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Propofol is a common anaesthetic agent used for oocyte retrieval procedures during in-vitro fertilization (IVF). The effect of Propofol in vitro on mouse oocyte maturation, fertilization and embryo cleavage was studied. In this study, 551 cumulus-free and 222 cumulus-enclosed oocytes from mice stimulated with pregnant mare's serum gonadotrophin (PMSG) were incubated for 30 min in medium containing 0, 100, 1000 or 10,000 ng/ml of Propofol prior to in-vitro maturation. Also, 325 cumulus-enclosed oocytes from mice stimulated to ovulate with PMSG/human chorionic gonadotrophin (HCG) were incubated for 30 min in similar concentrations of Propofol prior to IVF. Maturation, fertilization and cleavage rates were compared. A significant decrease in the in-vitro maturation rate was observed only when the cumulus-free and cumulus-enclosed oocytes were exposed to 10,000 ng/ml Propofol (P < 0.0074 and P < 0.0001 respectively). Fertilization and embryo cleavage rates were not significantly different compared with the controls. These findings give some reassurance with respect to human IVF. However, further studies on the potential effects of Propofol on implantation and pregnancy outcome following IVF are needed.

Ovaries of the seasonally-breeding snow leopard (Uncia uncia) were examined to determine whether they were responsive to exogenous gonadotrophins throughout the year. The potential of laparoscopic artificial insemination (AI) also was assessed for producing offspring. During the non-breeding, pre-breeding, breeding and post-breeding seasons, females (n = 20) were treated with a standardized, dual-hormone regimen given intramuscularly (600 I.U. of equine chorionic gonadotrophin followed 80-84 h later with 300 I.U. of human chorionic gonadotrophin (hCG)). Laparoscopy was performed 45-50 h after administration of hCG, and all ovarian structures were described. Females with fresh corpora lutea (CL) were inseminated, and anovulatory females were subjected to follicular aspiration to examine oocyte quality. Snow leopards responded to exogenous gonadotrophins throughout the year. Mean number of total ovarian structures (distinct follicles mature in appearance plus CL) did not differ (P > or = 0.05) with season, but the proportion of CL: total ovarian structures was greater (P < 0.01) for the breeding season compared with all other seasons. The proportion of females ovulating was greater (P < 0.05) during the breeding and post-breeding seasons than during the pre-breeding and non-breeding seasons respectively. No Grade-1 quality oocytes were recovered from follicles of anovulatory females. Serum concentrations of oestradiol-17 beta appeared elevated in all females, and neither oestradiol-17 beta concentrations nor progesterone concentrations differed (P > or = 0.05) among seasons. Of 15 females artificially inseminated, the only one that was inseminated in the non-breeding season became pregnant and delivered a single cub. This is the first successful pregnancy resulting from AI in this endangered species.

Twenty 3-6-week-old Merino lambs were given either 3, 1 or 0 treatments of 50 microg oestradiol benzoate and (48 h later) a 1.5 mg Norgestamet implant left in situ for 9 days (3-, 1- and 0CYCLE+G). On Day 7 after the last implant insertion, and on the same day for 0CYCLE+G, each lamb received 400 I.U. pregnant mare serum gonadotrophin and 6 mg follicle-stimulating hormone (FSH). The reproductive tracts were removed for oocyte collection 24 h after FSH. Reproductive tracts were also collected from 16-24-week-old lambs (n = 31) (0CYCLE-G). The number of antral follicles per ovary was similar for the 3-, 1- and 0CYCLE+G treatments. Similar rates of in vitro maturation and monospermic fertilization were obtained for all groups. The proportion of blastocysts per cleaved oocyte was higher for 1CYCLE+G (50.5%) than for 3CYCLE+G (32.9%), 0CYCLE+G (24.3%), and 0CYCLE-G (11.8%) (P < 0.05). Viable fetuses were obtained at Day 93 of pregnancy after transfer of embryos from all treatments. These results indicate that a single treatment with oestrogen and progesterone, prior to gonadotrophin stimulation, will increase the yield and developmental capacity of oocytes from prepubertal sheep.

The paucity of information and the unagreed consensus about the estrous cycle of the Mongolian gerbil (Meriones unguiculatus) have rendered such studies rather difficult. The estrous cycle of the Mongolian gerbils in this report were divided into 5 stages: Stage I, proestrus; Stage II, estrus (scattering); Stage III, estrus (gathering); Stage IV, metestrus and Stage V, diestrus. The normal estrous cycle in the Mongolian gerbils was 4-6 days long. In our experimental conditions, 67.9% of virgin females had a 4-6 day cycle, whereas 26.4% had an unsettled cycle and 5.7% assumed pseudopregnancy. The changing pattern of the estrous cycle and copulatory behavior of the Mongolian gerbils after hormone (PMSG: pregnant mare serum gonadotropin, 10 IU; hCG: human chorionic gonadotropin, 10 IU) injection were examined. The change in the estrous stage after hormone injection could be roughly classified into two types. The vaginal smear of virgin females injected with PMSG at Stage I, II or III changed to Stage V the next day and to Stage I or Stage II after 48 hr, but in the case of Stage IV or V, the smear changed to Stage I after 48 hr. The females injected with PMSG at Stage I, II or III copulated at from 13:00 to 23:00, whereas others injected at Stage IV or V scarcely copulated at all between 13:00 and 23:00. Mating of these females with the male midnight was not observed.

Combination regimens of equine chorionic gonadotrophin (eCG) and human chorionic gonadotrophin (hCG) are used in ovarian stimulation protocols for assisted reproduction of felids. In the present study, domestic cats exhibited decreased ovarian responsiveness after repeated administration of eCG-hCG and a possible immunological mechanism for this ovarian refractoriness was investigated. An ELISA was used to analyse sera from male, naive female and previously eCG-hCG-stimulated (1x, 3x and 4x) female cats for the presence of immunoglobulins binding to eCG, hCG and pig FSH (pFSH). The sera of cats receiving multiple eCG-hCG injections, at intervals of 44-50 days, displayed greater eCG and hCG-binding than did the sera of male, naive female or female cats stimulated once, and demonstrated variable affinity for pFSH. In preovulatory and postovulatory ovarian stimulation assays, mice injected with an eCG-antisera mixture had lower ovary masses than did mice injected with eCG-saline and fewer ovulated oocytes compared with mice treated with eCG-naive sera. Treatment of queens that were refractory to eCG-hCG with a pFSH-hCG regimen caused a rebound in development of ovarian follicles but not in oocyte maturity. These studies indicated that repeated treatment of domestic cats with eCG and hCG may cause an immunologically mediated refractoriness to ovarian stimulation. Although alternative gonadotrophin regimens may alleviate this refractoriness, a preferable strategy might be the avoidance of potential immunological complications through the cautious use of eCG and hCG in domestic and endangered nondomestic felids.

