Decreased embryonic survival of in-vitro fertilized oocytes in rats is due to retardation of preimplantation development

Immature female rats (60-65 g) were injected with 4 i.u. PMSG on Day -2 and allocated to 3 groups. On the evening of Day 0, rats in Groups I and II were allowed to mate. Embryos were collected on Day 4 (Group I, control morulae) or Day 5 (Group II, control blastocysts) and were transferred into the oviduct or uterine horn of Day-4 pregnant recipient rats. On the transfer side of the recipients, the bursa had been peeled from around the ovary to prevent endogenous oocytes from entering the oviduct. For Group III, unmated donors were killed 65-67 h after PMSG injection. Ovulated oocytes recovered from the oviducts were fertilized in vitro and transferred 16-18 h later. Embryos developing from in-vitro fertilized (IVF) oocytes were recovered on Day 5, separated into morulae (Group IIIm) and blastocysts (Group IIIb) and transferred into Day-4 pregnant recipients similar to control embryos. Some embryos from each group were used to determine the mean number of cells/embryo. Embryo recipients were killed on Day 20. After transfer, the development of IVF oocytes was retarded compared to control embryos. IVF morulae contained significantly fewer cells/embryo than did control morulae but were able to implant and grow to fetuses, in proportions similar to controls, if transferred into the oviduct of the recipients. These results suggest that the developmental potential of rat oocytes fertilized in vitro is limited due to asynchrony between the embryo and the uterine environment at the time of implantation, rather than possible defects incurred by the oocyte during the fertilization procedure.     

Effect of removal of the ovarian bursa of the rat on infundibular retrieval and subsequent development of ovulated oocytes

The ovarian bursa was peeled from around one ovary of each rat and the rats were killed 1, 2, 3, 4 and 5 weeks later. The proportion of rats that maintained a bursa-free ovary did not change over the 5-week period (80-89%). Ovulation from the peeled ovary occurred in all rats but oocytes (1-4) were found in the ipsilateral oviduct in only 18% of the rats. The presence of oocytes in the oviduct was normally associated with some degree of re-encapsulation of the ovary. In another experiment rats were mated within 1 week of removal of the bursa from around the ovary. Unilateral pregnancy resulted in 92% of the rats. In a third experiment fertilized oocytes from mated donor rats were transferred into the oviduct next to the peeled ovary in 15 mated recipients. Of 85 zygotes transferred, 51 survived to be viable fetuses on Day 20. A single fetus developing from an endogenous oocyte was found in the transfer uterine horn in only one rat. This preparation may be useful in studies which attempt to determine the viability of oocytes that have undergone various manipulations in vivo or in vitro.     

Effects of time of PMSG and fixed-time GnRH injections on estrus incidence and fertility in physiologically different ewes pre-treated with progestogen-impregnated vaginal sponge during the nonbreeding season

A 2 × 2 factorial study for hormonal treatment was designed in 85 seasonally anestrous ewes with physiologically different status (nonparous, dry, and postpartum). All ewes were pre-treated with 60 mg of 6-methyl-17-acetoxy-progesterone (MAP) vaginal sponge for nine days and divided into four groups: Group I (22 ewes) — an i. m. injection of 600 i.u. pregnant mare's serum gonadotropin (PMSG) at the sponge removal (Day 0) and a single i.m. injection of 100 ug synthetic gonadotropin releasing hormone (GnRH) at 36 h after the sponge removal; Group II (21 ewes) — a PMSG injection at Day 0 and a saline injection at 36 h after the sponge removal; Group III (21 ewes) — a PMSG injection two days before the sponge removal and the GnRH injection at 24 h after the sponge removal; and Group IV (21 ewes) — a PMSG injection at Day -2 and a saline injection at 24 h after the sponge removal. The treated ewes were allowed to mate once with rams for five days after treatment. Estrus incidence and lambing rates were low (69.4% and 27.1%, respectively), probably due to the mating system and poor body condition of ewes used in the study. No effect of PMSG injection two days before with-drawal of MAP sponge and the fixed-time GnRH injection were observed in estrus incidence, fertility, and prolificacy. The present study indicates that the physiological status of ewes combined with management systems including feeding and mating would be important for out-of-season breeding with hormonal treatment.     

