Pregnancies after fertilization of cow follicular oocytes, their incubation and their transfer to the cow uterus

128

The structure of oocytes aspirated from the dominant and its subordinate follicles was investigated from the achievement of follicular dominance to ovulation. Ovulation was induced in 18 heifers and 5 cows by injection of cloprostenol at days 8-14 (day 0 = day of ovulation), and follicular development was monitored by ultrasonography. The animals were slaughtered at days 3-11, but animals slaughtered on days 8-11 were given a second injection of cloprostenol at day 7 to allow ovulation of the dominant follicle of the first follicular wave. Oocytes were aspirated from the dominant (largest) and two largest subordinate follicles and processed for transmission electron microscopy, whereas the follicular fluids were analyzed for concentrations of estradiol-17 beta (E2) and progesterone (P4). Dominant follicular growth was associated with increase in the concentration of E2 and P4 in the follicular fluid, which was E2-dominated. From days 3-7, the dominant oocytes had pronounced junctional contacts with the cumulus cells and a nonundulating nuclear envelope but showed an increase in the number of lipid droplets and a decrease in the size of Golgi complexes, the size of cortical granule clusters, and the number of microvilli stacks. After cloprostenol injection on day 7, but before the anticipated LH surge, the dominant oocytes showed a reduced oocyte-cumulus contact, vacuolization of the nucleolus, undulation of the nuclear envelope, and dispersal of the mitochondrial clusters. The morphological alterations occurring in the dominant oocytes before the anticipated LH surge are suggested to be a prerequisite for the oocyte to achieve the competence to undergo final maturation. Subordinate follicles ceased growing at about days 3-4 and their follicular fluid had low E2:P4 ratio or was P4-dominated. Subordinate oocytes displayed degenerative features in their cumulus investment and nuclear activation and maturation especially after day 5. The structural changes associated with oocyte degeneration showed similarities with the processes seen before and during final maturation of the dominant oocytes.

Section: Discussionmentioning

confidence: 70%

Section: Introductionmentioning

confidence: 95%

Oocyte morphology in dominant and subordinate follicles

Assey¹,

Hyttel²,

Greve³

et al. 1994

128

“…Reports on in vitro maturation of bovine oocytes show that the developmental capacity of these oocytes is reduced after the culture period. The quality of the culture conditions has been improved during the last several years, and various research teams have reported live offspring after in vitro maturation and fertilization (Lu et al, 1987;Xu et al, 1987;Goto et al, 1988;Fayrer-Hosken et al, 0 1992 WILEY-LISS, INC.…”

Section: Introductionmentioning

confidence: 99%

Structural aspects of bovine oocyte maturation in vitro

Loos

Beneden

Kruip

et al. 1992

Bovine cumulus-oocyte complexes (COCs) were collected from 4-8 mm follicles and graded into four categories on their morphological characteristics. These four categories were matured in vitro and processed for transmission electron microscopy at 24 h after the onset of culture. The morphology of the four groups of oocytes was analysed and compared with that of oocytes that had matured in vivo and were collected 20-23 h after the preovulatory luteinizing hormone peak. After in vivo maturation, oocytes formed a homogeneous group with respect to their morphological characteristics. After in vitro maturation, the oocytes formed a heterogeneous group with respect to their morphology between as well as within the four categories of oocytes. Oocytes from the first three categories showed the same morphology after in vitro maturation. The fourth category showed some specific characteristics: 1) vacuolization, 2) flattening of cumulus cells, and 3) almost complete lack of cortical granules in some category 4 oocytes. These characteristics are interpreted as signs of degeneration. Besides these aspects of degeneration, other deviations from normal development were seen: 1) retraction of cumulus cell process endings from the oocyte without the breaking down of these processes, 2) retardation of some aspects of the cytoplasmic maturation, and 3) incomplete cumulus expansion. It is concluded that oocytes capable of development in vitro show a large morphological variability before the onset of culture. In vitro maturation systems can support normal development, but many oocytes show signs of degeneration and deviant development after in vitro maturation.

“…Lambs (Cheng et al, 1986;Crozet et al, 1987) and calves Xu et al, 1987;Goto et al, 1988;Stubbings et al 1988;Utsumi et al, 1988) can be produced from oocytes matured and fertilized in vitro. In vitro fertilized bovine eggs are transferred to rabbit (Hanada, 1985;Sirard et al, 1985) or sheep (Parrish et al, 1986;Lu et al, 1987;Sirard et al, 1988) oviducts to complete preimplantation development there, before they are transferred to final recipients.…”

Section: Introductionmentioning

confidence: 99%

“…In vitro fertilized bovine eggs are transferred to rabbit (Hanada, 1985;Sirard et al, 1985) or sheep (Parrish et al, 1986;Lu et al, 1987;Sirard et al, 1988) oviducts to complete preimplantation development there, before they are transferred to final recipients. Alternatively, the preimplantation period can be spent in vitro, in coculture with oviductal bovine cells (Goto et al, 1988;Lu et al, 1988;Eyestone and First, 1989;Rexroad, 1989;Ellington et al, 1990a,b;Kim et al, 1990). Calves were born after in vitro maturation, fertilization, and coculture (Lu et al, 1988;Aoyagi et al, 1990).…”

Section: Introductionmentioning

confidence: 99%

Birth of lambs after in vitro maturation, fertilization, and coculture with oviductal cells

Czlonkowska

Eysymont

Guszkiewicz

et al. 1991

Control ovine oocytes matured and fertilized in vitro were transferred to intermediate recipient ewes. After 5 days, 59% of eggs were recovered. Thirty-one (38%) reached morula/blastocyst stage. Twenty-one embryos at the morula or blastocyst stage were transferred to six recipient ewes, resulting in five pregnancies, of which four were maintained. Nine lambs were born (43%). In the experiment, 72 ooctyes matured and fertilized in vitro were cocultured for 5 days with sheep oviductal epithelial cells. Thirty-one eggs (43%) developed to the noncompacted morula stage. Transfer of 26 embryos to 11 recipient ewes resulted in two pregnancies (18%). Two male lambs were born. The result indicates that the coculture of in vitro matured and fertilized ovine eggs with sheep oviductal epithelial cells throughout the preimplantation period is compatible with further development to term.