Comparison between ejaculated and epididymal stallion sperm cryopreserved with the pellet method

In order to understand the freezability of epididymal sperm and its fertilising potential we compared the membrane integrity and the capacity to bind to the zona pellucida of frozen/thawed epididymal and ejaculated sperm. Epididymal semen (EpS) was harvested by flushing the epididymal tails of the testis obtained by standing castrations of 3-year old colts while ejaculated semen (EjS) was collected by artificial vagina. Semen was cryopreserved with the pellet method. Sperm viability and acrosome integrity were evaluated after thawing by incubating spermatozoa with propidium iodide and FITCPSA. Fertilising potential was assessed by glycerol-Hoechst staining of IVM horse and ovine oocytes coincubated with spermatozoa for 24 h. After thawing both EpS and EjS presented 50% viability, but a significant difference (Chi-Square test) was evidenced in the percentage of live acrosome reacted spermatozoa (EpS: 6%; EjS: 36.1%; p < 0:001). Zona binding test both with horse (EpS: 0.50.5; EjS: 0.30.5; p¼0.285) and ovine (EpS: 57.5; EjS: 3.65.6; p¼0.246) oocytes did not revealed any significant difference (ANOVA) between EpS and EjS. The results suggest that epididymal stallion spermatozoa can be cryopreserved successfully with a better membrane integrity and an equal capacity to bind to the zona pellucida in vitro compared to ejaculated spermatozoa.