Morphology and proportion of inner cell mass of bovine blastocysts fertilized in vitro and in vivo

The morphology and proportion of inner cell mass (ICM) of bovine blastocysts cultured in vitro or in vivo in rabbit oviducts after in-vitro fertilization of in-vitro matured follicular oocytes were compared with those of blastocysts fertilized in vivo by a differential fluorochrome staining technique. The delineation of each ICM cell was improved by the transfer of embryos derived from in-vitro fertilization to a rabbit oviduct although the cell-cell contacts of ICM cells were not as tight as those from in-vivo fertilization. The proportions (15.8 and 14.9%) of ICM in blastocysts cultured in vitro at early and expanded stages were significantly lower than those cultured in rabbit oviducts after in-vitro fertilization and fertilized in vivo. These results show that the transfer of bovine embryos derived from in-vitro fertilization to the rabbit oviduct increased the proliferation of ICM cells to the level of embryos fertilized in vivo although the cell-cell contact of ICM cell is not improved by the process.     

Changes in peripheral inhibin levels and follicular development following treatment of buffalo with PMSG and Neutra-PMSG for superovulation

Fourteen buffalo were synchronized by administration of a prostaglandin (PG) salt Lutalyse in a double injection schedule, with a single intramuscular (im) injection of 25 mg at Day -13, followed by 30 mg and 20 mg im 12 h apart on Day 0 of the experiment. The 30-mg PG injection was designated as 0 h of the experiment. Group I animals (n = 4) received saline and served as the controls, while animals in Groups II and III (n = 5 each) received PMSG (2500 IU im at -48 h. Group III animals were administered 5 ml Neutra-PMSG intravenously at 60 h. Blood samples were collected every 48 h from Day -12 to Day -4, every 24 h from Day -4 to Day 0, every 3 h from Day 1 to Day 4 and every 24 h from Day 5 to Day 10 of experiment for the measurement of peripheral plasma inhibin concentrations by RIA. The number of large follicles (> 10 mm diameter) in animals of Groups II and III was assessed by ultrasonography on Days -2, -1, 0, 1, 2, 5 and 7 of the experiment. Treatment with PMSG of Group II animals resulted in a significant increase (P < 0.05) in plasma inhibin concentrations over that of control animals of Group I at 24 to 99 h, with a peak inhibin concentration of 1.01 +/- 0.31 ng/ml at 48 h. Treatment with Neutra-PMSG in Group III animals caused a significant reduction (P < 0.05) in the peripheral inhibin concentrations at 84 to 120 h and in the number of large unovulated follicles at 168 h compared with that in Group II animals. Peripheral inhibin levels in Group III animals came down to those of Group I after 21 h of Neutra-PMSG treatment. These results suggest that treatment of buffalo with PMSG for superovulation causes a marked rise in peripheral inhibin concentrations. Administration of Neutra-PMSG after PG treatment reduces the peripheral inhibin concentrations and the number of large unovulated follicles.     

Role of cumulus cells and serum on the in vitro maturation, fertilization, and subsequent development of rat oocytes

Immature oocytes were collected from immature female rats (60-65 g) 40 h after injection with 6 IU pregnant mare's serum gonadotropin (PMSG). Oocytes were matured cumulus-intact (CI) or cumulus-free (CF) in medium supplemented with 0.5% bovine serum albumin (BSA) or 5-20% serum for periods of up to 24 h. After assessment for nuclear maturation, the oocytes were exposed to epididymal sperm for fertilization in vitro. In vitro-matured and ovulated oocytes undergoing fertilization were transferred to unilaterally pregnant recipients for embryonic and fetal development. The presence of cumulus cells and serum shortened (by 2 h) the time required for polar body emission by in vitro-matured oocytes and also helped to increase significantly the penetrability of the oocytes by spermatozoa. A high proportion (45.6%) of fertilized oocytes showed evidence of abnormal fertilization following maturation in the absence of cumulus cells. Oocytes matured CI before fertilization were able to develop to viable fetuses (57.8%) in proportions similar to ovulated oocytes (55.0%) after in vitro fertilization. These findings indicate an essential role for cumulus cells in promoting normal cytoplasmic maturation of oocytes necessary for pronuclear formation and subsequent developmental capability.     

