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Gagarinskiy E. L., Uteshev V. K., Fesenko E. E. Storage of grass frog Rana temporaria (Ranidae, Amphibia) oocytes in a carbon monoxide and oxygen mixture under pressure. Current Studies in Herpetology, 2025, vol. 25, iss. 3, pp. 159-164. DOI: https://doi.org/10.18500/1814-6090-2025-25-3-4-159-164
Storage of grass frog Rana temporaria (Ranidae, Amphibia) oocytes in a carbon monoxide and oxygen mixture under pressure
The development of a technology for hypothermic preservation of amphibian oocytes is one of the urgent tasks for cryobiology. The previously proposed method of “dry” hypothermic preservation in small sealed containers allowed us to increase the duration of refrigerated storage of unfertilized oocytes of the grass frog Rana temporaria up to 7 days (fertilization 40%; hatching 20%). The aim of this study was to increase the effectiveness of the previously proposed approach using the technology of storing biological objects under pressure of a gas mixture (oxygen and carbon monoxide), usually used for warm-blooded animal organs. In the CO+O2 6.5 atm group, a decrease in the quality of preserved oocytes occurred only on the 7th day of storage. The number of fertile oocytes (91±8%) and larvae (80±14%) during preservation for up to 4 days in the CO+O2 6.5 atm group did not significantly differ from the values obtained in the native control group of 95+4% and 91±5%, respectively. On the 7th day of preservation, a sharp decrease in the hatching rates was observed in the CO+O2 6.5 atm group compared to the control group. A series of additional experiments showed that reducing the pressure from 6.5 down to 2.5 atm (over atmospheric) increased fertility from 34±21% in the CO+O2 6.5 atm group up to 68±8% in the CO+O2 2.5 atm group, and the number of larvae from 12±11 to 23±10% during the seven days preservation of oocytes. Thus, we have demonstrated the possibility of using the gas preservation technology with a CO+O2 mixture to prolong hypothermic storage of amphibian oocytes for up to 12 days. The recommended pressure of the gas mixture in the chamber is 2.5 atm (over atmospheric).
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