Teofilina associada ou não à heparina como agente capacitante para produção in vitro de embriões bovinos

Abstract

The in vitro production (IVP) of bovine embryos despite already used on a commercial scale, presents high variability in the results, that can often be attributed to the fertilization step. Therefore, it is important to research factors related to semen, such as sperm capacitation process. The aim of the study was to evaluate theophylline or its combination with heparin as possible replacement capacitation agent in the development of in vitro produced embryos and in acrosome reaction of sperm cells. The experiment was carried out with 4 bulls and 3 treatments, with 12 experimental groups. Each bull was evaluated in all treatments as described below: Treatment 1 (T1): Heparin - 10mg / ml; Treatment 2 (T2): Theophylline - 5 mM; Treatment 3 (T3): Heparin (10 mg / ml) + theophylline (5mM). The bulls semen was thawed and exposed to incubation in three treatments for 0, 6, 12 and 18h, subsequently stained with Trypan blue / Giemsa and analyzed by electron microscopy for assessment of the acrosome reaction. For IVP of embryos capacitation agents added to fertilization medium were evaluated utilizing the same bulls. In sperm analysis, real acrosome reaction rate was higher (p<0,05) at 0h time while the higher dead spermatozoa rate (p<0,05) was observed at 12 h (84,46 ± 5,82 ) and 18h (86,75 ± 4,19). The IVP embryos rate (37,97 ± 13) and the hatching rate (33,50 ± 14) were higher (p<0,05) in heparin treatment. There was no influence of bull or treatments (p> 0.05) in the acrosome reaction analysis. The use of theophylline as a capacitation agent decreased embryo production rates in IVF, however was as efficient as heparin in acrosome reaction induction.