Pregnancies after co-culture of cumulus cells with bovine embryos derived from in-vitro fertilization of in-vitro matured follicular oocytes

Bovine follicular oocytes surrounded by cumulus cells for more than one-third of their surface were matured, fertilized and developed in vitro utilizing a co-culture system with bovine cumulus cells. Embryos developed into blastocysts were non-surgically transferred to the uteri of cows at Day 6, 7 or 8 (Day 0 = oestrus). Out of 6 recipient cows (19 blastocysts transferred), 3 became pregnant. One of the 3 pregnant cows carried twins. The results of this study demonstrated the viability of embryos obtained from in-vitro maturation of bovine oocytes followed by in-vitro fertilization and culture to the blastocyst stage in vitro.     

Pregnancies after in-vitro fertilization of cow follicular oocytes, their incubation in rabbit oviduct and their transfer to the cow uterus

Cow embryos fertilized in vitro (1-8-cell) (n = 113) were transferred surgically to the ligated oviduct of pseudopregnant female rabbits (31 +/- 4 h after 75 i.u. hCG). Rabbits were killed 99 +/- 5 h later and 67 embryos were recovered: 45 (67%) had cleaved at least once, 15 had reached the morula stage and 2 blastocysts were obtained. Transfer of cleaved embryos (2-8-cell) led to a high recovery rate (84%) compared to 39% for one-cell embryos. Of the embryos incubated for more than 99 h in the rabbit oviduct, 41% were at the morula stage. Embryos incubated in vivo (n = 21) (8-cell to blastocyst) were transferred to the uterus of 14 synchronized recipient heifers by a surgical (N = 5) or a non-surgical (N = 9) procedure: 6 pregnancies resulted (4 from the non-surgical procedures). In addition, 27 (2-8-cell) cow embryos developed in vitro were transferred to the oviduct of synchronized heifers by a paralumbar surgical approach on a standing animal, but no pregnancies resulted. It is therefore concluded that (1) the rabbit oviduct can be used to obtain cow eggs at an embryonic stage sufficiently advanced to permit transfer to the uterus of a synchronized recipient; and (2) the pregnancy rate after in-vitro fertilization and incubation in the rabbit oviduct are similar to results with fertilization in vivo.     

Successful pregnancies after transfer of embryos recovered from ewes induced to ovulate 24-29 days post partum

After lambing in late November, oestrus and ovulation were induced by using a CIDR device and PMSG in early weaned (N = 13) or lactating (N = 14) Border Leicester x Scottish Blackface ewes between 23 and 29 days after parturition. Ewes were intrauterine inseminated under laparoscopic visualization 54-55 h after CIDR-device withdrawal and eggs recovered on Day 3 of the cycle. Ovum recovery and fertilization rates were higher in lactating than in early weaned ewes, with fertilization being achieved as early as 24 days post partum in both groups. Of the 7 early weaned and 11 lactating ewes yielding eggs, fertilization occurred in 4 and 7 ewes respectively. A total of 20 embryos were transferred to the normal uterine environment of 15 recipient ewes in which the interval from parturition was greater than 150 days. Pregnancies were successfully established in 9 recipient ewes, resulting in the birth of 10 viable lambs. Prolactin concentrations were significantly higher (P less than 0.001) in lactating than in early weaned ewes throughout the study. Nevertheless, normal luteal function (as assessed by daily progesterone concentrations) was exhibited by 12 of 14 lactating and 8 of 13 early weaned ewes. Two post-partum donors in which the corpora lutea completely failed to secrete progesterone yielded fertilized eggs which developed to term when transferred to a normal uterine environment. The results show that sheep oocytes can be fertilized using laparoscopic intrauterine insemination as early as 24 days after parturition and that the resulting embryos are viable when recovered on Day 3 after oestrus and transferred to a normal uterine environment.     

