Characterization of viability, mitochondrial activity, acrosomal integrity and capacitation status in boar sperm during in vitro storage at different ambient temperatures

Abstract

Extended storage of unfrozen boar semen becomes an alternative because the use of frozen–thawed boar sperm results in low fertility. Sperm viability, mitochondrial activity, capacitation and acrosome integrity of freshly ejaculated boar semen stored in vitro for up to 48 h at 4°C, 15°C, 20°C and 39°C was characterized during the study. The viability of boar sperm was assessed by both Hoechst 33258 and SYBR-14/PI staining. Mitochondrial function was assessed by JC-1 staining. Capacitation status was determined by chlortetracycline (CTC)/Hoechst 33258 staining. The acrosome integrity was analysed with Coomassie blue staining. These data were derived from three ejaculates each from three crossbred boars. The viabilities assessed with SYBR-14/PI, Hoechst 33258 and JC-1 staining correlated highly (r > 0.980). In freshly ejaculated boar semen, 96 ± 1% of the sperm did not take up the Hoechst 33258, whereas 95 ± 2% were stained by SYBR-14 and 96 ± 2% of the sperm had mitochondria exhibiting positive JC-1 staining. Staining with CTC/Hoechst 33258 suggested that a high percentage of sperm became capacitated after 24 h storage at 15°C and 20°C. There were 62 ± 2% (15°C) and 89 ± 2% (20°C) capacitated sperm by 48 h. Moreover, most of the capacitated sperm were acrosome intact. These results suggest that SYBR-14/PI, Hoechst 33258 or JC-1 staining can be used to effectively evaluate the quality of boar sperm during in vitro storage.