不同电激活参数及CB处理对猪卵母细胞孤雌发育的影响

从猪卵巢上获取的卵母细胞经体外培养成熟42～44 h后去除颗粒细胞,选取成熟的卵母细胞进行孤雌激活培养.试验采用不同的场强、脉冲时程和脉冲次数进行对照试验,并比较了细胞松弛素B(CB)辅助激活对孤雌胚胎发育的影响.结果显示:1)当场强为0.8 kV.cm-1,选取4个脉冲时程(80、100、120、140μs)参数分别进行1次、2次脉冲激活时,100μs、1次脉冲囊胚率最高,为(42.22±5.38)%,但同其他组无显著差异(P>0.05).2)脉冲时程为100μs,选取4个场强(0.8、1.0、1.2、1.4 kV.cm-1)参数分别进行1次、2次脉冲激活时,1.4 kV.cm-1、1次脉冲囊胚率最高,为(44.04±0.68)%,但同其他组无显著差异(P>0.05).3)场强为0.8 kV.cm-1和1.4 kV.cm-1,选取4个脉冲时程(80、100、120、140μs)进行1次脉冲激活时,1.4 kV.cm-1+80μs组囊胚率(54.60±2.86)%均高于其他组,且极显著高于1.4 kV.cm-1+140μs、0.8 kV.cm-1+80μs和0.8 kV.cm-1+140μs组(P<0.01).4)用1.4 kV.cm-1、80μs、1次脉冲激活卵母细胞后,使用CB辅助激活4 h的囊胚率为(54.07±3.12)%,极显著(P<0.01)高于不使用CB组.以上结果说明,电激活中脉冲次数对猪孤雌胚胎发育无显著影响,且在本试验条件下,采用1.4 kV.cm-1的场强、80μs的脉冲时程,并使用CB辅助激活4 h,能够得到较高囊胚率的猪孤雌激活效率.

Effects of Different Electrical Stimulation Parameters and Cytochalasin B Treatment on Parthenogenetical Development of Porcine Oocytes

Oocytes were collected from porcine ovaries,cultured in vitro for 42-44 h,and cumulus cells were removed from oocytes.Matured oocytes were selected for parthenogenetical activation.The effect of different field strengths(FSs),pulse durations(PDs),electrical pulse numbers(EPNs),and the influence of cytochalasin B treatment on development of parthenogenetically activated porcine oocytes were investigated.In the first experiment,4 different PDs(80,100,120,140 μs) were applied with a single or two 0.8 kV·cm-1 pulse(s) to activate the oocytes.The results showed that a single 100 μs pulse of 0.8 kV/cm resulted in the highest blastocyst rate of(42.22±5.38)%,but it was not statistically different from the blastocyst rates of other groups(P>0.05).In the second experiment,4 different FSs(0.8,1.0,1.2,1.4 kV·cm-1) were used with a single or two 100 μs pulse(s) to stimulate the oocytes.The highest blastocyst rate [(44.04±0.68)%] was observed in the group applied with single 100 μs pulse of 1.4 kV·cm-1,but it was not statistically different from the blastocyst rates of other groups(P>0.05).In the third experiment,4 different PDs(80,100,120,140 μs) were used with a single 0.8 kV·cm-1 or 1.4 kV·cm-1 pulses to activate the oocytes.The blastocyst rate of the group treated with a single 80 μs pulse of 1.4 kV·cm-1 reached(54.60±2.86)%,which was significantly higher than that of other groups(P<0.01).In the fourth experiment,one group of oocytes was stimulated only by a single 80 μs pulses of 1.4 kV·cm-1,while the other groups was activated with a single 80 μs pulses of 1.4 kV·cm-1 followed by 4 h-cytochalasin B treatment.The results indicated that the blastocyst rate of the latter group was(54.07±3.12)%,significantly higher than that of the former group(P<0.01).The above results suggested that EPN had no significant effect on parthenogenetical development of porcine oocytes,and 4 h-cytochalasin B treatment to the oocytes after the electrical stimulation by a single 80 μs pulses of 1.4 kV·cm-1 resulted in a high parthenogenetical blastocyst rate.