Developmental Rate of Rabbit Parthenogenetic Embryos Derived Using Different Activating Protocols

  • Chrenek, P. (Research Institute of Animal Production) ;
  • Makarevich, A. (Research Institute of Animal Production)
  • Received : 2003.07.31
  • Accepted : 2004.01.27
  • Published : 2004.05.01


The present study compares development of rabbit embryos generated using different oocyte activation protocols and reconstructed with embryonic or cumulus cells as nuclear donor. In vivo matured oocytes were collected from New Zealand White rabbits at 16 h after ovulation treatment and were activated at18 h of post-ovulation treatment. The following schemes of oocytes activation were tested: 1) single electric pulse (EP, 3.2 kV/cm, 3${\times}$20 $\mu$s, 0.3 M mannitol)+5 min culture in the presence of 5 mM Ionomycin, 2) single electric pulse (EP, 3.2 kV/cm, (${\times}$20 $\mu$s, 0.3 M mannitol)+1 h culture in the presence of 2 mM 6-DMAP, and 3) three electric pulses 30 min apart. Cleavage rate, percentage of expanded and hatched blastocysts as well as total cell number of blastomeres of parthenogenetic embryos were significantly higher using either EP+6-DMAP or 3${\times}$EP schemes, comparing with EP+Ionomycin. Development rate up to hatched blastocyst stage of cloned rabbit embryos using the EP+6-DMAP for activation of nuclei were 19% for embryonic cell nuclei and 36% for cumulus cell nuclei. The best activation protocol optimalized in this study was the combined treatment "P+6-DMAP" which may be potentially used for nuclear transfer protocol.


  1. Collas, P. and J. M. Robl. 1990. Factors affecting efficiency of nuclear transplantation in the rabbit embryo. Biol. Reprod. 43:877-884.
  2. Ju, J. Ch., J. S. Yang, Ch. T. Liu, Ch. H. Chen, J. K. Tseng, P. Ch. Chou and S. P. Cheng. 2003. Differential influences in sizes and cell cycle stage of donor blastomeres on the development of cloned rabbit embryos. Asian-Aust. J. Anim. Sci. 16:15-22.
  3. Ozil, J. P. 1990. The partenogenetic development of rabbit oocytes after repetetive pulsatile electrical stimulation. Development 109:117-127.
  4. Ozil, J. P. and D. Huneau. 2001. Activation of rabbit oocytes: the impact of the Ca signal regime on development. Development 128:917-928.
  5. Heyman, Y., P. Chesne and J. P. Renard. 1990. Full-term reprogramming of frozen embryonic nuclei after nuclear transfer in rabbit species. C. R. Acad. Sci. Paris. 311:321-326.
  6. Mitalipov, S. M., K. L. White, V. R. Farrar, J. Morrey and W. A. Reed. 1999. Development of nuclear transfer and parthenogenetic rabbit embryos activated with inositol 1,4,5-triphosphate. Biol. Reprod. 60:821-827.
  7. Yin, X. J., T. Tani, Y. Kato and Y. Tsunoda. 2000. Development of rabbit parthenogentic oocytes and nuclear transferred oocytes receiving cultured cumulus cells. Theriogenology 54:1469-1476.
  8. Wakayama, T. and R. Yamagimachi. 2001. Effect of cytokinesis inhibitors, DMSO and the timing of oocyte activation on mouse cloning using cumulus cell nuclei. Reproduction 122:49-60.
  9. Chesne, P., P. Adenot, C. Viglietta, M. Baratte, L. Boulanger and J. P. Renard. 2002. Cloned rabbits produced by nuclear transfer from adult somatic cells. Nature Biotechnology 20:366-369.
  10. Dinnyes, A., L. Liu, Y. Dai, J. Xu, P. Zhou, V. A. Velde and X. Yang. 1999. Development of cloned embryos from adult rabbit fibroblast cells. Theriogenology 51:199 (Aabstr.).
  11. Stice, S. L. and J. M. Robl. 1990. Activation of mammalian oocytes by a factor obtained from rabbit sperm. Mol. Reprod. Dev. 25(3):272-80.
  12. Chang, M. C. 1954. Development of parthenogenetic rabbit blastocyst induced by low temperature storage of unfertilized ova. J. Exp. Zool. 125:127-149.
  13. Escriba, M. J. and F. Garcia-Ximenez. 2000. Use of a variable electrical pulsing sequence in rabbit oocytes activation. Reprod. Nutr. Dev. 40:261-269.
  14. Inoue, K., N. Ogonuki, Y. Yamamoto, Y. Noguchi, S. Takeiri, K. Nakata, H. Miki, M. Kurome, H. Nagashima and A. Ogura. 2002. Improved postimplantation development of rabbit nuclear transfer embryos by activation with inositol 1,4,5-trisphosphate. Cloning Stem Cells 4(4):311-317.
  15. Liu, C. T., C. H. Chen, S. P. Cheng and J. C. Ju. 2002. Parthenogenesis of rabbit oocytes activated by different stimuli. Anim. Reprod. Sci. 15, 70 (3-4):267-276.
  16. Fissore, R. A. and J. M. Robl. 1992. Intracellular Ca response of rabbit oocytes to electrical stimulation. Mol. Reprod. Dev. 32:9-16.
  17. Ha, R. J., X. J. Yin, D. W. Kang, J. K. Park, S. Y. Song, I. K. Kong, H. J. Lee and C. S. Park. 1998. Parthenogenetic activation and development of rabbit oocytes following electric stimulation or ionomycin plus 6-dimethylaminopurine. Theriogenology 49:153 (Abstr.).
  18. Ju, J. C., T. H. Chen, J. K. Tseng, C. Tsay, S. P. Yeh, P. C. Chou, C. H. Chen and C. T. Liu. 2002. Cytoskeletal patterns, in vitro maturation and parthenogenetic development of rabbit GV oocytes. Asian-Aust. J. Anim. Sci. 15(12):1695-1701.