Effect of elevated Ca(2+) concentration in fusion/activation medium on the fusion and development of porcine fetal fibroblast nuclear transfer embryos

Mol Reprod Dev. 2002 Apr;61(4):488-92. doi: 10.1002/mrd.10110.

Abstract

The present study examined the effect of elevated Ca(2+) concentration in fusion/activation medium on the fusion and development of fetal fibroblast nuclear transfer (NT) porcine embryos. Frozen-thawed and serum starved fetal fibroblasts were transferred into the perivitelline space of enucleated oocytes. Cell fusion and activation were induced simultaneously with electric pulses in 0.3 M mannitol-based medium containing 0.1 or 1.0 mM CaCl(2). Some fused embryos were further activated 1 hr after the fusion treatment by exposure to an electric pulse. The NT embryos were cultured in vitro for 6 days. Fusion and blastocyst formation rates were significantly (P<0.05) increased by increasing the Ca(2+) concentration from 0.1 mM (67.1 and 6.3%) to 1.0 mM (84.7 and 15.8%). However, no difference in the number of cells in blastocysts was observed between the two groups. A higher percentage of blastocyst was also observed when control oocytes were parthenogenetically activated in the presence of elevated Ca(2+) (19.3% vs. 32.4%, P<0.05). When the reconstituted oocytes were fused in the medium containing 1.0 mM CaCl(2), increasing the number of pulses from 2 to 3 or an additional activation treatment did not enhance the blastocyst formation rate or cell number in blastocysts. These results demonstrate that increasing the Ca(2+) concentration in the fusion/activation medium can enhance the fusion and blastocyst formation rates of fetal fibroblast NT porcine embryos without an additional activation treatment.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Calcium*
  • Cell Fusion
  • Cloning, Organism*
  • Culture Media* / chemistry
  • Embryo Transfer*
  • Female
  • Fibroblasts / physiology*
  • Fibroblasts / ultrastructure
  • Mice
  • Oocytes / chemistry
  • Oocytes / physiology
  • Swine / embryology

Substances

  • Culture Media
  • Calcium