Objectives: The increasing use of drugs for ovarian stimulation and the possibility of long-term risks has actualized a quantitative assessment of the use of such therapy. The aim of the study was to analyze the development in the sale of different types of drugs used for ovarian stimulation in Denmark during the last two decades., Material: Sale statistics of clomiphene citrate, cyclophenile, human menopausal gonadotropin (hMG), mare menopausal gonadotropin (mMG) and human chorionic gonadotropin (hCG) in Denmark 1973-1993., Methods: The number of defined daily doses (DDD) was calculated for each product group. On given assumptions the number of cycles of different treatment regimens and the number of treated women was calculated., Results: The sale has increased almost exponentially throughout the last two decades: Clomiphene citrate 11 fold, hMG 30 fold, and hCG 5 fold. Today, among women 15-44 years old, the estimated incidence rate of women treated with clomiphene alone is about 2.7/1,000/year, and the incidence rate of women treated with clomiphene/hCG and hMG/hCG account for about 3.1/1,000/year and 1.9/1,000/year, respectively., Conclusion: Any study concerning short- and long-term effects of ovarian stimulation have to consider this secular trend.

Ovulated oocytes are used to obtain estimates of germ cell aneuploidy following in vivo treatment with various chemicals. Such studies commonly use exogenous gonadotrophins to increase the number of ovulated oocytes and to induce ovulation during specific time periods. However, the gonadotrophin dosages have not been evaluated relative to the possibility that synergistic and antagonistic interactions may exist between the hormones and the chemical being studied. We, therefore, studied the interactions of different hormone dosages (5 IU pregnant mare's serum (PMS) and 2.5 IU human chorionic gonadotrophin (HCG), 7.5 IU PMS and 5 IU HCG, and 10 IU PMS and 7.5 IU HCG) alone (controls) and in combination with 0.2 mg/kg of the aneugen vinblastine sulfate (VBS) relative to the number of oocytes ovulated, the frequencies of metaphase I (MI), diploid and aneuploid oocytes. The results indicated that the proportions of oocytes ovulated were not significantly (P > 0.05) different among the various control and VBS groups. VBS, however, resulted in significantly (P < 0.01) higher levels of MI oocytes than in controls. In the VBS-treated females, higher frequencies (P < 0.05) of MI oocytes were found in the 7.5 IU PMS and 5 IU HCG and 10 IU PMS and 7.5 IU HCG groups as compared to the 5 IU PMS and 2.5 IU HCG group. The frequencies of diploid oocytes did not differ significantly (P > 0.05) within the control of VBS groups; whereas, when comparisons were made between each control and VBS group, only the difference between the 5 IU PMS and 2.5 IU HCG groups was significant (P < 0.05).(ABSTRACT TRUNCATED AT 250 WORDS)

Oestrus was synchronised in 57 Bos indicus heifers using norgestomet-oestradiol and pregnant mare serum gonadotrophin. Oestrus was detected by observations made at six-hourly intervals, using oestrogen-treated and chin-ball harnessed steers, heatmount detectors, tail-paint and visual observation. Heifers were inseminated once at either a fixed time of 49.2 +/- 0.4 h (mean +/- SE; n = 29) after implant removal or 12.6 +/- 1.5 h (n = 28) after oestrus was detected. The mean (+/- SE) time to the onset of oestrus was 47.1 +/- 1.9 h, while 90% of heifers recorded in oestrus were detected within 66 h of implant removal. Heatmount detectors were significantly more efficient at detecting oestrus than chin-ball harnessed steers, tail paint or visual observation (P < 0.001). A higher pregnancy rate was obtained in heifers inseminated after oestrus detection compared with heifers inseminated at a fixed-time (57.1 vs 34.5%; P = 0.043) and a higher pregnancy rate was obtained in heifers classified as easy to inseminate compared with heifers classified as difficult to inseminate (57.8 vs 0%, P < 0.001). We conclude that heatmount detectors are an efficient means of detecting oestrus in synchronised B indicus heifers and that pregnancy rates can be increased when insemination follows oestrus detection compared with a fixed-time insemination regimen.

Adult Wistar-Imamichi rats were induced to superovulate by an i.p. injection of PMSG followed by an i.p. injection of hCG 48 hr later. The doses of PMSG and hCG administered were 0, 75, 100, 150, 300 iu/kg body weight. In this study the doses were based on iu/kg body weight rather than iu/rat to facilitate accurate induction. The following results were obtained. In the adult rats (10 to 15 weeks old) PMSG induced an increase in the number of morphologically normal and fertilized oocytes, as well as in the total number of superovulated oocytes, dose-dependently upto 150 iu/kg. The increase was significant compared with control rats, but PMSG treatment had only half this effect in young rats (6 to 9 weeks old), compared with the adult rats. These results suggest that sensitivity to PMSG changes with age. The proportion of fertilized oocytes among the morphologically normal oocytes was not significantly different in young and adult rats, and both groups had high percentages. In view of the results of this study, the authors recommend that, to collect a large number of fertilized oocytes required for generation of transgenic rats, the induction of superovulation be performed in adult rats aged 10-15 weeks rather than young rats aged 6-9 weeks and giving PMSG at a dose of 150 iu/kg and hCG at a dose of 75 iu/kg.

