Ultrastructural study of the ability of seminal plasma proteins to protect ram spermatozoa against cold-shock.

The process of sperm cryopreservation, involving cooling, freezing, and thawing, induces serious detrimental changes in sperm function. The plasma and acrosomal membranes of spermatozoa are considered to be the primary site of these modifications due to thermal, mechanical, chemical, and osmotic stress. In previous studies, we demonstrated the ability of seminal plasma (SP) proteins to protect ram spermatozoa against cold-shock by using biochemical markers and scanning electron microscopy. In this study, we have attempted to examine the potential protective effect of SP proteins in membrane ultrastructure of ram spermatozoa subjected to cold-shock, by means of transmission electron microscopy (TEM). All the experiments were carried out with fresh spermatozoa freed from SP by a dextran/swim-up procedure. The high proportion of viable spermatozoa found in the swim-up obtained sample decreased drastically after the cold-shock treatment, and a considerable blebbing and vesiculation of the plasma and acrosomal membranes was found. The addition of SP proteins increased the sperm resistance to damage due to cold-shock (48% membrane-intact spermatozoa versus 15% in the control sample), and TEM analysis revealed that membrane alteration was prevented. This protective effect seems to be specific for SP proteins, as the addition of BSA did not provide any protection.