Henning Heiko, Nguyen Quynh Thu, Wallner Ulrike, Waberski Dagmar
Unit for Reproductive Medicine/Clinic for Pigs and Small Ruminants, University of Veterinary Medicine Hannover, Hannover, Germany.
Friedrich-Loeffler-Institut, Institute of Farm Animal Genetics, Neustadt am Rübenberge, Germany.
Front Vet Sci. 2022 Aug 5;9:953021. doi: 10.3389/fvets.2022.953021. eCollection 2022.
The optimum storage temperature for liquid-preserved boar semen has been empirically determined to be between 15 and 20°C. Lower temperatures provide an advantage to inhibit bacterial growth, but are regarded as critical due to the high sensitivity of boar spermatozoa to chilling injury. Higher storage temperatures are supposed to induce energy deficiency due to an insufficient depression of metabolic cell activity. However, experimental evidence for alterations of the sperm's energy status in relation to storage temperature and duration is missing. Therefore, we aimed to revisit the upper and lower storage temperature limits for liquid-preserved boar semen from the perspective of the sperm's energy metabolism. Ejaculates ( = 7 boars) were cooled down in Beltsville Thawing Solution (BTS) to 25, 17, 10, or 5°C and stored for up to 120 h. ATP and adenylate energy charge (EC) levels were assessed at storage temperature (24, 72, and 120 h storage) and after subsequent re-warming (38°C). Sperm quality and energy status remained at a stable level in samples stored at 25 and 17°C. Chilling to and storage at 10 or 5°C in BTS provoked cold shock in a subset of sperm as shown by a loss in viability and motility ( < 0.05), which was accompanied by a significant release of adenine nucleotides into the semen extender. Prolonged storage for 120 h resulted in significantly lower mean ATP concentrations in viable spermatozoa at 5 or 10°C compared to 17°C ( < 0.05). Cluster analysis revealed that the main sperm subpopulation, i.e., sperm with moderate speed and linearity, decreased from 50 to 30% ( < 0.05) in favor of slow-moving spermatozoa (5°C) or spermatozoa with a hyperactivation-like motility pattern (10°C). The results point to a sublethal imbalance in available ATP in a subset of the surviving sperm population, rather than a general decrease in available ATP in all spermatozoa. In conclusion, storing diluted boar semen at a stable temperature between 17 and 25°C is a safe procedure concerning the spermatozoa's energy status. Future concepts for hypothermic boar semen preservation below 17°C require measures which ameliorate the imbalanced energy status in viable spermatozoa.
通过经验确定，液态保存的公猪精液的最佳储存温度在15至20°C之间。较低的温度有利于抑制细菌生长，但由于公猪精子对冷休克损伤高度敏感，所以被视为关键因素。较高的储存温度可能会因代谢细胞活性抑制不足而导致能量缺乏。然而，关于精子能量状态随储存温度和时间变化的实验证据尚缺。因此，我们旨在从精子能量代谢的角度重新审视液态保存公猪精液的上下储存温度极限。采集射精样本（n = 7头公猪），在贝尔茨维尔解冻液（BTS）中冷却至25、17、10或5°C，并储存长达120小时。在储存温度下（储存24、72和120小时）以及随后复温（38°C）后评估ATP和腺苷酸能荷（EC）水平。储存在25和17°C的样本中，精子质量和能量状态保持稳定。在BTS中冷却至10或5°C并储存会使一部分精子受到冷休克，表现为活力和运动能力丧失（P < 0.05），同时伴随着腺嘌呤核苷酸大量释放到精液稀释液中。与17°C相比，在5或10°C下延长储存120小时导致存活精子中的平均ATP浓度显著降低（P < 0.05）。聚类分析显示，主要精子亚群，即具有中等速度和线性度的精子，从50%降至30%（P < 0.05），有利于慢速运动精子（5°C）或具有超活化样运动模式的精子（10°C）。结果表明，在存活精子群体的一部分中存在可用ATP的亚致死性失衡，而非所有精子中可用ATP普遍降低。总之，就精子的能量状态而言，将稀释的公猪精液储存在17至25°C的稳定温度下是安全的操作。未来低于17°C的公猪精液低温保存概念需要采取措施改善存活精子中失衡的能量状态。
