Effect Of Melatonin On Sperm Quality Of Heat Stressed Bull

As a heat stress source,high temperature has a serious impact on the sperm quality of bulls,which can lead to the decrease of ejaculation,the decrease of sperm motility and viability.Even lead to the production of frozen semen in summer is forced to stop for 2-3 months,which has become one of the important factors restricting the production of frozen semen in summer and the development of beef cattle industry,and is an important problem to be solved in the current animal husbandry production.Melatonin plays an important role in the reproductive process of male animals,such as spermatogenesis and sperm motility.However,the research and application of melatonin to protect the sperm quality under heat stress are few.Therefore,this experiment analyzes the influence of different seasons on semen quality of different breeds of bulls,taking Simmental as an example,.According to sperm activity,plasma membrane integrity,acrosome integrity,mitochondrial membrane integrity and other sperm quality indicators,the heat stress model of sperm in vitro was established to screen the best concentration of melatonin to protect the quality of heat stressed sperm.The possible receptor types of melatonin protecting sperm quality of bull under heat stress was also discussed.The main research contents and results are as follows:1.Semen quality of different bulls in different seasons.Three bulls of four breeds were selected from Henan Dingyuan breeding Co.,Ltd.for semen quality test in spring,summer,autumn and winter,and were investigated and counted the quality changes.The results showed that in summer,the ejaculation amount of different kinds of bulls decreased in different degrees,the rate of spermatogenesis and post freezing activity decreased significantly?P<0.05?,the sperm density decreased significantly?P<0.05?,and the rate of sperm deformity increased significantly?P<0.05?.2.Establishment of heat stress model of bull sperm in vitro.To investigate the changes of sperm acrosome integrity,plasma membrane integrity and mitochondrial membrane integrity in bulls cultured at different temperatures?35?,37?,39?,41?,43??for one hour.The sperm acrosome integrity was detected by hypotonic swelling test,inverted microscope,FITC/PNA fluorescent staining and PI/Rh123 fluorescent dye The integrity of mitochondrial membrane was detected by double staining.The results showed that the expression level of sperm quality indexes was the highest at 37?,and decreased significantly with the increase of temperature ?P<0.05?.3.To investigate the effect of different concentrations of melatonin on sperm quality under heat stress.Different concentrations of melatonin(0,10^-3mol/L,10^-4mol/L,10^-5mol/L,10^-6mol/L, 10^-7mol/L)were added to culture in vitro under heat stress for 1H.The effect of melatonin concentration on sperm quality under heat stress was detected and analyzed.The results showed that when the concentration of melatonin was 10^-33 mol/L,the sperm quality of bulls was significantly improved?P<0.05?.4.Melatonin protects the receptor type of heat stressed sperm quality.10^-77 mol/L lzindole?total melatonin inhibitor?and 10^-77 mol/L 4-p-pdot?melatonin receptor 2 antagonist?were used to detect the changes of sperm quality in each group.The results showed that the expression level of sperm quality indexes in luzinole group was significantly lower than that in melatonin group?P<0.05?,while that in 4-p-pdot group was not significantly different from that in melatonin group?P<0.05?.Compared with the melatonin treated group,the sperm quality indexes decreased significantly?P<0.05?.It was found that melatonin could protect the sperm quality of Simmental cattle under heat stress by activating MT1.