An attempt at intracytoplasmic sperm injection of frozen-thawed minke whale (Balaenoptera bonaerensis) oocytes.
Little is known about the characteristics of fertilisation events in minke whales. Cryopreserved minke whale oocytes and spermatozoa do not fertilise in a standard IVF. This study was conducted to investigate the pronucleus formation ability of cryopreserved minke whale oocytes and their subsequent development following intracytoplasmic sperm injection (ICSI). In experiment 1, frozen-thawed minke whale immature oocytes were cultured for in vitro maturation (IVM) in a maturation medium (TCM199) supplemented with either porcine follicle stimulating hormone (pFSH)/estradiol-17beta (E2) or pregnant mare's serum gonadotropin (PMSG)/human chorionic gonadotropin (hCG). After 120 h of IVM, oocyte survival was examined before ICSI, and showed no significant difference in morphological normality (24-36%) between the two IVM media. Two-cell embryos (two oocytes from 21 sperm-injected oocytes) were obtained when the maturation medium was supplemented with pFSH/E2 or PMSG/hCG. In experiment 2, cryopreserved maturing oocytes were investigated for the effects of repeat-culture (2 h or 24 h) on survival before ICSI. Pronuclear formation and development were examined for the effects of sperm pretreatment with dithiothreitol (DTT) and oocyte activation with ethanol at ICSI. A frequency of 49-69% of frozen-thawed maturing oocytes was used for ICSI. Although oocyte activation did not produce a significant difference in survival, pronucleus formation and embryonic development, 2- and 4-cell cleaved oocytes were observed after injection of sperm pretreated with DTT.
Asada
M
M
Obihiro University of Agriculture and Veterinary Medicine and The Institute of Cetacean Research, Japan.
Wei
H
H
Nagayama
R
R
Tetsuka
M
M
Ishikawa
H
H
Ohsumi
S
S
Fukui
Y
Y
D000818:Animals; D015925:Cryopreservation; D004229:Dithiothreitol / Q000378:metabolism; D004625:Embryo, Nonmammalian / Q000502:physiology; D005260:Female; D012739:Gonadal Steroid Hormones / Q000502:physiology; D008297:Male; D009865:Oocytes / Q000502:physiology; D020554:Sperm Injections, Intracytoplasmic / Q000379:methods*; D013094:Spermatozoa / Q000502:physiology; D014907:Whales
D012739:Gonadal Steroid Hormones; D004229:Dithiothreitol