Chemical activation of parthenogenetic and nuclear transfer porcine oocytes using ionomycin and strontium chloride.
Effective protocols for oocyte activation are crucial for study of parthenogenetic development and to produce nuclear transfer reconstructed embryos. This study investigated the use of ionomycin (ION) and strontium chloride (Sr(2+)) in the activation of parthenogenetic and nuclear transfer porcine oocytes. In-vitro-matured oocytes with a polar body were treated with varying concentrations of ION, Sr(2+) or its combinations, and then fixed or cultured to assess activation and development rates, respectively. Ionomycin concentrations of 10 and 15 microM resulted in more frequent oocyte activation and the 15 microM in advanced development compared to 5 microM (71.8 and 70%vs. 47.5%; P=0.04, and 43.7%vs. 19.3%; P=0.008, respectively). Oocytes treated with 10, 20 or 30 mM of Sr(2+) for 2 or 4h displayed a pronuclear formation rate ranging from 46.7 to 70%. When employed after a 5 min treatment with 10 or 15 microM ION, exposure to 10 mM Sr(2+) for 4 h resulted in higher pronuclear formation than did the 20 mM concentration (82 and 88.6%vs. 63.3 and 73.2%; P=0.03). Nuclear transfer reconstructed oocytes treated with 15 microM/5 min ION followed by 10 mM/4 h Sr(2+) resulted in a higher development to blastocyst stage compared to those treated with 15 microM ION alone (17.7 vs. 11.3%; P=0.06). In conclusion, we inferred that the inclusion of Sr(2+) in the activation protocol can benefit the development of nuclear transfer reconstructed porcine oocytes.
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Department of Animal Science, McGill University, 21111, Lakeshore road, Ste-Anne-de-Bellevue, Québec, Canada, H9X 3V9.
Lalonde
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A
Bordignon
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D000818:Animals; D002118:Calcium / Q000502:physiology; D019976:Cloning, Organism / Q000379:methods / Q000662:veterinary; D047108:Embryonic Development / Q000502:physiology; D005260:Female; D015759:Ionomycin / Q000494:pharmacology*; D053652:Nuclear Transfer Techniques / Q000662:veterinary*; D009865:Oocytes / Q000502:physiology*; D010312:Parthenogenesis / Q000187:drug effects / Q000502:physiology*; D013324:Strontium / Q000494:pharmacology*; D013552:Swine / Q000502:physiology*
D015759:Ionomycin; C025700:strontium chloride; D002118:Calcium; D013324:Strontium