title: Effect of holding technique and culture drop size in individual or group culture on blastocyst development after ICSI of equine oocytes with low meiotic competence.
abstract: The effect of medium-to-embryo ratio on blastocyst development of equine embryos from oocytes with compact cumuli was evaluated in the present experiment. In addition, two methods for holding oocytes before in vitro maturation were compared. In Experiment 1, oocytes cultured with roscovitine for 16-18h before maturation were fertilized by intracytoplasmic sperm injection and cultured individually in 2.5, 5, 10 or 50microl droplets. In Experiment 2, oocytes were either cultured with roscovitine or held in a modified M199 with 20% serum at room temperature (EH treatment) for 16-18h, then matured, fertilized and cultured in groups at 5microl medium per embryo. In Experiment 3, oocytes were held in the EH treatment, then were matured and fertilized. In Study 3.1, injected oocytes were cultured individually in drop sizes as for Experiment 1; in Study 3.2, groups of 2-7 oocytes were cultured in fixed drop sizes of 5 or 50microl. Blastocyst development rates of individually-cultured embryos were not significantly different among drop sizes in either Experiment 1 or 3 (15-29%). In Experiment 2, blastocyst rates were not significantly different between holding treatments (17-23%). In Experiment 3, for group-cultured oocytes, blastocyst development was not significantly different between 5 and 50microl drops (39 and 27%, respectively). In conclusion, compact-cumulus oocytes may be effectively held in the EH treatment before maturation, and single culture of equine embryos yields acceptable blastocyst development. The greatest blastocyst rate (39%) was obtained with group culture in a 5microl droplet.
authors.affiliation: Department of Veterinary Physiology and Pharmacology, College of Veterinary Medicine and Biomedical Sciences, Texas A&M University, College Station, TX 77843, United States.
authors.forename: Y H
authors.initials: YH
authors.lastname: Choi
authors.forename: L B
authors.initials: LB
authors.lastname: Love
authors.forename: D D
authors.initials: DD
authors.lastname: Varner
authors.forename: K
authors.initials: K
authors.lastname: Hinrichs
mesh_terms: D000818:Animals; D001755:Blastocyst / Q000502:physiology*; D018929:Cell Culture Techniques; D003470:Culture Media / Q000737:chemistry / Q000494:pharmacology; D005260:Female; D006736:Horses; D008540:Meiosis / Q000502:physiology*; D009865:Oocytes / Q000166:cytology / Q000187:drug effects / Q000502:physiology*; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D011687:Purines / Q000494:pharmacology; D000077546:Roscovitine; D013048:Specimen Handling; D020554:Sperm Injections, Intracytoplasmic / Q000662:veterinary*
chemical_list: D003470:Culture Media; D047428:Protein Kinase Inhibitors; D011687:Purines; D000077546:Roscovitine