Effects of an antiandrogen, flutamide, on oocyte quality and embryo development in rats superovulated with pregnant mare's serum gonadotropin

Increased oocyte degeneration in rats is associated with enhanced ovarian androgen secretion following superovulation with pregnant mare's serum gonadotropin (PMSG). To determine whether androgens may be causally related to oocyte degeneration, we examined the effects of an antiandrogen, flutamide, on oocyte quality and embryo development after induction of superovulation with PMSG. Immature female Sprague-Dawley rats were used for two experiments. In the first experiment, the females received either 5 mg flutamide or vehicle alone 30 and 36 h after 40 IU PMSG and were killed at 48, 60, and 72 h. Although total number of oocytes was not significantly different between the two groups, flutamide significantly reduced the percentage of degenerate oocytes ovulated at all the time intervals examined (p less than 0.01). In the second experiment, the females were given 4 IU PMSG (control) or 40 IU PMSG with either 5 mg flutamide or vehicle. The rats were mated and then killed on Days 2, 3, 4, and 5 of pregnancy. Compared to control, flutamide did not effectively prevent the early loss of preimplantation embryos nor the developmental retardation which took place in the vehicle-treated rats after Day 2. However, flutamide treatment was associated with a significant decrease in embryo degeneration and a significant increase in the percentage of cleaved embryos on Day 2 (p less than 0.01). Compared to levels associated with the vehicle regimen, ovarian and/or serum androgen levels in the flutamide-treated rats significantly decreased at 60 h (p less than 0.01) and on Day 2 (p less than 0.05).(ABSTRACT TRUNCATED AT 250 WORDS)     

Fertilizability of Superovulated Eggs by Estrous Stage-independent PMSG/hCG Treatment in Adult Wistar-Imamichi Rats

We investigated the fertilization and developmental ability of superovulated eggs obtained from adult Wistar-Imamichi (WI) rats, by using pregnant mare serum gonadotropin (PMSG) and human chorionic gonadotropin (hCG) treatment. Female WI rats, 11–13 weeks of age, were divided into four groups by estrous stage (metestrus [ME], diestrus [DE], proestrus [PE], or estrus [E]). PMSG (150 IU/kg) and hCG (75 IU/kg) were injected at an interval of 48 or 55 h and the female rats were mated with mature male rats. The ovulated eggs were collected 20, 24, and 27 h after hCG injection. Regardless of the estrous stage at the time of PMSG injection, the treated rats mated and ovulated similar to the untreated spontaneously ovulated rats (S group). Although the proportion of fertilized eggs in the E- and PE-treated groups was less than the S group 20 h after hCG injection, the proportion was not different among all treated and S groups 24 h after hCG injection. The proportion of fertilized eggs using in vitro fertilization and the proportion of offspring obtained from 2-cell stage embryo transfer did not differ among the treated and S groups. In comparison with PMSG/hCG-treated immature rats, mating and ovulation rate of adult rats were significantly higher. The proportion of fertilized eggs obtained from mated rats did not differ between immature and adult rats. These results demonstrate that adult WI rats are good egg donors for reproductive biotechnological studies using unfertilized or fertilized eggs.     

Interference with the preovulatory luteinizing hormone surge and blockade of ovulation in immature pregnant mare's serum gonadotropin-primed rats with the anti-androgenic drug, hydroxyflutamide

