Abstract
This study evaluated the effect of melatonin supplementation of in vitro maturation media on in vitro maturation (IVM) and in vitro fertilization (IVF) rate of buffalo oocytes. Cumulus oocytes complexes (COCs) were aspirated from follicles of 2-8 mm diameter. In experiment I, COCs were matured in IVM medium supplemented with 0 (control), 250, 500, and 1000 μM melatonin for 22-24 hours in CO2 incubator at 38.5°C with 5% CO2 and at 95% relative humidity. The maturation rate did not differ in media supplemented with melatonin at 250 μM, 500 μM, 1000 μM and control (0 μM). In experiment II, the matured oocytes were fertilized in 50 μl droplets of Tyrode’s Albumin Lactate Pyruvate (TALP) medium having 10 ug/ml heparin for sperm (2 million/ml) capacitation. The fertilization droplets were then kept for incubation at 5% CO2, 39°C and at 95% relative humidity for 18 hours. The fertilization rate was assessed by sperm penetration and pronuclear formation. Fertilization rate was improved when maturation medium was supplemented with 250 μM melatonin compared to control. In conclusion, melatonin supplementation to serum free maturation media at 250 μM improved the fertilization rate of buffalo oocytes.
Highlights
Buffalo is important for ecologically disadvantaged agricultural systems as it provides meat, milk and working power but is essential as a livestock source
The oocytes are more exposed to light, air and chemicals during in vitro handling that is responsible for generation of reactive oxygen species (ROS)
The oocyte developmental competence was reported to be improved by increasing the antioxidant capacity of oocytes during in vitro maturation (IVM), as the high lipid content makes buffalo oocytes/embryos sensitive to oxidative damages (Boni et al, 1992)
Summary
Buffalo is important for ecologically disadvantaged agricultural systems as it provides meat, milk and working power but is essential as a livestock source. Due to low maintenance requirements and good ability of feed conversion, buffaloes are considered ideal for low input systems and for the low cost production systems (Zicarelli, 1994). In spite of these qualities, the production potential of our dairy buffalo is low compared to dairy cattle in developed countries. Free radicalscavenging antioxidants exist within the follicular and oviductal fluid that is able to protect the oocytes against oxidative stress (Wang et al, 2002), this system becomes insufficient under in vitro conditions. It has been reported that melatonin directly protects the oocytes of human and mouse from oxidative stress (Tamura et al, 2008). Present study was designed to assess the role of melatonin in in vitro maturation media on in vitro maturation (IVM) and in vitro fertilization (IVF) of buffalo oocytes
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