Freeze-fracture observations on mammalian oocytes

Freeze-fracture studies on mammalian oocytes have been hampered by the relatively small numbers of cells available at a given time as well as by difficulties encountered in effectively freezing these large, watery cells. We have nevertheless pursued this area because of the benefits of visualizing membrane faces involved in various fusion reactions by the freeze-fracture method. Our observations indicate no overall change in intramembranous particle (IMP) distribution before and after sperm penetration, although the question of possible alterations of these structures at the precise locus of sperm attachment remains open. Preliminary statistical analysis indicates that there is a much higher IMP density on the P face than on the E face of the plasma membrane and that the microvillar membranes bear more IMPs than those of the intermicrovillus regions. Probes of lipid subclasses were used to determine the distribution of cholesterol and anionic lipid in the egg plasma membrane. Filipin and tomatin showed extensive complex formation in microvillus as well as nonmicrovillus regions, whereas anionic lipids (using polymyxin B) have been difficult to detect on the oocyte surface. These results are discussed relative to current views of membrane fusion mechanisms.