The effects of pregnant mare serum gonadotrophin (PMSG) dose and PMSG antiserum on superovulation in crossbred beef cows were studied. In experiment I, three groups were treated with 1200, 2400 or 3600 IU of PMSG and 48 h later with prostaglandin (PGF). The mean numbers of corpora lutea (CL), unovulated follicles, and total ova/embryos collected increased as the PMSG dose increased. The percent of fertilized ova and transferable embryos was lowest in the highest dose group (p < 0.05). In experiment II, all cows received 2500 IU of PMSG; groups 1 and 2 were treated with sheep anti-PMSG serum at 48 h or 60 h after PGF; group 3 cows were PMSG-only controls. The number of CL was lowest and the number of unovulated follicles highest in the PMSG-only group (p < 0.05). The number of CL was higher in group 2 (anti-PMSG at 60 h) than in the control group, with the anti-PMSG at 48 h not different from the other groups. Numbers of total ova/embryos, fertilized ova, and transferable embryos were higher (p < 0.05) in both antiserum-treated groups relative to the PMSG-only group. We conclude that superovulation of beef cows with PMSG and treatment with PMSG antiserum will induce a higher superovulatory response and will result in higher CL numbers and fewer unovulated follicles. Further, the variability in the superovulatory response to PMSG treatment was still evident when PMSG antiserum was administered.

Morphological changes in ovaries and hormonal changes as well as changes in Na, K, Ca, Mg, and P blood plasma levels were observed after superovulation induced by the administration of PMSG and the neutralization of this hormone with monoclonal antibodies Neutra-PMSG administered either 72 or 108 h later. The introduction of Neutra-PMSG 108 h after PMSG injection clearly decreases the number of surviving non-ovulated follicles with a diameter > 10 mm (1.7 +/- 0.8 vesicle in an individual cow on the average) in comparison to the group without Neutra-PMSG (19.4 +/- 9.5). The efficiency of ovulation in the group treated with Neutra-PMSG in the 108th h of the experiment (8.2 +/- 4.6 corpora lutea per cow on the average), did not differ statistically from the group treated with PMSG only (12.4 +/- 10.6). Early administration of Neutra-PMSG (72h), totally inhibits superovulation. Observations showed, that injection of Neutra-PMSG in the 108th h, caused a considerable decrease in the estradiol level, beginning with the 120th h of the experiment. Determination of the progesterone blood plasma level reflects the number of corpora lutea and can be helpful in evaluating the effects of superovulation. Superovulation did not effect the level of Na, K, Ca, Mg, and P in the blood plasma.

A method is described for the non-surgical transfer of embryos in pigs. Embryos at the 8-cell to the hatched blastocyst stage, recovered on days 4 to 7 of the oestrous cycle by flushing the oviducts and uteri of superovulated donors, were transferred transcervically into the uterine body of anaesthetised recipient gilts using a sterile disposable plastic spiral catheter and an embryo transfer cannula. Fifty-eight non-surgical transfers have been performed and six pregnancies were established. Eight and three normal fetuses were recovered from two recipients slaughtered between 35 and 45 days after embryo transfer. Three recipients came to term and gave birth to litters of two, six and seven living piglets. One recipient aborted between 45 and 60 days of gestation.

Our methodology for obtaining zona-free ova from golden hamster females (Mesocricetus Auratus Waterhouse) is described here. Five groups of 5 females were superovulated injecting intraperitoneally pregnant mare gonadothropins (PMG) at different dosages. The control group received 0.25 ml of distilled water. We harvested 653 ova from the stimulated females with an average of 32.65 per individual (range 16-52). The control group produced a total of 50 ova with an average of 10 (range 0-14). PMG optimum dose was 25 IU with an average of 39 (S.D. +/- 10.93) (n = 195 ova). The best conditions for denuding the cumulus was a 0.01% hyaluronidase solution with an average time of 10 minutes (S.D. +/- 03'56"). The optimum conditions for digesting the zona was 0.20% trypsin during 03'20" (S.D. +/- 01'19"). The proof that the ova were not damaged by the procedure was by means of a supravital stain (Eosin Y). All mature ova (N = 681) were not affected by enzymes or manipulation. Final preparation used a 1% acetic acid fixation and staining with 0.01% methylene blue, both acting for 1-3 minutes. Animal gametes give us the opportunity to substitute human ova in biology of reproduction studies and experiments, avoiding technical, ethical and moral problems.

Oestrus was induced in 14 anoestrous beagle bitches by intramuscular injection of PMSG in a dose of 20 I.U./kg once daily on five consecutive days, followed by an additional single i.m. injection of 500 I.U. of hCG on the fifth day. The day on which the first injection was given was counted as Day 1 of the experiment. Between the fourth and sixth day, the bitches began to attract the attention of male dogs and between Days 9 and 15 all bitches came in heat. Matings occurred on two to ten occasions, and six of the bitches conceived. The maximum concentrations of oestradiol in the plasma were in most cases reached on Days 10 or 12 and ranged from 42 to 195 pg/ml. In all cases progesterone concentrations rose sharply between Days 10 and 20. The incretion phase of the corpora lutea was noticeably brief in the non-gravid bitches; in five of these eight bitches, anoestrous values below 2 ng/ml were already obtained before the 65th day of the experiment. These results indicate that the administration of PMSG on five consecutive days supplemented by a final single injection of hCG stimulates the ovaries adequately to afford good prospects of conception. The concentrations of ovarian hormones are discussed.

The yield of embryos from goats treated with single injections of FSH combined with low doses of PMSG was compared with that from goats treated with conventional superovulatory regimens based on multiple doses of FSH. In one experiment, 101 female goats were assigned to 10 groups. One group received six injections of FSH-P (FSH-P control); a second group received eight injections of Ovagen (Ovagen control). The remaining eight groups conformed to a 2x2x2 factorial design in which goats received a single injection of a low or high dose of either FSH-P or Ovagen in combination with a low or high dose of PMSG. All goats were run with harnessed, entire bucks. Ovaries were examined by laparoscopy and embryos were recovered from goats that had ovulated and been mated. The mean number of normal embryos collected per goat was significantly greater in the FSH-P control group in all except those groups treated with either low or high doses of PMSG combined with a low dose of Ovagen or a low dose of PMSG combined with a high dose of Ovagen. Most goats ovulated and ovulation rates were not significantly different from the mean of the FSH-P control, except for those treated with a low dose of FSH-P combined with either low or high doses of PMSG. Embryo recovery rates were significantly lower in groups treated with a high dose of PMSG combined with a high dose of either Ovagen or FSH-P, or with low doses of PMSG and FSH-P, than in the FSH-P control.(ABSTRACT TRUNCATED AT 250 WORDS)