The involvement of androgens in the control of ovulation has been assessed by administration of the androgen antagonist, hydroxyflutamide, to prepubertal rats treated with pregnant mare's serum gonadotropin (PMSG) to induce first estrus and ovulation. Without human chorionic gonadotropin (hCG) injection, only 46% of rats that received six 5-mg, s.c. injections of hydroxyflutamide at 12-h intervals, beginning an hour before s.c. injection of 4 IU PMSG on Day-2 (Day 0 = the day of proestrus), had ovulated a mean of 1.3 +/- 0.4 oocytes per rat when killed on the morning of Day 1, whereas 92% of sesame oil-treated controls had ovulated a mean of 6.9 +/- 0.6 oocytes. After i.p. injection of hCG at 1600 h on Day 0, 92% of hydroxyflutamide-treated rats ovulated a mean of 8.3 +/- 1.2 oocytes compared to 100% of controls, which ovulated 7.3 +/- 0.4 oocytes per rat: these groups were not significantly different from each other, nor from control rats that received no hCG. Thus, exogenous hCG completely overcame the inhibitory effect of hydroxyflutamide on ovulation. Rats treated with PMSG and hydroxyflutamide without hCG were killed either on the morning of Day 0 to determine serum and ovarian steroid levels or on the afternoon of Day 0 to determine serum LH levels. Serum levels of estradiol-17 beta and testosterone in hydroxyflutamide-treated rats were significantly higher (178% and 75%, respectively; p less than 0.01) than levels observed in controls on the morning of Day 0. Ovarian concentrations of the steroids were also elevated in hydroxyflutamide-treated rats (p less than 0.01 for testosterone only).(ABSTRACT TRUNCATED AT 250 WORDS)     

Effects of culture duration and time of gonadotropin addition on in vitro maturation and fertilization of red deer (Cervus elaphus) oocytes

Immature red deer (Cervus elaphus ) oocytes (n = 1208) were collected from 1 to 4 - mm diameter follicles on ovaries and then cultured for 16, 20, 24 or 28 h (Groups I to IV) in TCM 199 supplemented with 10% FCS, 1 x 10(6) granulosa cells/ml and 1 mug/ml estradiol at 39 degrees C under 5% CO(2) in air. Gonadotropins (10 mug/ml, FSH and LH) were added to the culture medium at the start of culture (0 h) or after 6 h. Approximately one-third of the oocytes were examined for maturation, and the remainder were fertilized in vitro with frozen-thawed semen collected from a stag by electroejaculation. In vitro fertilized oocytes (n = 309) from four of the maturation treatment (Groups II and III in both gonadotropin treatments) were cultured for 7 d and examined for cleavage. Oocytes cultured for 16 h (Group I) had lower (P < 0.001) maturation rates (4.7%) than those in the longer culture durations (Groups II to IV: 68.9%). Culture for 20 (Group II) and 24 h (Group III) resulted in higher (P <0.001) fertilization rates than culture for 16 (Group I) and 28 h (Group IV) (18.3, 20.5, 7.1, 7.8%, respectively). The time of gonadotropin addition did not affect maturation or fertilization rates, but its addition at 6 h increased (P < 0.05) the percentage of oocytes cleaving (5.7 vs 12.5%). Oocytes cultured for 20 h (Group II) and with the delayed addition of gonadotropins cleaved most readily (18.2%). No embryos developed beyond eight-cell stage.     

Effects of in-vitro fertilization, culture, freezing and transfer on the ability of mouse embryos to implant and survive

The survival after transfer of frozen-thawed mouse blastocysts obtained from culture of in-vitro fertilized oocytes or 1- and 2-cell ova was compared. About 10% of transferred embryos developed to term in each group and there was no difference between embryos fertilized in vitro or in vivo. In addition to embryonic loss due to transfer, in-vitro cultivation and freezing reduced the proportion of fetuses considered viable at Day 15 of pregnancy (29.8 versus 50.7% and 26.3 versus 50.7% respectively). When used together these procedures had an additive effect on fetal wastage (18.4 versus 50.7%). In-vitro culture also entailed a significant increase of resorbing implantation sites (10.2 versus 4.3%). The re-expansion rate after freezing and thawing of blastocysts grown in vitro was paradoxically greater than that of blastocysts grown in vivo (85.8 versus 54.6%).     

Effect of endometriosis on ovulation, ovum pickup, fertilization and tubal transport in the rabbit

In 25 rabbits (Group E) endometrium from the right uterine horn was transplanted onto the peritoneum. In 25 rabbits (Group C) fat was transplanted. After a recovery period of 12 weeks the does were mated, and killed 24 h later. In Group E the implants had changed into cysts of 5-15 mm in diameter. Histological examination revealed endometrial glands and stroma in every specimen. Periadenexal adhesions did not develop in any animal. No differences were found between Groups C and E in the number of corpora lutea, the recovery rate, the fertilization rate and the transport of fertilized ova. These findings indicate that endometrial implants in the rabbit have no influence on the ovulatory mechanisms, the pickup function of the oviduct, the fertilization rate or on the transport of fertilized ova. Taking into account the restrictions of a rabbit model, it is suggested that the decreased fecundity in mild endometriosis in the human may be caused by disturbances in post-fertilization events, i.e. development of the pre-implantation embryo or implantation.     

