Electron microscope studies of sperm incorporation into the golden hamster egg

Abstract

Hamster eggs freed from the cumulus oophorus and zona pellucida were inseminated in vitro with capacitated spermatozoa. As soon as the spermatozoon reaches the surface of the egg cytoplasm, numerous microvilli of the egg surround the spermatozoon. Those microvilli attached to the acrosomal collar region of the spermatozoon quickly spread over the inner acrosomal membrane of this region, dislodging the outer acrosomal membrane and the overlying sperm plasma membrane. At the same time, microvilli fuse with the plasma membrane of the postnuclear cap region of the spermatozoon, and the egg cytoplasm quickly flows around the nucleus of that region of the spermatozoon, The inner acrosomal membrane persisting on the sperm head never fuses with the egg (microvillar) plasma membrane. This membrane is destined to disintegrate and be absorbed into the egg cytoplasm. Incorporation of the sperm flagellum into the egg cytoplasm takes place by fusion between plasma membranes of the spermatozoon and egg.Penetration of mammalian eggs by spermatozoa has been studied by electron microscopy in the rat (Szollosi and Ris, '61; Pik6, '64, '67, '69; Pik6 and Tyler, '64; Merker and Koehler, '68; Merker and Neubert, '69), rabbit (Moricard et al., '62; Hadek, '63a,b, '64, '66, '69; Austin, '63; Bedford, '67, '68), mouse (Stefanini et al., '69), golden hamster (Hadek, '64, '69; Barros and Franklin, '68; Yanagimachi and Noda, '70a) and Chinese hamster (Pik6, '64).In normal fertilization of the mammalian egg, only one spermatozoon enters the egg cytoplasm. Because of the lack of an appropriate technique to control the time of sperm penetration into the egg, the chance of obtaining thin sections with penetrating spermatozoa in favorable orientations for interpretation is limited. This difficulty can be overcome by inseminating the eggs in vitro with fully capacitated spermatozoa after removal of the egg investments (cumulus oophorus and zona pellucida). Many spermatozoa enter the egg almost immediately after insemination and their heads develop into male pronuclei (Yanagimachi and Noda, '70b). Since more than one spermatozoon (occasionally more than 20) enter one egg, thin sections revealing the spermatozoa in various phases of incorporation with the eggs are obtained easily. This paper is a record of observations on fertilization in such "naked" eggs inseminated in vitro with capacitated spermatozoa. Although such fertilization can not be considered normal, our observations with the phase-contrast microscope on living materials showed that there is no substantial difference between normal ( i n vivo) fertilization and polyspermic (in vitro) fertilization of the naked eggs as far as the early behavior of fertilizing spermatozoa is concerned. MATERIALS AND METHODSMethods for collection of unfertilized eggs from the golden hamster, removal of their egg investments (cumulus oophorus and zona pellucida) and insemination of the "naked eggs with capacitated spermatozoa in vitro have been described (Yanagimachi and Noda, '70b OBSERVAT...

Induction of Nuclear Decondensation of Mammalian Spermatozoa in Vitro

“…In spite of this, the nucleus swells and its chromatin begins to decondense very rapidly as the spermatozoon is incorporated into the egg cytoplasm (Austin, 1961;Yanagimachi & Noda, 1970;Bedford, 1972). As yet, the mechanism underlying this phenomenon has not been directly examined.…”

In vitro and in vivo studies of mammalian sperm nuclear decondensation

“…Transmission electron microscopy of early stages of decondensation (Fig. 4) revealed chromatin fibrils extending from a condensed mass in the postacrosomal region of the nucleus, similar to the morphology of the sperm nucleus during early stages of fertilization [Yanagimachi and Noda, 1970;Bedford, 19721. Incubation of sperm nuclei in DTT and anionic detergents such as sodium dodecylsulfate, deoxycholate, and sarkosyl also resulted in nuclear decondensation .…”

Singular features of fertilization and their impact on the male reproductive system in eutherian mammals

“…Perhaps most striking in this regard is that, after negotiating the egg coat, eutherian spermatozoa do not fuse conventionally with the oolemma via the apex of the inner acrosomal membrane (IAM), but rather fuse by the mid-region of the sperm head, followed by the egg's 'phagocytosis' of the acrosomal domain (Yanagimachi & Noda 1970a, Bedford 1972. Underlying this mode, the eutherian acrosome alone displays an attenuated stable posterior region -the equatorial segment, which is unique to eutherian mammals and whose significance lies in its ability to preserve a section of fusogenic plasma membrane overlying it ).…”