POSTTHAW PLASMA-MEMBRANE INTEGRITY OF BULL SPERMATOZOA SEPARATED WITHA SEPHADEX ION-EXCHANGE COLUMN

Previous experiments have established that filtration of bovine semen through a Sephadex ion-exchange column improves its quality before and after freezing. The present study was conducted to determine the post -thaw membrane integrity of bull spermatozoa separated with a Sephadex ion-exchange column and to determine the kind of protection to sperma tozoa is provided by glycerol during freezing and thawing. Semen from Holstein bulls diluted in TEST-yolk extender (with and without glycero l) was filtered through a Sephadex ion-exchange column and frozen in l iquid nitrogen (-196 degrees C). After thawing, there were more normal acrosomes in filtered spermatozoa than nonfiltered (P<0.01). Post-tha w plasma membrane integrity and swelling ability in a hypoosmotic solu tion revealed that the filtered spermatozoa had a stronger (P<0.005) p lasma membranes than the nonfiltered. Filtered spermatozoa demonstrate d higher zona-free hamster oocyte penetration than the nonfiltered (30 .5 vs 11.5%; P<0.0005). Spermatozoa extended in TEST-yolk without glyc erol had the lowest (P<0.001) normal acrosomes, intact plasma membrane s and swelling ability. Plasma membrane over the post-acrosomal region of the head and post-midpiece region of the tail was more sensitive t o damages caused by freezing and thawing than acrosomal and midpiece r egions of spermatozoa. Glycerol in the extender provided significant ( P<0.05) protection to the sensitive regions of filtered and nonfiltere d spermatozoa during freezing and thawing. Filtered plus glycerolated spermatozoa had the highest (P<0.01) normal acrosomes, intact plasma m embranes and swelling ability. In conclusion, the pre-freezing filtrat ion of bovine semen harvested the spermatozoa possessing stronger plas ma membranes which enabled them to endure freezing and thawing stresse s. The addition of glycerol to the extender protected the post-acrosom al region of the head and post-midpiece region of the tail of spermato zoa from freezing and thawing shocks. (C) 1997 by Elsevier Science Inc .