title: Nuclear morphology, diameter and meiotic competence of buffalo oocytes relative to follicle size.
abstract: The nuclear morphology, diameter and in vitro meiotic competence of buffalo oocytes was compared relative to follicle size. Cumulus-oocyte complexes (COCs) were collected from 1-<2, 2-<3, 3-<4, 4-<6 and 6-<8 mm follicles from abattoir ovaries. Cumulus cells were removed using 3 mg mL(-1) hyaluronidase in saline and repeated pipetting. Denuded oocytes were measured, fixed in 3% glutaraldehyde, stained with 4,6-diamidoino-2-phenylindole and evaluated for nuclear morphology, namely the stage of germinal vesicle (GV) development before in vitro maturation (IVM). The COCs from >2-mm follicles were matured in vitro in their respective size groups for 24 h in Medium 199 supplemented with 10 microg mL(-1) follicle-stimulating hormone, 10 microg mL(-1) luteinizing hormone, 1.5 microg mL(-1) oestradiol, 75 microg mL(-1) streptomycin, 100 IU mL(-1) penicillin, 10 mM HEPES and 10% fetal bovine serum. Matured oocytes were fixed, stained and evaluated for GV status and meiotic development. The number of oocytes collected from follicles 1-<8 mm in diameter averaged 1.82 per ovary. Oocytes from follicles 1-<2 mm (107.7 +/- 1.6 microm), 2-<3 mm (108 +/- 1.1 microm) and 3-<4 mm (114.6 +/- 1.3 microm) in diameter were smaller in diameter (P < 0.05) than oocytes from follicles 4-<6 mm (124.4 +/- 1.3 microm) and 6-<8 mm (131.9 +/- 1.4 microm) in diameter. A majority of oocytes (P< 0.05) from <4-mm follicles was at the initial stages of GV development (GV-I, II and III), whereas oocytes from 4-<6- and 6-<8-mm follicles were at the final stages of GV-IV (35.0 and 21.6% respectively) and GV-V (49.1 and 67.5% respectively). Poor IVM rates of 32.0% and 32.7% to metaphase (M)-II were observed for oocytes isolated from 2-<3- and 3-<4-mm follicles, respectively, whereas significantly (P< 0.05) more oocytes from 4-<6- and 6-<8-mm follicles reached M-II (67.1% and 79.1% respectively). In conclusion, buffalo oocytes displayed a size-dependent ability to undergo meiotic maturation and we suggest that oocytes from >4-mm follicles should be considered in buffalo in vitro fertilization systems for better results.
authors.affiliation: Department of Theriogenology, Faculty of Veterinary Sciences, University of Veterinary and Animal Sciences, Lahore, Pakistan.
authors.forename: Muhammad Rizwan
authors.initials: MR
authors.lastname: Yousaf
authors.forename: Kazim Raza
authors.initials: KR
authors.lastname: Chohan
mesh_terms: D000818:Animals; D002020:Buffaloes; D002467:Cell Nucleus / Q000648:ultrastructure*; D048429:Cell Size; D005260:Female; D005307:Fertilization in Vitro; D008540:Meiosis; D009865:Oocytes / Q000166:cytology* / Q000648:ultrastructure; D006080:Ovarian Follicle / Q000166:cytology* / Q000648:ultrastructure