Effect of melatonin supplementation to the maturation media on in vitro cumulus cells expansion and nuclear maturation rates of buffalo oocytes

The aim of this study is to determine the effect of the melatonin addition to the maturation medium (MM) on in vitro maturation of buffalo oocytes (IVM). Maturation is evaluated by the cumulus cells expansion and nuclear maturation. The cumulus oocyte complexs (COCs) were harvested by aspiration of antral follicles (2-8 mm diameter) of slaughter buffalo ovaries, COCs with homogeneous cytoplasm and at least three layers of cumulus cells were selected. The COCs were incubated for 24h in TCM–199 medium with melatonin (10, 30 or 50ng/ml) or without addition (control). After 24 h in culture media, the maturation rate of oocytes was determined by evaluating the expansion degree of the cumulus-oocyte complex (COCs) in each treatment, then classified into: fully expanded, partially expanded or not expanded, and byevaluation stages of oocyte nuclear maturation (GV, GVBD,  MI, MII or degenerated). Results illustrate that supplementation maturation media with 10 and 30, 50 ng/ml melatonin resulted in increasing the cumulus cell expansion of buffaloes oocytes by  86.4, 82.3, 83.2 %, respectively, as compared with oocytes cultured with melatonin - free medium (73.3%), The respective differences were significant. Majority of this improvement in the expansion rates of oocytes was observed in the rate of fully expanded oocytes, On the contrary, the rate of immature (unexpanded) oocytes incubated in melatonin-free medium was significantly (P<0.01) higher (26.7%) than those incubated in maturation media with 10 or 30 or 50 ng/ml melatonin (13.8, 17.7, 16.9 %, respectively). Moreover, results indicated that addition of melatonin to the maturation medium significantly improved the percentage of oocytes of first polar body (matured to MII) as compared with those incubated without melatonin, the highest percentage of those oocytes was obtained with 10 ng/ml melatonin (58.7 %), however, the least percentage was in those oocytes cultured with melatonin - free medium (48.6%) with significant differences (P<0.05). On the other side, the percentage of oocytes of GVBD, MI or degenerated stages did not significantly affected with melatonin addition (10. 30 or 50 ng/ml) to culture media as compared with those cultured   without melatonin medium. It could be concluded that addition of melatonin to the MM improved cumulus expansion and nuclear maturation rate of buffalo oocytes.