This study determined the individual and combined effects of extracellular adenosine triphosphate (ATP) and quercetin (QUE) on the quality of post-thawed sperm and the fertilization potential of Lohi rams. In experiment 1, semen samples from four Lohi rams were pooled and extended with different concentrations of ATP or QUE (control; no ATP or QUE, 1 or 2 mM ATP and 10 or 20 mu M QUE). In experiment 2, pooled semen samples were extended with various combinations of ATP and QUE (control; no ATP and QUE, 1 mM ATP + 10 mu M QUE, 1 mM ATP + 20 mu M QUE, 2 mM ATP + 10 mu M QUE and 2 mM ATP + 20 mu M QUE). All samples in both experiments were cryopreserved and analyzed for post-thawed sperm quality. In experiment 3, the best combination of ATP and QUE from experiment 2 was to extend semen, which was then used for laparoscopic insemination in estrussynchronized ewes (n = 83). The results of experiment 1 showed that 1 mM ATP and 20 mu M QUE treatments resulted in higher total motility, progressive motility, viability, plasma membrane intactness (PMI), and motion kinetics (VCL, VSL, VAP, LIN, and STR) compared to other treatments (p < 0.05). In experiment 2, the 1 mM ATP +10 mu M QUE-treated group exhibited significantly higher total and progressive motility, PMI, and motion kinetics (VSL, VCL, VAP, STR, and BCF) compared to the control group (p < 0.05). In experiment 3, the fertilizing potential of sperms treated with 1 mM ATP +10 mu M QUE was greater than that of untreated controls (58.1% vs. 27.5%, respectively, p-value = 0.012). In conclusion, the quality of post-thawed ram semen is enhanced when the extender is supplemented with extracellular 1 mM ATP and 20 mu M QUE, whether used separately or in combination with 1 mM ATP and 10 mu M QUE. Furthermore, the inclusion of 1 mM ATP and 10 mu M QUE together in the extender significantly improves in vivo fertility in Lohi ram.