Objective: Our null hypothesis was that the angiotensin II antagonist saralasin does not reduce the number of ovulations in the rat ovarian perfusion model., Study Design: Ovaries from pregnant mare's serum gonadotropin-stimulated immature rats were perfused with nutrient media to which luteinizing hormone and 3-isobutyl-1-methylxanthine had been added to induce ovulation. Test perfusions were treated with saralasin 1 mumol/L (n = 0.5) and compared with controls (n = 5) with the Student t test. Perfusions with both saralasin and angiotensin II and dose-response evaluations were performed., Results: Saralasin-treated ovulations were 6.6 +/- 1.3 (mean + SEM) compared with 18.6 +/- 3.9, p < 0.02. The effects of saralasin could be reversed with the addition of an equimolar amount of angiotensin II. Dose-response evaluations showed a progressive inhibition of ovulation at 10(-8) to 10(-6) mol/L., Conclusion: The angiotensin II antagonist saralasin inhibits ovulation in a dose-dependent fashion; this effect is canceled by the addition of equimolar concentrations of angiotensin II.

The descent and localization of eggs and embryos in individual segments of the reproductive tract of superovulated cows were studied in this work. For the induction of superovulation, serum gonadotropin (PMSG, Ivanovice in Haná) at a dose of 2,500-3,500 I.U. was used, in combination with 0.5 mg of Cloprostenol (Oestrophan, Spofa), administered 48 hours after gonadotropin treatment. The start of superovulation fell on days 9 to 12 of the sexual cycle and was conditioned by the presence of the corpus luteum (CL). After the onset of the heat, 2-3 inseminations were carried out using fresh semen. Donor cows were slaughtered 3, 4, 5, 6, and 7 days after the second insemination and isolated reproductive organs (Fig. 1) were divided into five segments (two on oviducts and three on uterine horns) by the applied ligature. In laboratory conditions superovulation response was determined accurately, the volume of ovaries was assessed according to water displacement and the segments of oviducts and uterus were rinsed with TCM 199 or PBS supplemented with FCS. 3, 4, 5, 6, and 7 days after insemination (Tab. I). 18.1 (+/- 3.55), 12.4 (+/- 0.91), 19.2 (+/- 2.86), 20 and 23 (+/- 2.44) CL on average were recorded, which corresponded to the ovulation of 64, 50, 56, 71 and 72 percent of stimulated follicles (Fig. 2). Within 3 to 7 days after insemination nearly triple enlargement of ovaries was also observed (Tab. I, Fig. 3). During the lavage of individual segments of the tubular reproductive tract, 38 per cent of eggs and embryos were detected in the uterus as early as 3 days after insemination (Tab. II). Unfertilized eggs and degenerated embryos were found in the 2nd and 3rd uterine segment, embryos at the stage of 8-16 blastomeres were localized in the 1st and 2nd segment of the uterus. Four days after insemination (Tab. III), about 64 per cent of eggs and embryos at the stage up to 16 blastomeres were found in the uterus, but embryos up to 32 blastomeres were still flushed out of the oviduct. On day 5 after insemination, 92 per cent of eggs and embryos were released into the uterus, being localized mostly in the cranial and medial part of the uterus (Tab. IV). 7.5 per cent of recovered eggs and embryos at the stage of early or compacted morulae were still detected in the oviducts.(ABSTRACT TRUNCATED AT 400 WORDS)

The characteristics and regulation of oocyte maturation in the normal menstrual cycle is reviewed and the mechanisms by which ovarian hyperstimulation with exogenous gonadotrophins, used in assisted reproductive techniques, can disturb this process, is discussed. It is postulated that derangements induced in the maturation of oocytes in the hyperstimulated cycle result in the recovery of poor quality oocytes at egg-pickup which have a reduced potential to undergo normal fertilisation and subsequent embryonic development.

A total of 950 female Wistar rats in 81 groups were involved in this study. Different groups of rats were stimulated with PMSG (0, 10 & 20 IU) at diestrus followed, 48-52 hr later, by different doses of HCG (0, 10, 20, 30 & 40) for ovulation induction. The dose-dependent effects of HCG, either with or without the use of PMSG for stimulation of multiple follicular development, on the quality of oocytes and their in vitro fertilisability, quality of Day 2-embryos, viability of pregnancy and status of embryos retrieved on Day 2, 3, 4 or 5 of pregnancy in different subgroups of rats were examined. Results showed that more oocytes and embryos fertilised in vivo were retrieved from rats supraphysiologically stimulated with 20 IU of PMSG. The addition of HCG did not increase the number of ovulated oocytes or Day-2 embryos. In other words, the number of oocytes or embryos produced is dependent on the dose of PMSG administered during diestrus rather than on the dose of HCG given for ovulation induction. Hence, no increase in the amount of HCG is required to effectively ovulate bigger cohort of preovulatory follicles in supraphysiologically stimulated rats. As was shown earlier, in vitro and in vivo fertilisation rates were reduced when higher doses of PMSG were used. Similarly, these rates were reduced when increasing doses of HCG were used in rats not previously stimulated with PMSG. When higher doses of HCG were used in rats stimulated earlier with PMSG (10 and 20 IU), the in vitro but not the in vivo fertilisation rates were further reduced.(ABSTRACT TRUNCATED AT 250 WORDS)

Follicles were sampled at three different times after treatment with 1200 iu pregnant mares' serum gonadotrophin (PMSG) or 12 mg ovine follicle-stimulating hormone (FSH), and from untreated control animals. The meiotic status and protein synthesis of the oocyte from each follicle was determined and correlated with the intrafollicular concentration of oestradiol and progesterone. Significantly higher amounts of oestradiol were present in PMSG-treated animals at sponge withdrawal than in FSH-treated and control goats. Twenty hours later, both oestradiol and progesterone concentrations in the PMSG group were higher than those in the FSH group, and were equivalent to control animals at the onset of oestrus. At 18 h after the administration of human chorionic gonadotrophin (hCG), oestradiol decreased markedly in all three treatment groups, whereas progesterone remained significantly higher in PMSG-treated follicles. Although these high concentrations of intrafollicular steroids were associated with a higher incidence of premature condensation of chromatin in oocytes, the two events were not causally related. Moreover, cytoplasmic maturation was not prematurely activated in these oocytes and a changed pattern of protein synthesis was observed in oocytes from all three treatment groups after the hCG injection. Whereas disturbances in follicular steroidogenesis of oestradiol and progesterone occur in vivo in goats superovulated with PMSG, they do not underlie the premature activation of the initial stages of nuclear maturation in oocytes but are associated with normal cytoplasmic maturation.

