Sperm tail entry into the mouse egg in vitro

Abstract

Cumulus‐free mouse eggs were placed on microscope slides and inseminated with capacitated mouse spermatozoa. Fertilization could then be observed through the phase contrast microscope and recorded by time‐lapse cinematography. Following the penetration of the fertilizing spermatozoon through the zona pellucida and the fusion of the sperm head with the vitelline membrane, the entire sperm tail gradually entered the vitellus. The time required for tail incorporation into the vitellus as measured in 49 eggs varied from 3 h 3 min to 5 h 49 min, with a mean time of 4 h 23 min. When tail incorporation began, the greater part of the flagellum was still outside the zona pellucida; occasionally it slipped into the perivitelline space, but generally it remained outside the zona and shortened by degrees as incorporation proceeded. The motility of the fertilizing spermatozoon declined abruptly very soon after fusion of the sperm head with the vitellus and remained at a very low level during the 3–6 h required for tail incorporation. Sperm motility, therefore, does not appear to be the main determinant in tail incorporation and the primary mechanism responsible for it remains unclear. As the sperm tail slowly entered the vitellus, the second meiotic division was completed with concomitant extrusion of the second polar body. Key stages in second polar body formation were correlated with events in tail incorporation. Differences between fertilization in vitro and in vivo are discussed.

Mammalian gamete interactions: What can be gained from observations on living eggs?

“…) Notwithstanding differences in its position at the start of incorporation,the sperm tail is usually incorporated into the vitellus in its entirety whether penetration has occurred in vitro or in vivo. (Some instances of incomplete incorporation have been noted during in vitro fertilization in the mouse and hamster [Gaddum-Rosse et al, 1982;Yang et al, 1972;Yanagimachi, 19811. ) Only two of the mammalian species studied so far are exceptions to this generalization: the Chinese hamster and the field vole (Austin and Walton, 1960;Pickworth et al, 1968;Yanagimachi et al, 1983).…”

“…Although mouse eggs undergoing in vitro fertilization do not display such prominent surface movements as rat eggs, there is nonetheless a n interesting change detectable by time-lapse cinematography: an increase in the rate of motion of granular components within the egg cytoplasm after fusion (Gaddum-Rosse et al, 1982). Furthermore, Waksmundzka et al (1984) have observed that the surface of fertilized mouse eggs changes from smooth to slightly undulate about 7-10 hr after activation.…”

“…The greater the length of tail that enters the space, the greater the chances that oocyte rotation will permit the remainder of the tail to enter. Finally, one should bear in mind that most of the observations on the mouse and rat described above (Sato and Blandau, 1979a;Gaddum-Rosse et al, 1982 were made on eggs penetrated in vitro. The extent to which the in vitro environment affects the final position of the sperm tail at the time of gamete fusion needs to be explored.…”

“…During in vitro fertilization of mouse eggs (Sato and Blandau, 1979a;Gaddum-Rosse et al, 1982) and rat eggs (Gaddum-Rosse et al., 1984), the major part of the flagellum is usually outside the zona pellucida at the time of sperm-egg fusion. As incorporation progresses, the proportion of the flagellum remaining outside the zona gradually decreases.…”

“…Where the tail is outside the zona at the time of gamete fusion, the process must be more complicated. Timelapse films of mouse eggs during tail incorporation suggested that the egg cortex rotated away from the site of entry of the flagellum so that, as each portion of the flagellum entered the vitellus, it was "received" into a new portion of the cortical cytoplasm (Gaddum-Rosse et al, 1982). The true mechanism of tail incorporation under these circumstances remains to be determined.…”