Variables affecting the yield and developmental potential of embryos following superstimulation and in vitro fertilization in rhesus monkeys

Three follicular-stimulation protocols were compared to evaluate the yield and quality of oocytes obtained from rhesus monkeys. Five animals received a high-dose regimen of PMSG (protocol I), three received a lower-dose regimen (protocol II), and two received Clomid-Pergonal (protocol III). Oocytes were recovered at laparoscopy after HCG injection, fertilized in vitro, and cultured up to the blastocyst stage. Yields of mature oocytes were 17.2 ± 13.0 (80% of total recovered), 6.7 ± 6.6 (41%), and 4.5 ± 2.5 (90%) per stimulated cycle for protocols I, II, and III, respectively. Of mature oocytes, 72%, 45%, and 89% were fertilized for protocols I, II and III, respectively. Protocol I produced the most fertilized oocytes per stimulated cycle (11.6 ± 11.6) and the greatest E₂ production (approximately fourfold that maximally expected for an unstimulated cycle). For the combined protocol I and II results, there was a significant correlation (P ? 0.05) between mean embryo development score and E₂ production. Fertilized oocytes from protocol I yielded 7.8 ± 8.0 morulae and 6.8 ± 7.2 early zonal blastocysts per cycle. After transfer of nine singleton embryos to surrogate recipients, one live birth resulted. We conclude that our high-dose PMSG regimen offers the best means at present for obtaining susbstantial numbers of developmentally competent oocytes in rhesus monkeys and that more extensive use of rapid serum E₂ assays for monitoring both stimulated cycles and those of potential surrogate recipients could help to predict the success of in vitro fertilization and embryonic development following embryo transfer in rhesus monkeys.     

Activation of quiescent ovaries by administration of PMSG after LH-RH analogue treatment in heifers

The effect of pregnant mare serum gonadotrophin (PMSG) treatment on activation of quiescent ovaries was examined in heifers. Groups of thirteen, twenty and twelve heifers which showed ovulation within 2 d and corpus luteum (CL) development after injection with a luteinizing hormone releasing hormone analogue (LH-RH-A) were supplementally injected with 500 IU of PMSG (Group I); 500 IU of PMSG and 500 mug of Prostaglandin F(2alpha) analogue (PGF(2alpha)-A; Group II); and 500 mug of PGF(2alpha)-A (Group III) on Day 6 after the injection of 200 mug of LH-RH-A (Day 0), respectively. Estrus appeared in 33.3 to 45.0% of the heifers of the respective groups after the treatment. Ovulation occurred at a significantly (P<0.01) higher rate in Groups I (100%) and II (90.0%) than in Group III (41.7%). The ovarian cyclic activity was initiated in all the heifers that ovulated. Plasma progesterone levels decreased significantly (P<0.05) to about 1 ng/ml on Day 8 and Day 7 in Group I and Groups II and III, respectively. Plasma estradiol-17beta (E(z)) levels increased significantly (P<0.05), reaching a peak on Days 7 to 7.5 in Groups I and II but not in Group III. It is concluded that PMSG treatment stimulates maturation and E(z) secretion of a follicle, thus promoting ovulation and the onset of ovarian cyclic activity.     

