Temporary inhibition of meiosis and chemical lipolysis in oocyte: innovations in bovine embryos in vitro production

Abstract

The cilostamide and forskolin have been effective in delaying meiosis to be activators of cAMP. The forskolin is also used to decrease the amount of lipid droplets present in the cytosol of oocyte during IVM. This project aims to evaluate the effect of forskolin and cilostamide as attenuation of spontaneous meiosis strategy and thus to improve the in vitro production of bovine embryos, as well as evaluating the chemical induction of lipolysis during IVM of oocytes to improve the quality and cryotolerance of IVP embryos. In Experiment I, the oocytes will be matured with forskolin (50 µM) and cilostamide (20 µM) for 6 h, alone or together to cause a temporary inhibition of the spontaneous meiosis resumption, and then be matured in the absence of these drugs for 18 h to meiosis progression and also in the presence or absence of FSH (1 mg / mL). After 24 h of IVM, their nucleus will be evaluated. They will be evaluated for production of embryos (cleavage and blastocyst rate, apoptotic cells of embryos) after IVF and IVC. In Experiment II, oocytes will be matured with forskolin (50 µM) in three periods to evaluate lipolysis: 6 h with forskolin and 18 h without forskolin; 18 h with forskolin and 6 h without forskolin; and 24 h with forskolin. These same groups will be evaluated in the presence or absence of FCS (10 %). After maturation, the lipids are quantified with Nile Red technique to determine the degree of lipolysis. In the second stage the same groups will be evaluated after IVF and IVC in relation of embryos production (cleavage and blastocyst rate), embryo quality (apoptosis rate) and vitrification survive (reexpansion and apoptosis rate). (AU)