1. Pig eggs were examined with the ordinary and phase-contrast microscope in search for clues about the process of sperm penetration of the zona pellucida.
2. The following observations were made, (i) The zona pellucida consists of three concentric layers--the inner-zona, the middle-zona, and the outer-zona. (ii) The fertilizing spermatozoon makes a narrow slit in the zona by penetrating through it. The slit has a particular shape referred to as ‘the penetration curve’. (iii) Extra spermatozoa (i.e. spermatozoa other than the fertilizing spermatozoon) enter the zona and follow a course very similar to that of the fertilizing spermatozoon, but they can penetrate it only as far as the middle-zona. Extra spermatozoa have not been observed in the inner-zona, indicating that in this layer the ‘zona reaction’ is normally absolute, (iv) A filament extending from the apex of the sperm head has been observed in spermatozoa embedded in the zona. It is suggested that the sperm path through the zona is determined by this filament, which is therefore termed the sperm-penetration filament (SPF).
3. On the basis of these observations the following sequence of events in the passage of a spermatozoon through the zona is suggested. First, the spermatozoon attaches to the zona; it then ejects into the zona the SPF. The SPF, in turn, guides the spermatozoon on its curved course through the zona.