Abstract
Changes in sperm structure and function occur during the processing of semen. The present study was designed to investigate the effect on buck sperm during different stages of semen preparation including dilution, cooling, equilibration and freeze-thawing. Semen ejaculates from three mature bucks (replicates = 5) were diluted with tris-citric acid egg yolk glycerol extender at 37 ºC, cooled to 4 ºC over 90 min, equilibrated at 4 ºC for 2 h, transferred to 0.5 mL straws, placed in nitrogen vapour, frozen and thawed and then analysed. Sperm samples were assessed for percentage motility, acrosomal and plasma membrane integrity, live sperm, and morphology after dilution, cooling, equilibration and thawing. Mean percentage motility after dilution (86.0 ± 1.4%) was reduced significantly (p < 0.05) due to cooling and equilibration (77.6 ± 1.3% and 74.6 ± 1.4% respectively); furthermore, it decreased significantly (p < 0.05) after freezing and thawing (42.3 ± 2.5%). Mean percentage of live sperm was higher (p < 0.05) after dilution (89.3 ± 1.4%)compared with cooling (84.8 ± 1.8%) and equilibration (80.2 ± 2.5%) and further reduced (p < 0.05) after freezing and thawing (56.0 ± 3.4%). Sperm morphology dropped significantly (p < 0.05) from 96.4 ± 0.3% after dilution to 88.8 ± 1.3% at cooling and further decreased (p < 0.05) after freezing and thawing (81 ± 1.9%). Mean percentage of sperm with normal plasma membrane after dilution (82.2 ± 1.1%) was significantly reduced (p < 0.05) at cooling or equilibration (73.8 ± 1.8) and further decreased (p < 0.05) after freezing and thawing (50.1 ± 2.9%). The percentage of sperm with normal acrosomes did not differ significantly due to dilution, cooling or equilibration (85.8 ± 1.7%, 83.2 ± 1.6%, 81.7 ± 1.8%) but was significantly reduced after freezing and thawing (45.2 ± 2.8%). In conclusion, frozen thawed sperm showed maximum damage to motility, morphology, plasma membrane and acrosome integrity following cooling.
Highlights
About 90% of goats are found in Asian countries, including China, India, Pakistan and Bangladesh (Iqbal et al 2008)
Some of the key semen assays related to functional significance and fertility include hypo-osmotic swelling as a measure of plasma membrane integrity (Jeyendran et al 1984), the presence of a normal acrosomal ridge (Gillan, Evans & Maxwell 2005), and sperm morphology, which has been considered as a robust clinical test (Blom 1973)
The objective of the present study was to assess the damage to sperm motility, plasma membrane integrity (PMI), and the normal apical ridge (NAR) of the acrosome, live-dead and morphology of buck sperm after dilution, cooling to 4 oC, equilibration at 4 oC and thawing after freezing
Summary
About 90% of goats are found in Asian countries, including China, India, Pakistan and Bangladesh (Iqbal et al 2008). Goats are embedded in the culture and are socially accepted to alleviate poverty, in developing countries. Against this background, research has been undertaken on reproductive biotechnology including artificial insemination using fresh or frozen semen. The greatest obstacle to the exploitation of frozen semen is that the freeze-thawing process of mammalian sperm generally leads to a decrease in motility and viability of sperm cells as a result of damage to membrane integrity and ultrastructure (Watson 2000). It has been demonstrated that cryopreservation leads to a decrease in sperm motility in the goat (Dorado et al 2009). Some of the key semen assays related to functional significance and fertility include hypo-osmotic swelling as a measure of plasma membrane integrity (Jeyendran et al 1984), the presence of a normal acrosomal ridge (Gillan, Evans & Maxwell 2005), and sperm morphology, which has been considered as a robust clinical test (Blom 1973)
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