Five hundred and eight mature female Wistar rats divided into 35 different groups were stimulated with pregnant mare serum gonadotrophins (PMSG) (0, 5, 10, 20 & 40 IU) at the late diestrus stage to induce multiple follicular development. No chorionic gonadotrophin (CG) was used for ovulation induction. The quality of oocytes and their in vitro fertilisability, quality of Day 2-embryos, viability of pregnancy and status of fetuses on Day 14 of gestation and status of embryos retrieved on Day 2, 3, 4 and 5 of pregnancy in different subgroups of rats were examined. Results showed that more oocytes and embryos fertilised in in vivo were retrieved from rats supraphysiologically stimulated with 20 IU of PMSG. However, concurrent with the larger number, higher proportions of abnormal oocytes and embryos were found. High doses of PMSG caused lower in vitro fertilisability of oocytes and greater degrees of embryonic degeneration. Although, the number of oocytes and Day 2-embryos were higher in the 20PMGS dose group, the pregnancy rate was significantly reduced to 27%. In the 40PMSG group no viable pregnancy was noted. Most embryo demise occurred by day 3-5 of pregnancy, probably within the oviducts and before the implantation stage. In rats supraphysiologically stimulated with 20 and 40 IU of PMSG, the number of morphologically normal looking embryos was greatly reduced by Day 3-5 of pregnancy. In the 40PMSG group, there were no embryos retrieved by Day 4 and 5.(ABSTRACT TRUNCATED AT 250 WORDS)

The specific activity of NAD(+)-dependent 15-hydroxyprostaglandin dehydrogenase (PGDH) was found to increase in the ovaries of pregnant and pseudopregnant rabbits. The mean specific activity of cytosolic ovarian PGDH in 14- to 28-day pregnant rabbits was 24.3 +/- 8.1 nmol NADH formed/min/mg protein (n = 16) using PGE1 as substrate whereas in nonpregnant rabbits the specific activity was 1.5 +/- 0.8 nmol NADH formed/min/mg protein (n = 8). The reaction was dependent on NAD+; NADP+ did not support the reaction. In grouping the PGDH activities from pregnant rabbits into second (14-18 days) and third (2-28 days) trimester periods, no significant difference between values was found (26.1 +/- 8.9 vs 23.4 +/- 8.1 nmol NADH formed/min/mg protein, respectively). Western blot analysis of the ovarian cytosol using an antibody which was made to the purified lung PGDH of pregnant rabbits recognized an ovarian protein of identical molecular mass (30 kDa). Ovarian PGDH activities were also examined in rabbits treated with pregnant mare's serum gonadotrophin (PMSG) and human chorionic gonadotrophin (hCG) to induce a state of superovulatory/pseudopregnancy and only on day 11 following hCG treatment was an increase in PGDH specific activity observed. On day 11, the specific activity was 14.8 +/- 4.3 nmol NADH formed/min/mg protein whereas values on days 10 and 12 were only 1.1 +/- 1.1 and 1.0 +/- 0.8, respectively. PGDH activities on days 3, 7 and 16 were also low.(ABSTRACT TRUNCATED AT 250 WORDS)

On the day of diestrus, female mice, syrian hamsters and rats, showing regular 4-day estrous cycles, were injected with hCG or PMSG and were inspected for the presence of ovulation the following day. The dose level expected to cause an effect in 50% of the animals (ED50) was calculated using the Van der Waerden method. When hCG was injected into i. v., i. p. and s. c., the ED50 values per animal and per body weight (kg) in parenthesis were as follows; 0.2 (7.7), 0.3 (11.5) and 0.7 (26.9) I. U. for mice, 1.0 (9.5), 1.8 (17.1) and 2.6 (24.8) I. U. for syrian hamsters and 1.3 (4.6), 3.5 (12.3) and 7.5 (26.3) I. U. for rats, respectively. In PMSG study, the ED50 values per animal and per body weight (kg) in parenthesis were as follows: i. v., 0.8 (30.8); i. p., 2.0 (76.9); s. c., 2.8 (107.7) I. U. for mice, i. v., 3.6 (34.3); i. p., 8.0 (76.2); s. c., 13.2 (125.7) I. U. for syrian hamsters and i. v., 6.0 (76.8); i. p., 20.8 (73.0); s. c., 76.8 (269.5) I. U. for rats, respectively. From these results, the intravenous ED50 value was lower than other routes in three rodents with hCG or PMSG. In all injection routes, the ED50 value for mouse was lower than others. However, there were not significant differences in the ED50 values per body weight (kg) among three rodents. In particular, subcutaneous ED50 of hCG and intraperitoneal ED50 of PMSG were almost same values among three rodents, respectively. Given that the ED50 value per body weight (kg) in one of three rodents is determined, its value may be possible to be extrapolated to remaining two rodents.

The superovulatory response to gonadotrophin treatment during different months of the year was investigated in Kambing kacang goats, a tropical breed, in Malaysia. Sixty-three cycling does, fitted with progesterone impregnated intravaginal sponges for 17 days, received two days before sponge withdrawal, an intramuscular injection of either 10, 15 or 20 mg of follicle stimulating hormone (FSH) or 500, 1000 or 1500 iu of equine chorionic gonadotrophin (eCG). The dose of FSH was divided into four decreasing daily doses and each daily dose was subdivided into two and administered at 07.00 and 19.00. Fifty-four does detected in oestrus were mated with fertile bucks. The ovarian response was determined by laparoscopy and eggs were recovered surgically five or six days after oestrus. The ovulatory response (mean +/- standard deviation) based on corpora lutea was higher in the FSH (13.4 +/- 8.4 corpora lutea per doe, n = 20) than the eCG-treated groups (6.4 +/- 5.1 corpora lutea per doe, n = 25) but the difference was not significant (P greater than 0.05). Does responded to gonadotrophins throughout the year with more than 50 per cent of does responding during the rainy months compared with less than 35 per cent responding during the dry months. This difference was statistically significant (P less than 0.05). Egg recovery was better in the FSH (6.8 +/- 5.3 per doe, n = 20) than the eCG groups (3.0 +/- 3.8 per doe, n = 21) but the difference was not significant (P greater than 0.05).

