Assay of capacitated, freeze-damaged and extended stallion spermatozoa by filtration

Abstract

Results of stallion semen assay with glass wool/Sephadex filters are highly correlated with fertility. A hypothesis was tested that mechanically damaged spermatozoa are trapped in glass wool while capacitated spermatozoa are trapped in Sephadex. Another objective was to assay stallion semen in extenders known to vary in their ability to maintain sperm fertility with these filters, and for sperm motility. In Experiment 1, ejaculates were split so that a portion was used for fresh semen filtration and another portion plunged into liquid nitrogen repeatedly to maximize the number of broken acrosomes. The remainder of the ejaculate was inseminated into the uterus of an estrous mare which was flushed 6–8 hours later. Semen from each treatment was filtered through cotton with and without Sephadex, and through glass wool with and without Sephadex. Ejaculates were also washed and incubated in a capacitation medium. Aliquotes were filtered every 2 hours for 10 hours through glass wool and Sephadex. In Experiment 2, stallion semen diluted in egg yolk-lactose, skim milk-glucose and Triscitrate-yolk was evaluated for motility and with glass wool and Sephadex filters daily for 4 days. In Experiment 1, more frozen-thawed spermatozoa were trapped in glass wool than in Sephadex filters. Filters containing Sephadex trapped all uterine-incubated cells, while cotton did not affect the trapping of spermatozoa. In the capacitation medium, the passage of cells through all filters decreased with time, and glass wool and Sephadex had additive effects. In Experiment 2, fewer spermatozoa passed Sephadex filters than glass wool filters after storage in the Tris-citrate extender, which is known to be detrimental to the fertility of stallion semen although it did maintain sperm motility. We suggest that Sephadex traps spermatozoa with capacitation-like changes.