A high frequency of fertilization in premature and mature coelomic toad eggs after enzymic removal of vitelline membrane

The jelly-less coelomic eggs of the toad, Bufo bufo, even if chromosomally mature and reactive to parthenogenetic stimuli, are not fertilized or fertilized in a low frequency under the conditions which assure the sperm entry into dejellied uterine eggs. A high frequency of fertilization was obtained, however, when coelomic eggs from which the vitelline membrane had been digested away by pronase or hatching enzyme were inseminated in the presence of polyvinylpyrrolidone (PVP) or the diffusible egg-jelly component. Experimental evidence is presented to show that the conditions important for inducing this success in fertilization are the complete removal of the vitelline membrane and insemination in the presence of one of the following sperm ‘capacitating’ substances: the diffusible jelly component, the non-dialysable jelly component, PVP, dextran, or Ficoll. Fertilization obtained by this method was monospermic, as defined both by cytological examination and by the mode of early cleavage. However, the eggs invariably ceased development at the gastrula stage, because of the lack of the mechanical support normally provided by the egg envelopes. These results indicate that toad eggs acquire the full ability, without any influences from the oviduct, both to react to a penetrating sperm and to prevent polyspermy. When eggs undergoing maturation (premature eggs) were demembranated and inseminated in the presence of PVP, they did not show a genuine cleavage. Cytological examination proved the occurrence of highly polyspermic fertilization in these premature eggs, together with a tendency for the rate of pronucleus formation by the incorporated sperm to increase as the eggs were closer to the stage of full nuclear maturity. On the basis of these results, a possible role of the oviduct in affecting the vitelline membrane-egg surface interrelationship is discussed, as well as the changing reactivity of maturing oocytes to spermatozoa, and vice versa.