Abstract: 560

IN VITRO DEVELOPMENT OF DOMESTIC CAT EMBRYOS AFTER RECOVERY OF OOCYTES FROM OVARIES EITHER FRESHLY COLLECTED OR STORED 24 H AT 4°C.

C Pope^{1 *}, R Schmid¹ , E O'Brien¹ , B Dresser^{1 *}
Audubon Inst Center for Research of Endangered Species, New Orleans, Louisiana ¹

Temporary storage of ex situ ovaries is often the only option if oocyte recovery and in vitro embryogenesis is an objective. Therefore, we compared in vitro development of domestic cat cumulus oocyte complexes (COC) recovered from ovaries held either at ~22°C for 2-5 h after ovariectomy (fresh, 55 sets) or after storage in Hepes-saline at 4°C for 24 h (stored, 88 sets). COC with uniformly medium to dark ooplasm were selected and held at ~38°C in Hepes (15 mM) TCM 199+BSA. For in vitro maturation (IVM) oocytes were cultured (500 l) in TCM 199+3 mg/ml BSA, 2 mM glutamine, lactate/pyruvate, 1 IU/ml hCG, 0.5 IU/ml eCG and gentamicin at 38°C in 5% O₂, 5% CO₂, 90% N₂ (=5% O₂). After 24 h IVM, COC were inseminated with fresh or cooled (4°C) spermatozoa in modified Tyrode's (Ty) +6 mg/ml BSA under oil. Fifteen h later, oocytes were rinsed and cultured in Ty+3 mg/ml BSA+nonessential AA (NEAA) at 38°C in 5% O₂. Two ± 1 day later, uncleaved oocytes were removed and embryos were cultured in Ty+10% FBS+NEAA+essential AA. On day 7 IVC, number of embryos developing into blastocysts was recorded and embryos were fixed for determination of cell number by fluorescence microscopy. Fewer oocytes (P<0.05) were recovered from stored ovaries (mean=4.4/set) than from fresh ovaries (mean=9.9/set). Cleavage frequency was not different between oocytes from fresh (40%) or stored (56%) ovaries. In vitro development to the morula (>16 cells)/blastocyst stages was similar between groups—38%/41% and 37%/32% for fresh and stored ovaries, respectively. Likewise, mean total cell number and blastocyst cell number of embryos from fresh ovaries was 89 and 122 vs 86 and 121 for embryos from stored ovaries (P>0.05). In summary, although fewer COC were recovered from stored ovaries, in vitro developmental competence was not reduced as compared to that of oocytes from fresh ovaries.

    This abstract is being presented on Tuesday, August 3 at 4:00 PM at Todd 133.