed by enzymatic or mechanical isolation of secondary follicles and their further culture as individual follicles, will allow follicular growth to progress in vitro to a stage where oocytes can be matured in vitro. Specific Aim #1 is to test the hypothesis that bovine and baboon follicles can develop in vitro from the primordial to the secondary stage provided that culture conditions can be improved to extend survival. Specific Aim #2 is to develop methods for growing bovine secondary follicles to the stage of competence for IVM. These experiments are crucial steps towards the ultimate goal of using the large numbers of primordial follicles in mammalian ovaries as a resource for genetic improvement of domestic animals, preservation of endangered species, and alleviation of human infertility.