17 African dwarf goats were treated with 4 different superovulation regimes. The results of the treatments were assessed spectroscopically and checked laparoscopically. The quantitative basis of the investigation is too small to allow statistically well-founded statements, but is indicative of distinctive trends. A proportion of 73% CL and CLA in the overall ovarian changes after treatment with 750 IE PMSG (2 days before removal of the sponge) and 125 micrograms PGF2 alpha (at the time of the sponge removal) proved an acceptable method of treating African dwarf goats as regards the requirement of labour and material as well as the superovulation effect.

Sminthopsis crassicaudata is a small (approximately 16 g) polyovulatory dasyurid marsupial which has the potential to become an important model species. This study examined the use of exogenous hormone treatment to manipulate the breeding of S. crassicaudata and as a means to obtain timed developmental stages for further study. Two thirds (21/32) of the females treated with 1.0 or 5.0 I.U. of pregnant mare serum gonadotrophin (PMSG) had ovulated when the contents of their reproductive tracts were examined 5 or 6 days later. Only one of eight females treated with 0.2 I.U. PMSG had ovulated in the same period. Although a similar proportion of animals treated with 1.0 I.U. and 5.0 I.U. ovulated, the ovulation rate was significantly lower when the higher dose was administered (mean of 10.5 ovulations per female v. 3.8 ovulations per female). In addition, the ovaries of 6/8 of the animals treated with 5.0 I.U. PMSG had luteinized follicles with degenerating oocytes, evidence of over-stimulation. Follicular luteinization also occurred in 4/8 animals treated with 1 I.U. PMSG. Oocyte maturation and ovulation occurred following PMSG stimulation without injection of synthetic gonadotrophin releasing hormone (GnRH). Treatment with a 10-micrograms dose of GnRH following PMSG seemed to have no effect on the outcome. Of the females that had ovulated by Day 6, three quarters had mated and some had fertilized eggs and two-cell embryos in the oviducts and uteri. In a further series of experiments the subsequent development of embryos conceived after PMSG treatment was assessed. Two thirds of treated females mated within 7 days of treatment and 60% of these matings yielded embryos when examined 11 days after PMSG. However, full-term development was only achieved in one animal. Gonadotrophin treatment of S. crassicaudata thus may have application as a means to obtain mature or maturing oocytes, cleavage stage embryos and blastocysts, but at this stage it appears not to offer promise as a method to achieve full-term development.

We have recently found that Tokishakuyakusan (TS), Keishibukuryogan (KB) or Unkeito (UT) inhibits in vivo DNA polymerase alpha activity in the rat uterus stimulated by PMS. In this study, uteri resected 24 h after injection of PMS on day 27 of age were incubated in vitro with 20 micrograms/ml of extract of TS, KB or UT for 4 h. The DNA polymerase alpha activity in uteri tended to decrease after the addition of TS, KB or UT with significant difference (P < 0.05) compared with TS-, KB- and UT-untreated control groups. These results suggest that TS, KB or UT, especially KB, tends to inhibit directly the enzyme activity in rat uterus.

The present study was conducted to investigate the mode of action of danazol by monitoring the first ovulation, serum luteinizing hormone (LH) levels and ovarian prostaglandin (PG) F2 alpha metabolism in pregnant mare serum gonadotropin (PMSG)-primed immature female rats. When danazol (750 mg/kg) was given p.o. once a day for 5 days (day 24-28), the occurrence of the first ovulation, the increase in capacity to form 13,14-dihydro-PGF2 alpha and PGF2 alpha levels induced by PMSG (5 IU) injected on day 26 were clearly inhibited on day 29. Danazol also markedly suppressed the LH surge occurring on day 28. Although the danazol-induced blockage of ovulation was restored by injection of human chorionic gonadotropin, the number of oocytes was significantly decreased as compared with that of controls. The present data indicate that the inhibitory actions of danazol on ovulation and ovarian PGF2 alpha metabolism may occur via some direct effects on the ovary in addition to the suppression of gonadotropin release from the pituitary gland.

In our previous study, a drastic change in terminal saccharides of glycoconjugates of the hamster zona pellucida associated with oocyte maturation was observed using light microscopic methods of lectin cytochemistry. To understand the mechanism of this change, in the present study, the correlation between the cytochemical appearance of saccharide residues in the zona pellucida and nuclear maturation was examined. Immature hamsters were treated with PMSG and hCG to induce follicular development and ovulation. The animals were euthanized 0 to 26 hrs. after the injection of PMSG or 0,1,2,3,4,5,7,9 or 11 hrs. after the injection of hCG, and ovaries were dissected out, fixed, paraffin embedded and sectioned serially. Every other paraffin section was stained with hematoxylin to observe the status of nuclei and to classify follicular growth and only the fully developed preovulatory follicles were examined in experiments. The peroxidase-labelled lectin-diaminobenzidine procedure was applied to sections. The lectins employed were WGA, SBA, MPA, UEA-I, LotusA and AAA. Germinal vesicle breakdown was observed within 3 hrs. after the administration of hCG. A positive reaction of WGA, SBA or MPA for zonae pellucidae in the fully developed preovulatory follicles appeared 1 hr. after hCG injection, and remained so for the next 10 hrs. UEA-I, Lotus A and AAA reactions were negative for all of the zonae pellucidae observed. The data indicate that the synthesis of saccharide residues such as GlcNAc and GalNAc forming zona components in the follicles is not triggered by germinal vesicle breakdown.

