ACROSOME STATUS AND MORPHOLOGY OF HUMAN SPERMATOZOA BOUND TO THE ZONA-PELLUCIDA AND OOLEMMA DETERMINED USING OOCYTES THAT FAILED TO FERTILIZE IN-VITRO

It is known that only acrosome-reacted spermatozoa can fuse with the oolemma during normal fertilization with zona pellucida-intact oocytes. The aim of this study was to determine if the oolemma of human zona pellucida-free oocytes selectively binds spermatozoa with normal morphology and a reacted acrosome. Oocytes that failed to fertilize in vitro because of severe sperm defects were used. The zona pellucida was removed with acidic (pH 2-3) saline. Sperm samples were obtained from normal fertile donors and normozoospermic men. Motile spermatozoa were selected with a swim-up technique and 2 x 10(6)/ml incubated with oocytes. The results from 23 experiments showed that at 2 h there was a significantly higher mean percentage of acrosome-reacted spermatozoa bound to the zona pellucida (mean +/- SD, 42 +/- 22) than in the insemination medium (27 +/- 12). In contrast, all spermatozoa bound to the oolemma at 2 h were acrosome reacted. Furthermore, each fresh zona pellucida had > 100 spermatozoa bound compared with an average of 28 (range 4-81) spermatozoa bound per zona pellucida-free oocyte. There was no significant difference in the zona pellucida-induced acrosome reaction between fresh (45 +/- 21) and salt-stored (35 +/- 22) zonae pellucidae. The percentage with normal morphology was significantly higher for spermatozoa bound to the zona pellucida (84 +/- 13) and oolemma of zona pellucida-free oocytes (71 +/- 25) than for spermatozoa in insemination medium (39 +/- 11) (P < 0.01). Extending the time of incubation of spermatozoa with zona pellucida-intact oocytes increased the proportion of spermatozoa undergoing the acrosome reaction (n = 6, 2 h, 41 +/- 23; 3 h, 53 +/- 31; 4 h, 61 +/- 34). However, there was a large variation in the percentage of acrosome reactions among spermatozoa bound to the zona pellucida between individual sperm samples. In conclusion, only acrosome-reacted spermatozoa can bind to the oolemma of zona pellucida-free oocytes. Both fresh and salt-stored human zonae pellucidae are equally effective in inducing the acrosome reaction. Both the zona pellucida and the oolemma of zona pellucida-free oocytes selectively bind spermatozoa with normal morphology.