Abstract
Rat sperm freeze-dried in a solution containing 10mM Tris-HCl and 1mM EDTA (TE buffer) can be preserved at 4°C, and oocytes injected with these sperm develop into offspring. However, the development of oocytes injected with freeze-dried sperm was quite low. We studied optimal culture conditions to improve the developmental ability of oocytes injected with freeze-dried sperm. The oocytes injected with fresh sperm were pre-cultured in modified Krebs-Ringer bicarbonate (mKRB) or modified rat 1-cell embryo culture medium (mR1ECM) with BSA before being cultured in mR1ECM. A high proportion of fertilized oocytes after pre-culturing in mKRB or mR1ECM/BSA developed to blastocyst compared to no pre-culturing (mKRB: 50% vs. 28%, mR1ECM/BSA: 49% vs. 18%). Pre-culturing in mKRB also led to a higher proportion of oocytes developing into blastocyst after the injection of freeze-dried sperm compared to that of oocytes without pre-culturing (32% vs. 15%). Moreover, offspring were obtained from oocytes injected with freeze-dried sperm preserved at 4°C for 6 months (17%) and 1 year (16%). This study demonstrated that culture conditions soon after the injection of sperm markedly influenced the subsequent development of oocytes. Furthermore, rat sperm freeze-dried in TE buffer could be preserved at 4°C for long term without deterioration.
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