Oocyte recovery, maturation and fertilization in vitro in the puma (Felis concolor)

Eight female pumas were treated i.m. with 1000 (N = 5) or 2000 (N = 3) i.u. PMSG followed 84 h later by 800 i.u. hCG. Eggs were recovered 24-26 h after hCG from ovarian follicles by using laparoscopy and transabdominal aspiration. Mature eggs were inseminated in vitro 4-6 h later whereas immature eggs were cultured for 24 h and then inseminated. Electroejaculates from 3 pumas were diluted with mKRB before insemination to evaluate the influence of sperm concentration on fertilization. Seven of 8 pumas responded with follicle development, and 140 eggs were recovered from 145 follicles (96.6%; 77 mature, 43 immature, 20 degenerate eggs; mean +/- s.e.m., 20.0 +/- 5.9 eggs/female). Overall fertilization rate was 43.5% (total eggs fertilized = 40) despite using inseminates containing 82-99% pleiomorphic spermatozoa. Of the 36 immature oocytes matured in vitro and inseminated, 12 were fertilized even though 50% of the inseminating spermatozoa contained an acrosomal defect. Fertilization rate of mature oocytes collected from follicles appeared unrelated (P greater than 0.05) to PMSG dose or number of spermatozoa/inseminate. This study demonstrates that a high proportion of follicular eggs can be recovered laparoscopically from adult pumas treated with PMSG and hCG. These gametes are capable of being fertilized in vitro (immediately or after maturation in vitro) even with low quality semen with a high incidence of sperm pleiomorphisms.     

Superovulation of immature rats by continuous infusion of follicle-stimulating hormone

Immature female rats were infused s.c. continuously over a 60-h period with partially purified porcine pituitary follicle-stimulating hormone (FSH) preparations differing in degree of purity and having widely divergent luteinizing hormone (LH):FSH potency ratios as defined by radioreceptor assays. Rats infused with the more purified FSH preparation (FSH-A) ovulated a mean of 60-85 oocytes per rat on the morning of the third day (Day 1) after FSH infusion was begun (on Day -2). The same total dose of FSH administered as a single s.c. injection or as twice daily injections over the same 60-h period resulted in ovulation in only a minority of treated rats (3/16), with none achieving ovulation rates approaching those of rats infused continuously. High fertilization rates (80% of ovulated oocytes) were observed in superovulated rats joined with fertile males on the evening of the second day of infusion (Day 0). Of the 67 +/- 7 fertilized ova per rat retrieved from oviducts flushed on Day 1, 52 +/- 8, or 80%, were accounted for as morulae or blastocysts recovered when oviducts and uteri were flushed on the morning of Day 5, demonstrating essentially normal developmental rates and high survival rates in reproductive tracts of superovulated females during the preimplantation period. Infusion of rats with the same dose of a less well-purified FSH preparation (FSH-E) containing 20 times as much LH activity, or injection of rats with a superovulatory dose of pregnant mare's serum gonadotropin (PMSG) (40 IU), were much less effective in causing superovulation, with ovulation rates of 17 +/- 6 and 34 +/- 8 oocytes/rat, respectively, compared to 79 +/- 9 oocytes/rat infused with the FSH preparation (FSH-A) containing lower LH activity.(ABSTRACT TRUNCATED AT 250 WORDS)     

Developmental capacity of bovine oocytes collected from ovaries of individual heifers and fertilized in vitro

Bovine follicular oocytes from individual heifers (n=49) were separately matured, fertilized with frozen-thawed spermatozoa and cultured with cumulus cells. Although there were great variations in the number (mean+/-SD=19.1+/-11.9) of oocytes collected from individual heifers and the percentages of the oocytes cleaved 48 hours after insemination (mean+/-SD=69.5+/-18.4) and developed to the morula stage 7 days after insemination (mean+/-SD=10.9+/-10.9), there were significant correlations between the numbers of oocytes collected and cleaved (the correlation coefficient: r= 0.9336) or developed to morula stage (r=0.6633), indicating that oocytes from different heifers have the same developmental ability after in vitro fertilization. Ten morulae and 12 blastocysts which were obtained 7 and 8 days after insemination were transferred, one by one, to each uterine horn of 11 recipients. At Day 60 of pregnancy, 8 (80%) fetuses were identified in 4 (80%) of 5 recipients into which 10 embryos were transferred at Day -1 of synchrony. However, only 3 (25%) fetuses were identified in 2 (40%) of 6 recipients into which 12 embryos were transferred at Day 0 or +1 of synchrony.