Effect of microinjection and two types of electrical stimuli on bovine sperm‐hamster egg penetration

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Abstract
These experiments were designed to test the effects of an electrofusion and an electroporation pulse on bovine sperm‐hamster egg development. In experiment 1, single motile sperm were injected into the perivitelline space of each egg. A 4,500 V/cm, 30 μsec fusion pulse (FP) was applied while sperm‐egg membrane contact was maintained. It was observed that single motile sperm were rendered immotile immediately after FP application whereas nonpulsed single motile sperm remained motile for up to 36 h postinjection. In addition, both motile and sanicated spermatozoa were injected directly into the ooplasm prior to receiving an FP to determine whether the FP was detrimental to sperm viability. In experiment 2, to induce the acrosome reaction, an 1,150 V/cm electroporation pulse was applied to washed bovine sperm suspended in TALP medium containing 5 mM Ca2+. Treated and nontreated sperm were coincubated with zona‐free hamster ova, and sperm‐penetrating ability was measured. Results from experiment 1 indicate that FP failed to induce sperm‐egg fusion (0/69). FP did not, however, inhibit decondensation or pronuclear formation of sperm injected into hamster egg ooplasm. Single motile sperm injected into the ooplasm resulted in development of both pulsed (19/28) and nonpulsed (21/28) groups. Sonicated tail‐free sperm heads injected into the ooplasm resulted in no detectable difference between treated (18/30) and nontreated (19/30) groups. In experiment 2, treatment of sperm with electroporation pulse +5 mM Ca2+ increased zona‐free hamster ova penetration scores over nontreated sperm within bulls (P < .05). These results indicate that an electroporation pulse in conjunction with high Ca2+ rather than an electrofusion pulse facilitates sperm penetration and early development.