Fernández Silvina, Sestelo Adrián, Rivolta Miguel, Córdoba Mariana
1 Instituto de Investigación y Tecnología en Reproducción Animal, Química Biológica, Facultad de Ciencias Veterinarias, Universidad de Buenos Aires, Chorroarín 280 (1427) Capital Federal, Argentina.
Zoolog Sci. 2013 Dec;30(12):1110-6. doi: 10.2108/zsj.30.1110.
The aim of this research was to evaluate two different diluents for sperm cryopreservation and to study functional parameters in relation to the response to heparin, lysophosphatidylcholine and progesterone, in frozen-thawed semen of fallow deer (Dama dama) during the reproductive season (brama). In this way, fallow deer can be used as a biological model of endangered cervids. Semen was obtained by electroejaculation. Heparin, progesterone and lysophosphatidylcholine were used as capacitation and acrosome reaction inducers, respectively. Capacitation and acrosome reaction were evaluated by chlorotetracycline epifluorescence technique (CTC), membrane integrity by Hypo-osmotic swelling test (HOS) and viability and acrosome integrity by trypan blue stain/DIC. Data was analyzed by ANOVA and Tukey Test (P < 0.05). Semen was cryopreserved in different diluents and Fructose-Tris-Glycine extender was selected. Capacitation with heparin at different incubation times determined that the highest capacitation percentage was obtained at 45 minutes incubation. Progesterone (1 'M) and lysophosphatidylcholine in heparin capacitated sperm induced acrosome reaction (P < 0.05). This study contributes to improve cryopreservation methods and to increase the knowledge about capacitation and acrosome reaction in vitro in deer spermatozoa, allowing an advance in the development of reproductive biotechnologies.
本研究的目的是评估两种不同的精子冷冻保存稀释剂，并研究在繁殖季节（发情期）梅花鹿（黇鹿）冻融精液中，与对肝素、溶血磷脂酰胆碱和孕酮反应相关的功能参数。通过这种方式，梅花鹿可作为濒危鹿科动物的生物学模型。精液通过电射精获取。肝素、孕酮和溶血磷脂酰胆碱分别用作获能和顶体反应诱导剂。获能和顶体反应通过金霉素落射荧光技术（CTC）评估，膜完整性通过低渗肿胀试验（HOS）评估，活力和顶体完整性通过台盼蓝染色/微分干涉对比显微镜评估。数据通过方差分析和Tukey检验进行分析（P < 0.05）。精液在不同稀释剂中冷冻保存，并选择了果糖 - 三羟甲基氨基甲烷 - 甘氨酸稀释液。在不同孵育时间用肝素进行获能测定，结果显示在孵育45分钟时获得最高获能百分比。在肝素获能的精子中，孕酮（1μM）和溶血磷脂酰胆碱诱导顶体反应（P < 0.05）。本研究有助于改进冷冻保存方法，并增加对鹿精子体外获能和顶体反应的了解，从而推动生殖生物技术的发展。
