A factor potentiating serotonin in the induction of germinal vesicle breakdown in surf clam oocytes

Summary Simultaneous addition of an aliquot of body fluid obtained from the surf clam,Spisula solidissina, enhanced oocyte germinal vesicle breakdown induced with serotonin but not with KCl. When the body fluid and serotonin were added sequentially to the oocytes, potentiation did not occur. Body fluids of both males and females were effective at a 200-fold dilution. The factor is stable when treated with heat, acid, base, trypsin and pronase. It is hydrophobic and not dialyzable through tubing with a molecular weight cutoff of 1000 daltons. The factor is probably not a protein.17 November 1986Acknowledgments. This work was supported by the U.S.-Japan Cooperative Science Program, NSF INT-8211350, and Rockefeller Foundation Grant GAPS 8506. The authors thank Drs H. Shirai, T. Kishimoto, K. Sano, E. Sato and H. Ueno for valuable discussion.     

基于USB的卵母细胞电压钳同步数据采集和分析软件设计

介绍了基于USB接口的卵母细胞电压钳系统的数据采集分析系统,应用Delphi语言开发了系统的人机操作实验界面,探讨了电压钳的USB同步传输、数据采集和和显示的实现方法.在细胞模型测试中,电压钳放大器的跨膜电流分辨达到了0.01nA,噪声测试低于10pA.在非洲爪蟾卵母细胞表达Kv4.2钾通道实验中,记录跨膜电流波形清晰,数据分析可靠.与国外同类仪器相比:该系统操作简单,价格低,数据采集和分析方便,适用于卵母细胞表达实验.     

Effects of different nuclear transfer and activation methods on the development of mouse somatic cell cloned embryos

A group of adult somatic cell cloned mice were obtained by using cumulus cells as nuclei donor cells. To study the effect of different nuclear transfer (NT) and activation methods on the development of mouse cloned embryos, embryos were reconstructed using two traditional NT methods (electrofusion and direct injection) and four activation treatments (electric pulse, ethanol, SrCl2 and electric pulse combined with SrCl2). The data showed that the efficiency of reconstruction using the direct injection method is significantly higher (90.7%) than that of the electrofusion method (49.7%). Parthenogenetic embryos can develop to blastocyst stage with three activation conditions, including ethanol, electric pulse and SrCl2; however, the rates of development to blastocyst after ethanol and electric pulse acti-vation (52.4%, 54.2%) are significantly lower than after SrCl2 activation (76.9%). Treatment of embryos for 6 h with 10 mmol/L SrCl2 was found to be the best condition for activation of parthenogenetic as well as reconstructed embryos. By contrast, reconstructed embryos failed to develop to blastocyst stage after being activated by ethanol. The use of either injection or electrofusion for embryo reconstruction affected the pre-implantation development. However, after transfer in pseudopregnant mice, cloned mice were obtained from both methods.     

卵母细胞老化对小鼠体细胞克隆胚发育的影响

就小鼠卵母细胞的卵龄对克隆胚的体外发育能力的影响进行了检测,以确定最佳的取卵时间,以及取卵后进行核移植操作的可耐受的时间。采用PMSG和hCG超排B6D2F1雌鼠。在体内老化实验中,分别于注射hCG后13,15,17,20h取卵用于核移植操作;在体外老化实验中,所有的卵均于注射hCG后13h取出并培养,然后在13,15,17,20h进行核移植操作。每个时间点的核移植操作在1h内完成,重构的胚胎培养1h后进行激活。结果显示:在体内老化实验中,注射hCG后13h取卵,获得的克隆囊胚发育率最高,为56.0%,15h取卵仍维持了其支持胚胎体外发育的能力,但17h及更长时间后取的卵,重构后的囊胚发育率显著下降(18.1%)。注射hCG后15h取的卵孤雌发育率最高(囊胚发育率为90.1%),并在17h仍维持较高的孤雌发育能力,在20h显著下降。在体外老化试验中,于注射hCG后13h取卵,分别于13,15,17h重构,都获得了较高的囊胚发育率(分别为57.7%,52.2%,46.3%),而在注射hCG后20h重构,囊胚发育率显著下降(14.8%)。体外老化的孤雌胚在注射hCG后22h激活时囊胚发育率显著下降。结果表明:卵母细胞在体内和体外老化都影响克隆胚的体外发育,最佳取卵时间为注射hCG后13h,取卵后立即用于重构可获得最佳囊胚发育率;体内、外老化卵支持重构胚较好发育的时间间隔是不同的,体内老化卵支持重构胚发育能力从注射hCG后17h开始下降,体外老化卵则发生在20h。这些结果可以指导操作,有利于在应用核移植进行其他基础研究时降低操作引起的误差。