Present investigation was conducted to study the ovarian response and embryo recovery using different PMSG dose levels. Six rabbits each were assigned randomly to treatment 1 (PMSG 50 IU + hCG 100 IU), treatment 2 (PMSG 75 IU + hCG 100 IU) and the control group (no hormone administered). PMSG injection (im) was followed 68 hr later by natural mating to a fertile rabbit buck and human chorionic gonadotrophin (hCG) injection iv post coitum. Embryos were recovered 96 hr post coitum by a modified surgical method. Mean number of ovulations in the control group differed significantly from treatment 1 and 2, but no significant difference was observed between treatments 1 and 2. Mean embryo recovery percentage was lowest in treatment 2 and highest in the control group. The higher dose PMSG (treatment 2) was observed to be more disturbing in terms of recovery of embryos as well as their morphology.

The effects of application of differentiated PMSG rates (Antex Leo, Denmark) were investigated in 30 ewes of the Slovak Merino breed in the period of May--June in experimental conditions on the induction of superovulation and on the speed of egg transport through the sexual organs. After administration of PMSG at rates of 750, 1000 and 1500 i. u. per head/day, increases were recorded in the average weight of ovaries, number of large follicles and ovulations. Flushing the eggs from different parts of the sexual organs indicated that the higher the PMSG rate, the faster was the egg transport.

The morphological changes were examined in the present study in mouse cumulus-oocyte complexes and granulosa cells induced by the administration of pregnant mare serum gonadotropin (PMSG) and human chorionic gonadotropin (hCG). Dissolution of the germinal vesicle (GVBD) occurred in oocytes at 3 hr after hCG administration. Partial expansion of the cumulus cell investment was observed in the cumulus-oocyte complexes, especially those with the oocytes having undergone GVBD. Cumulus expansion due to the deposition of intercellular materials stained with colloidal iron proceeded up to 8 hr after hCG administration. The number of cells in the follicles containing matured oocytes is smaller than that in the follicles containing intact germinal vesicles in the granulosa cell layer, which indicates that the dispersion of cells occurs during meiotic maturation not only in the cumulus-oocyte complexes but also in the granulosa cell layer. Examination of the expansion of mouse cumulus-oocyte complexes by electron microscopy revealed the abundant deposition of intercellular materials and the retraction of the cytoplasmic processes joining the cumulus cells to the oocyte at 3 hr after hCG administration.

The effect of aromatizable androgens (testosterone and androstenedione) and naturally occurring 5 alpha-androstane, 3 alpha 17 beta-diol and 5 alpha-androstane, -3 beta, 17 beta-diol on oestradiol secretion by granulosa cells isolated from preovulatory follicles of PMSG-primed immature rats was investigated. The amount of oestradiol secreted by granulosa cells in the absence of exogenous aromatizable androgen in a 4h incubation was negligible. However, the addition of testosterone or androstenedione resulted in concentration dependent increases in oestradiol secretion. The 5 alpha-reduced androgens inhibit oestradiol and oFSH-stimulated oestradiol secretion by the granulosa cells in the presence of exogenous testosterone. The least potent of the androgens tested in causing this effect being the 5 alpha-androstane-3 alpha, 17 beta-diol. This result suggests that the naturally occurring 5 alpha-reduced androgens have a direct effect on androgen-aromatizing enzymes. The effect of these androgens may have an important connotation with respect to the control of the onset of puberty and regulation of ovarian oestradiol secretion within the microenvironment of an ovarian follicle.

An account is given in this paper of organometric and histometric findings obtained, on three farms, N, B, and D, from ovaries, uteri, and oviducts of biotechnologically treated gilts and adult sows, using differentiated PMSG doses (600, 800, and 1,000 IU on gilts and adult sows of N and B; 500, 1,000, and 1,500 IU on gilts of D). Ovulation potentials were within the biological normal in response to low dosage (with an average of 12 to 15 follicles in gilts and 17 in adult sows). The 800 IU dose caused significant stimulation, which had to be interpreted as overstimulation for PMSG-sensitive probands of N. Ovarian reaction and induced cycle should by duly considered for interpretation of histometric findings.

In twenty ewes of the Slovak Merino breed coming from a demonstration farm at Zemplínska Teplica, to the age of two-three years biometrical variations of the sex organs and overall follicular response to PMSG and PGF2 alpha administration were investigated in the autumn period (October-November). In the ewes of all groups the heat was synchronized by i. m. administration of PGF2 alpha (Oestrophan Spofa) at a dose of 125 micrograms in the interval of 11 days. On the ninth day the ewes of the second and fourth group were given 1000 i. u. of PMSG (Antex Leo, Denmark), the ewes of the third group 750 i. u. of PMSG. The ewes of the third and fourth group were administered at the same time 50,000 i. u. vitamin A (Axerophtol Spofa) (each group). The weight and dimensions of the sex organs were investigated. The results demonstrate that the administration of PMSG in the mating period increases significantly the weight of oviducts and horns of uterus in the ewes while there are no variations of the weight of the other sex organs and of their length, or it is lower than in the ewes of the control group. The overall follicular response was not influenced by the higher weight of ovaries.

One single PMSG batch, Pregmagon-Dessau, was compared with other PMSG doses on 2,126 adult sows and 1,700 gilts from five farms. Additional comparisons were made with other Pregmagon or Prolosan batches on 1,178 adult sows and 1,026 gilts. All experiments were made in the context of a programme for ovulation synchronisation and term-oriented insemination. Doses of 700, 800, 900, and 1,000 IU were tested on adult sows and 600, 800, and 1,00 on gilts, with the mean dose of 800IU being found to be sufficient or superior to other doses for fertility results. Doses for adult sows could be adjusted and related to present and previous litter numbers, with the dosage being between 700IU (treatment for third litter) and 700 to 800IU (treatment following average to high previous litter. The preferential dose for gilts was found to be 800IU. Higher PMSG doses (up to 1,000IU) should not be used in synchronisation of ovulation and term-oriented insemination unless extraordinary conditions were safely established.

