SEQUENTIAL RELEASE OF ACROSOMAL MEMBRANES AND ACROSOMAL ENZYMES OF RAM SPERMATOZOA

Abstract

Summary. A method involving the treatment of ram spermatozoa with MgCl2 followed by Hyamine 2389 and Triton X-100 is described, which gave selective removal of the acrosomal membranes and acrosomal enzymes. The effect on the individual membranes of ram spermatozoa was evaluated by electron microscopy. Treatment with MgCl2 removed or altered the integrity of the plasma and outer acrosomal membranes, allowing the release of material from the acrosome. The inner acrosomal membrane and the electron-dense material of the equatorial segment were frequently unaffected by this treatment. Subsequent treatment of these spermatozoa with Hyamine and Triton removed the inner acrosomal membrane and the electron-dense material from the equatorial segment. The MgCl2 extract contained acrosomal proteinases and hyaluronidase. The detergent extract contained sperm neuraminidase. The specific activities and yields of the enzymes obtained in the MgCl2 step were much higher than those of the enzymes obtained by the detergent treatment. Slight alterations in the conditions of the treatment had different effects on the acrosomal membranes in that one or more of the membranes were removed or altered in varying proportions resulting in the release of one or more enzymes. Only the plasma membrane and the acrosome appeared to be affected by these treatments as selected mitochondrial enzymes were not detectable in the extracts.