Abstract
Polymyxin B (PMB) is beneficial for boar semen storage since it neutralizes the endotoxin of bacteria. However, the direct effect of PMB on boar spermatozoa has been unknown. This study aimed to examine the effect of PMB on acrosomal exocytosis, an essential process for successful fertilization in boar spermatozoa. Ejaculated spermatozoa stored with BTS extender at 17°C were washed and incubated with 0–100 μM PMB for 20 min and then examined for % total motililty, vigor grade and viability. None of the parameters was significantly different between 0 and 50 μM PMB with a gradual decline at higher concentrations. Thus the effect on acrosomal exocytosis was investigated at 0–50 μM of PMB. Spermatozoa were preincubated with PMB for 10 min, incubated for stimulation of acrosomal exocytosis with Ca2+ and the calcium ionophore A23187 and then fixed with glutaraldehyde at 5, 10 and 15 min. Preincubation with PMB at 0.01–50 μM and 0.05–50 μM resulted in significant enhancement of acrosomal exocytosis at 10 min and 15 min of incubation, respectively. Preincubation with PMB followed by incubation without A23187 did not affect acrosomal exocytosis. These results suggest that PMB exerts effects on the acrosomal exocytosis triggered by Ca2+ and A23187 in boar spermatozoa.
Highlights
During the last decades, the use of artificial insemination (AI) in commercial pig herds by means of fresh diluted semen has increased considerably (Dominiek, Alfonso, Tom, Philip, & Ann, 2011)
This study investigated the effect of Polymyxin B (PMB) on acrosomal exocytosis triggered by calcium and the calcium ionophore A23187 in boar spermatozoa that were freed from seminal plasma, extender, and from LPS
The results showed that PMB enhanced acrosomal exocytosis triggered by Ca2+ and A23187 at a concentration that did not affect sperm motility and viability suggesting that PMB can accelerate boar spermatozoa to undergo acrosomal exocytosis
Summary
The use of artificial insemination (AI) in commercial pig herds by means of fresh diluted semen has increased considerably (Dominiek, Alfonso, Tom, Philip, & Ann, 2011). Okazaki et al (2010) reported that addition of PMB to stored boar spermatozoa is beneficial to attenuate decrease in motility. While PMB is beneficial for boar semen storage via antibacterial action, the direct effect of PMB, not via the neutralizing effect of PMB on bacteria, is totally unknown. Because acrosomal exocytosis is an essential step to fertilization, information on the direct effect is of significance to employ PMB for boar semen storage. This study investigated the effect of PMB on acrosomal exocytosis triggered by calcium and the calcium ionophore A23187 in boar spermatozoa that were freed from seminal plasma, extender, and from LPS. Since the direct effect of PMB on boar spermatozoa is totally unknown, its effect on motility and viability was first examined. The PMB concentrations found to be without effect on these traits were used to investigate the effect of PMB on acrosomal exocytosis in order to avoid false assessment by degenerative acrosome loss
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