In vitro production of llama ( Lama glama) embryos by intracytoplasmic sperm injection: Effect of chemical activation treatments and culture conditions

Assisted reproductive technologies in the llama ( Lama glama) are needed to provide alternative methods for the propagation, selection and genetic improvement; however, recovery of adequate quantity and quality of spermatozoa for conventional IVF is problematic. Therefore, an effort was made to adap...

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Published in:	Animal reproduction science 2007-06, Vol.99 (3), p.342-353
Main Authors:	Sansinena, M.J., Taylor, S.A., Taylor, P.J., Schmidt, E.E., Denniston, R.S., Godke, R.A.
Format:	Article
Language:	eng
Subjects:	Activation animal breeding Animals Camelid Camelids, New World - embryology cell culture Cell Culture Techniques - veterinary chemical treatment Cleavage Stage, Ovum Co-culture Coculture Techniques Culture Media Embryo embryogenesis epithelial cells extraction Female females ICSI in vitro culture in vitro fertilization injection intracytoplasmic sperm injection ionomycin Ionomycin - pharmacology Lama Lama glama laparoscopy Llama llamas Male meiosis methodology Oocyte oocytes Oocytes - drug effects Oocytes - physiology oviducts Sperm Injections, Intracytoplasmic - veterinary spermatozoa ultrasonography
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Summary:	Assisted reproductive technologies in the llama ( Lama glama) are needed to provide alternative methods for the propagation, selection and genetic improvement; however, recovery of adequate quantity and quality of spermatozoa for conventional IVF is problematic. Therefore, an effort was made to adapt the intracytoplasmic sperm injection (ICSI) procedure for the in vitro production of llama embryos. The specific objectives of this study were: (1) to determine in vitro maturation rates of oocytes recovered by transvaginal ultrasound-guided oocyte aspiration (TUGA) or flank laparotomy; (2) to evaluate the effects of activation treatments following ICSI; (3) to evaluate the development of llama ICSI embryos in CR1aa medium or in an oviduct cell co-culture system. Llamas were superstimulated by double dominant follicle reduction followed by oFSH administered in daily descending doses over a 3-day interval. Oocytes were harvested by flank laparotomy or TUGA and matured in vitro for 30 h. Mature oocytes were subjected to ICSI followed by no chemical activation (Treatment A), ionomycin only (Treatment B) or ionomycin/DMAP activation (Treatment C). More oocytes were recovered by flank laparotomy procedure compared with TUGA (94% versus 61%, P < 0.05) and a greater number of oocytes harvested by flank laparotomy reached the metaphase-II stage (77% versus 44%, P < 0.05). After ICSI, the proportion of cleaved and 4–8-cell stages embryos was significantly greater when injected oocytes were activated with ionomycin/DMAP combination (63% and 38%, respectively, P < 0.05). The co-culture of ICSI embryos with llama oviduct epithelial cells resulted in progression to morula (25%) and blastocyst (12%) stages; whereas, all embryos cultured in CR1aa medium arrested at the 8–16-cell developmental stage.
ISSN:	0378-4320 1873-2232