Copyright q American Society of Andrology Influence of Sperm Pretreatment on the Efficiency of Intracytoplasmic Sperm Injection in Pigs

ABSTRACT: The purpose of this study was to determine the influ-ence of sperm pretreatment on the efficiency of intracytoplasmic sperm injection (ICSI) in pigs. This was done by examining the effect of 1) the conservation method (fresh vs frozen); 2) the sperm treat-ment preinjection (resuspension in Dulbecco phosphate-buffered sa-line (DPBS) vs selection by a Percoll gradient); and 3) the acrosomal and live or dead status of the spermatozoa (by incubation with or without calcium ionophore, 1 mM and 5 mM). In vitro matured porcine oocytes were injected with treated spermatozoa according to each experiment. All the experiments were done with non–artificially ac-tivated oocytes. The percentages of activation and cleavage were higher (68 % vs 43 % and 63 % vs 43%, respectively, P,.05) in oocytes injected with fresh vs frozen spermatozoa. The DPBS treat-ment allowed higher cleavage proportions than the Percoll treatment (P,.05). Moreover, a boar effect was observed in the percentage of developing blastocysts. None of the studied parameters was af-fected by the acrosomal or the live or dead status of the sperma-tozoa injected. In conclusion, the use of fresh semen is recom-mended for porcine ICSI, as well as careful selection of the boar; Percoll treatment is only recommended for poor-quality samples or for removing toxic agents, and no exogenous form of activation or induction of the acrosome reaction is necessary for porcine oocytes to develop a male pronucleus and cleave up to the 2-cell stage after ICSI, although experimental conditions to reach the blastocyst stage need to be investigated further. Key words: Capacitation, sperm cryopreservation, acrosome re-action, early development