Study On Cryopreservation Of Boar Semen

To obtain better boar frozen-thawed semen for artificial insemination, in this study, two-way cross experiment was designed to investigate the effect of pretreatment in room-temperature extenders on cryopreservation of boar semen and the effect of glycerol equilibration time on six room-temperature extenders and three kinds of cryoprotective basic media according to the motility of sperm frozen-thawed. The effects of three disaccharides (trehalose, sucrose, milk sugar) added in the diluents and low-density lipoprotein (LDL) extracted from egg yolk replacing egg yolk completely on the quality of sperm frozen-thawed were studied intently considering the motility, the viability of Propidium Iodide (PI) staining, the mitochondrial activity of Rhodamine 123 (R123) staining, the tail-coiling spermatozoa percentage of hypo-osmotic swelling test (HOST), the acrosome integrity of Fluorescein isothiocyanate-labeled peanut agglutinin (FITC-PNA) staining and Chlortetracycline (CTC) staining respective, the percentage of capacitated spermatozoa before capacitating, the potential of capacitation as parameters for evaluating. The results were summarized as follows: 1. The effects of cryoprotective basic media on the motility of sperm frozen-thawed were not significant in two protocols of glycerol equilibration time (P>0.05, n=18); However, the effects of pretreatment through room-temperature extenders were so marked (P<0.01, n=9) and the cooperation effects between room-temperature extenders and basic media were exhibited distinctly (P<0.01, n=54). It was proposed that the seminal paste and media'ingredient and the co-action among them would be responsible for the results mentioned above. 2. Glycerol equilibration time affected room-temperature extenders markedly (P<0.05, n=9) but not basic media, especially BTS, Zorlesco, Schonow and Glucose-Caffeine-Sodium Pyruvate extenders. The results showed the fitting glycerol equilibration time was 2-3h for BTS but 5-10min for the three extenders latter. 3. The motility and viability of sperm frozen-thawed were affected markedly and the difference between treatments was so significant (P<0.01, n=39); The motility and viability were higher after freezing-thawing in trehalose and sucrose than in milk sugar (P<0.05, n=39) since the motility could be maintained at 0.4 in 25% prime concentration, although the motility was decreased drastically by high concentration of disaccharide (50% and 100%). 4. In terms of acrosome integrity significant difference was found between treatments (P<0.01, n=39). The sperm samples frozen in trehalose obtained the highest intact acrosome percentages (the value respective was 63.41%,64.41%,65.08%,66.47% in four treatments of trehalose), which were significantly different from those of samples frozen in sucrose, milk sugar and TCG solutions. 5. Trehalose could decrease distinctly the percentage of spermatozoa capacitated caused by freezing and thawing. However, sucrose and milk sugar seemed to be not, even the percentage was increased at 75% and 100% concentration in milk sugar solution. 6. Capacitation potential was found to improve significantly with the addition of three disaccharides in the diluents (P<0.05, n=39), especially at 25% concentration. 7. The motility, viability, intact acrosome spermatozoa percentage and capacitation potential of sperm frozen-thawed were all found to improve markedly in LDL by replacing egg yolk completely, which were higher than in TCG control group. And there were very significant differences between treatments (P<0.01, n=27). 8. In terms of motility and viability of frozen-thawed sperm the results were even more than those of the best three disaccharide groups obtained from LDL group at 8% and 9% concentration and the prime concentration of LDL was 9%. 9. The percentage of spermatozoa capacitated in freezing-thawing process was observed to decrease distinctly in LDL added in the diluents (P<0.01, n=27).