Effects of glycerol concentration on the motility of equine spermatozoa after thawing

Abstract

The aim of this study was to evaluate the effect of different glycerol concentrations on stallion sperm motility after thawing. For statistical analysis 228 ejaculates were used. The semen was filtrated to remove gel fraction; macroscopic and microscopic evaluation was done. After evaluation the ejaculates were centrifuged, the supernatant was removed and the spermatozoa were re-suspended in French diluent with different concentrations of glycerol (2.0; 2.5; 4.0 and 6.0%). The choice of concentration of glycerol for a particular ejaculate was completely random. The spermatozoa were packed into 0.5 ml straws and placed for 2 h in a fridge (4 °C). Then the straws were placed in liquid nitrogen vapor (-80 to -100 °C) and after 10 min plunged into liquid nitrogen and stored at -196 °C for at least 48 h. The selected straws were individually thawed in a 38 °C water bath for 30 s prior to post-freezing analysis. Two progressive motilities using phase contrast microscopy (magnification × 400) were recorded: motility II immediately after thawing and motility III after 2 h incubation in a 38 °C water bath. The Spearmen/Kendall rang correlation test was selected to prove whether there is a correlation between the selected indices (glycerol concentration and motility II and motility III). Nonparametric multiple group analysis (Steel-Dwass test) was applied for finding the differences between groups. The Spearman/Kendall rang correlation proved a relationship between motility II and glycerol concentration. It can be stated that in this study the best glycerol concentration for freezing equine spermatozoa is with a concentration of 4.0% glycerol.

Effect of sperm and glycerol concentration on epididymal sperm motility parameters after thawing

“…Samples were evaluated at 60, 120 and 180 min of post-thaw incubation at different sperm and VG concentrations. This is quite different from other studies that evaluated motility indicators immediately after thawing (Macías García et al 2012;Neuhauser et al 2019) or 120 min after thawing (Olaciregui et al 2014;Mráčková et al 2017).…”

“…Because VG causes injury to spermatozoa during the cryopreservation process (Fahy et al 1990), its usage is usually limited to concentrations lower than 2.5% (Macías García et al 2012). Other studies have shown that concentrations higher than 2.5% may have better results (Mráčková et al 2017). Most of the studies were performed at relatively low sperm concentrations of 100-200 × 10 6 sperms per ml (Macías García et al 2012;Olaciregui et al 2014;Neuhauser et al 2019).…”

“…Most of the studies were performed at relatively low sperm concentrations of 100-200 × 10 6 sperms per ml (Macías García et al 2012;Olaciregui et al 2014;Neuhauser et al 2019). The previous study by Mráčková et al (2017) showed that the VG concentration and its harmfulness to sperm is closely related to the concentration of sperm used in the insemination dose for freezing. In the same study, the authors found that the best VG concentration for freezing equine epididymal sperm was 4%.…”

“…Insemination doses prepared with these concentrations consist of several straws (usually 5-10) which is impractical in terms of storage space and handling during mare insemination. The study of Mráčková et al (2017) showed that the concentration of VG and its harmfulness to sperm is closely related to the concentration of sperm used in the insemination dose for freezing, so it would be advantageous to use a higher concentration of sperm for freezing.…”