Inhibition of Interaction between Perivitelline Membrane and Sperm by Anti-Quail ZPC Antiserum

Abstract

The extracellular investment of oocytes in a number of species contains specific adhering molecule for spermatozoa ("sperm receptor") to which spermatozoa bind as a prelude to fertilization. However, little is known about the nature of sperm receptors in avian species. In quail, the oocyte at the time of fertilization is bounded by the perivitelline membrane, which is a three dimensional network of fibers consisted of two glycoproteins with molecular masses of 33kDa and 175kDa. In order to elucidate the sperm receptor in the perivitelline membrane of quail oocyte, we examined the effects of antiserum against 33kDa protein, homologous protein of mouse sperm receptor, ZPC, on the interaction of spermatozoa to the perivitelline membrane.
Ejaculated semen was obtained from male quail during natural mating with female. A piece of the perivitelline membrane of the largest follicle was incubated for 40min at 39C with 2×10⁶ spermatozoa/ml with or without 100μg/ml soybean trypsin inhibitor. The membrane was stained with Schiff's reagent for counting the number of perforation. Adherent spermatozoa were stained with 4', 6-diamidino-2-phenylindole and counted under a fluorescent microscopy. Sperm-induced perfora-tion was observed by the incubation of the perivitelline membrane with spermatozoa, and the binding of spermatozoa was observed by the incubation in the presence of trypsin inhibitor. Sperm binding, as well as perforation of the perivitelline membrane by spermatozoa, was inhibited by the addition of the antiserum against quail ZPC. These studies suggest the importance of quail ZPC for the sperm adhesion to the perivitelline membrane during fertilization.