In vitro fertilization with normal development in the sheep

Abstract

Ovine tubal (n = 87) and ovarian in vitro matured oocytes (n = 99) were fertilized in vitro with ejaculated spermatozoa capacitated for 8 h in modified defined medium buffered with Hepes. High levels of fertilization were obtained as assessed by development to two- to six-cell stage within 40 h (75.8% for ovulated and 62.6% for in vitro matured oocytes). Electron microscope analysis of oocytes 20-22 h after insemination indicated that in vitro fertilization approximated the in vivo events. Embryos (two- to six-cell) were transferred surgically to the oviducts of pseudopregnant rabbits. Three days later, 42 (from ovulated oocytes) and 15 (from in vitro matured oocytes) embryos were recovered; 26 (61.9%) and 10 (66.6%), respectively, had cleaved at least once. Embryos incubated in vivo (n = 20 from ovulated oocytes; n = 9 from in vitro matured oocytes) were transferred surgically to the uteri of seven and four recipient ewes resulting in four and two pregnancies, respectively, from which three and one, respectively, have been maintained (greater than 3 months). The first lamb resulting from the in vitro fertilization of an ovulated oocyte was born. In addition, six embryos (two- to four-cell) from tubal oocytes and ten embryos (two- to six-cell) from in vitro matured oocytes were directly transferred to the oviducts of two and three ewes, respectively. Two pregnancies resulting from in vitro matured fertilized oocytes are in progress (greater than 3 months).

Procedural improvements for in vitro production of viable uterine stage caprine embryos

“…Good results have been reported at 38.5"C (Crozet et al, 1987). Sperm incubation temperatures, 22°C or 38.5"C, for 5 h were not associated with significant differences (P < 0.05) in fertilization rates but viability was positively affected subsequently by sperm incubation at 38.5"C, at least under the best conditions used in the present work (Table 2).…”

“…A maturation interval of 27 h was found to be superior for caprine oocytes (DeSmedt et al, 1992;Keskintepe et al, 1993;Martin0 et al, 1994). The best insemination intervals for mammalian IVMFC were indicated as 24 h for bovine (Younis et al, 19911, 18 h for sheep (Crozet et al, 1987), and 18-24 h for goat (DeSmedt et al, 1992, Keskintepe et al, 1993. The results presented here lanta, GA.…”

Embryo transfer and related technologies in sheep reproduction

“…In this system, the semen was resuspended in medium containing oestrus sheep serum. This serum supplementation was also used by Crozet et al [29] and still remains the most effective capacitating factor in sheep. Later, Pugh and co-workers [85] reported the birth of the first lambs following IVM [18].…”

Mammalian oocyte maturation

“…The successful co-culture of cumulus-enclosed oocytes and granulosa cells have proved the basic role of these cells in the acquisition of oocyte cytoplasmic competence at the final stages of its maturation (rabbit, Motlik and Fulka, 1982 ;ovine, Staigmiller and Moor, 1984 ;Crozet et aL, 1987 ;bovine, Critser et aL, 1986 Osborn, 1983 ;Osborn and Moor, 1983). This conclusion fits well with the observations of Soupart (1975) who showed that normal male pronucleus formation only occurred in human oocytes when estradiol and then estradiol plus 17 aOH progesterone were added sequentially in the culture medium.…”