Delayed sperm incorporation into parthenogenetic mouse eggs: sperm nucleus transformation and development of resulting embryos.

In this study we examined the effect of experimentally induced asynchrony between male and female pronuclei on male pronucleus formation and developmental potential of the resulting mouse embryos. We demonstrate that when the interval between oocyte activation and sperm incorporation is up to 1.5-2 hr, the spermatozoa transform into normal pronuclei. These male pronuclei can replicate their chromosomes during the first embryonic cell cycle and are transcriptionally competent. During the first cleavage these "delayed" male pronuclei condense into discrete mitotic chromosomes and when resulting embryos are transplanted into oviducts of pregnant females at least some of them can develop to term. In contrast, when sperm nuclei are introduced into parthenogenetic eggs 3 hr or more after activation, their transformation into pronuclei is significantly impaired, and they neither replicate nor transcribe. During the first mitosis they form a group of condensed chromatin, which is displaced into one of the resulting blastomeres leading to formation of haploid/diploid mosaic embryos. These mosaic embryos have poor developmental potential: only a few can reach blastocyst stage in vitro and no full-term development of such embryos was observed after transfer into pregnant females. We conclude that the cytoplasmic factors that make possible the transformation of a sperm nucleus into a functional male pronucleus exhaust within 1.5-2 hr after fertilization and that the male genome which had skipped the first cell cycle cannot become a functional partner in the embryonic genome.

Mouse oocytes penetrated by sperm at GV or GVBD stage lose the ability to fuse with additional spermatozoa.

Mouse oocytes penetrated by spermatozoa during germinal vesicle (GV) breakdown undergo maturation and are arrested at metaphase of the second meiotic division despite the presence of sperm nuclei within the ooplasm. When these oocytes were re-inseminated, none was penetrated by newly added spermatozoa. When GV oocytes were inseminated and cultured in the presence of dibutyryl cAMP, the oocytes remained at GV stage, yet they did not permit entry of additional spermatozoa. These observations suggest that the plasma membrane of maturing oocytes is modified by precociously penetrating spermatozoa independently from cortical granule exocytosis. Sperm components incorporated into the oocytes seem to be responsible for the modification of the oocyte's plasma membrane.

Sperm entry into fertilised mouse eggs.

We have investigated the oolemma block to polyspermy in the mouse. Zona-free and zona-intact eggs were fertilised and subsequently re-inseminated (the latter following zona pellucida removal). The 'perivitelline' block to polyspermy in zona-intact eggs renders motile sperm in the perivitelline space unable to bind to the oolemma. This is not connected with irreversible changes in the egg plasma membrane, because freshly added sperm can still fuse with such eggs freed from the zona. Fertilised eggs eventually lose the ability to fuse with sperm within 1 h, while still being able to bind many sperm.