Beneficial effects of melatonin on canine oocyte nuclear maturation via reduction of oxidative stress

Unlike other domesticated animals, in vitro maturation of canine oocytes results in poor nuclear maturation to the metaphase II stage and high oocyte degeneration. The high-fat content of canine oocytes is the likely cause; it predisposes them to oxidative stress and deleterious reactive oxygen species (ROS). Melatonin (MTN) (potentially acting as a powerful antioxidant) was reported to support in vitro cultured oocytes in other species. In this work, canine cumulus oocyte complexes (COCs) were collected after routine ovariohysterectomy. Immunocytochemistry for expression of melatonin receptors (MTNR-1A and 1B) revealed that both receptors were highly expressed in canine oocytes and there was lower expression in the cumulus cells. Canine COCs matured in vitro in melatonin-supplemented culture at 100 nM concentration in low (5%) O2 incubator had a lower percentage at GV stage (6.7% ± 4.2 vs 19.8% ± 3), a higher MII stage (32.3% ± 6.4 vs 15.81% ± 8.1), lower degeneration (20.5% ± 3.2 vs 45.2% ± 5.15) and higher meiotic resumption (GVBD-MII; 56.2% ± 8.6 vs 19% ± 3), and produced lower ROS (determined after DCHFDA staining) (all P < 0.005) than oocytes cultured in high O2 (20%) incubator. The expression of ROS-regulating genes GPX-1 and catalase was significantly reduced in oocytes cultured with melatonin in low O2 compared with high O2 (P < 0.05). Importantly, the oocyte maturation rate increased significantly when G-IVF PLUS, a commercial culture medium used in human oocyte culture, was supplemented with MTN (P < 0.05). These results support the main objective of this study to analyze the beneficial effects of melatonin and low oxygen tension on both health and the developmental competence of in vitro matured canine oocytes.