Treatment was applied, on two farms, to 272 gilts and 169 adult sows, 24 hours after discontinuation of Suisynchron application to the gilts or weaning of piglets of adult sows. Included were 600, 800 or 1,000 IU of PMSG for the gilts and 750, 1,000 oder 1,250 IU of the same batch (Pregmagon-Dessau) for the adult sows for ovarian stimulation. This was followed by application of 500 IU of HCG to the gilts or 300 micrograms Gn-RH vet. "Berlin-Chemie" + 300 IU HCG to the adult sows for synchronised ovulation. Laparotomy was performed on the gilts on the fifth day from PMSG application and on the adult sows on the fourth day. Average ovariotropic stimulation was "mild" in response to lower doses (14.8 follicles in gilts and 20.4 follicles in adult sows) but was "pronounced" in response to medium doses (19.4 or 22.7 follicles, respectively). The pharmacological concept of "mild" exogenic ovarian stimulation by low dosage at the threshold of effectiveness should deserve more attention, last but not least, for breeding aspects. The two groups of animals on both farms differed considerably from one another with regard to reactivity, which cannot be etiologically interpreted for the time being.

Uptake of L-alanine, L-lysine, and choline into both preantral and antral mouse oocytes was enhanced by follicular cells. Follicular cells also enhanced glycine uptake into oocytes at the preantral stage of development, but no effect of these cells was observed at the antral stage. Glycine uptake was predominantly Na+ dependent and inhibited almost completely by 10 mM sarcosine, moderately by proline and its analog pipecolate, and poorly or not at all by other amino acids. By these criteria, glycine transport was mainly via system Gly in follicular cells and the oolemma at both the preantral and antral stages. Moreover, an increase in glycine transport via the oolemma between the preantral and antral stages was more than threefold larger than was the increase in transport of alanine or lysine. This relatively large increase in glycine-specific transport in the oolemma appears to obscure the ability of follicular cells to enhance glycine uptake into antral oocytes. In contrast to other amino acids, leucine uptake into oocytes was not enhanced by follicular cells unless 14 other amino acids were also present at their concentrations in mouse serum. An inhibitor of gap junctional communication, 18-alpha-glycyrrhetinic acid, abolished follicular cell-enhanced uptake of glycine and choline into preantral oocytes. Therefore, the extent to which follicular cells enhance uptake of a particular amino acid into oocytes depends on at least three physiologically important variables. Namely, enhancement may depend on the stage of follicular development, the presence of other amino acids in the environment, and gap junctional communication.

Ultrasonographic assessment of ovarian follicular maturity was reportedly associated with atypically early ovulation in women; related studies reported reduced litter sizes in rats. To confirm these findings, mice which were midway between ovulatory gonadotropin (LH or human chorionic gonadotropin) stimulation and ovulation, were sham- or ultrasound-treated periovarially for 5 min. Exposure was at a spatial average, temporal maximum intensity of 60 W/cm2. Carrier frequency in the pulse was 2.2 MHz, pulse length was 10 microseconds, and pulse repetition frequency was 200 Hz. Spatial average, temporal average intensity was 0.12 W/cm2. At autopsy, ultrasound- and sham-treated groups responded similarly in proportions ovulating and in mean ova ovulated. Combined experiments had a 97% chance of detecting a significant (greater than 1 h) advance in ovulation time, had it occurred. Thus, our adequately sensitive mammalian ovulatory tests revealed no association of ultrasound with decrease in ovum number or acceleration in ovulation time (as reported in humans).

A new procedure for studying the effect of various drugs on ovulation in vivo is described. In golden hamsters (Mesocricetus auratus), each ovary is enclosed within a complete bursa that is continuous with the oviduct. Drugs can be applied topically to ovaries by injection of a solution into a bursal cavity of an anesthetized hamster several hours before ovulation; the contralateral ovary serves as an in vivo control and receives no treatment. After ovulation, the number of ruptured follicles on experimental and control ovaries are compared. Data presented show that: (1) the operation and injection procedure per se do not affect ovulation; (2) normal saline is a suitable vehicle for administration of drugs by this route; (3) compounds with molecular weights of less than 1,000 daltons rapidly penetrate all layers of the follicle wall; (4) solutions injected into the bursal cavity 6 hours before ovulation remain there until ovulation. The advantages and limitations of this technique are discussed.

An attempt of granulosa cell tumor (GCT) induction by prolonged administration of exogenous pregnant mare's serum gonadotropin (PMSG) to young, mature, and middle-aged Balb-C mice resulted mainly in stromal luteal cell proliferation. 48.9% of young mice, 64.5% of mature mice and 65% of middle-aged mice developed luteal cell proliferation. This effect seemed to be dependent on duration of treatment, mainly in young mice. Young mice injected for less than 6 months developed significantly less luteal cell proliferation than those injected for more than 6 months. Only 1 GCT was found. This study seems to support the contention that besides gonadotropin stimulation, other factors may be involved in GCT development. A large number of PMSG-injected mice developed generalized lymphosarcoma, the mechanism of which remains unknown.

Immature rats can be induced to ovulate with pregnant mare serum (PMS) as long as they weigh over 60 g. In the rats weighing less than 60 g 5-hydroxytryptamine (5HT)-60 micrograms/rat intraventricularly or 2 micrograms/rat in the paraventricular nucleus (PVN)-stimulated ovulation. Injections into several other hypothalamic nuclei were ineffective. In rats weighing greater than 60 g, lesions of the PVN or injection of 10 micrograms/rat p-chlorophenylalanine into the PVN inhibited ovulation. The hypothalamic levels of 5HT and 5-hydroxy indole acetic acid (5HIAA) were generally lower in the afternoon and evening in the less than 60 g when compared with the greater than 60 g rats. It is possible that increased 5HT activity is required to stimulate ovulation and that its site of action is the PVN.

A field experiment was conducted to examine the effect of anti-oestradiol-17B antibody titre on the oestrous and ovulatory responses of ewes to low (600 i.u.) or high (1200 i.u.) doses of pregnant mare's serum gonadotrophin (PMSG). Merino ewes were treated with intravaginal sponges and were subsequently used as vehicle-treated controls or were immunized to produce reciprocal anti-oestradiol-17B antibody titres less than 1000 or greater than 1000. Ewes were then treated with PMSG and the incidence of oestrus and ovulation, ovulation rate, and yield of embryos recorded. Treatment of immune ewes with 1200 i.u. PMSG resulted in both a higher proportion of ewes ovulating and a higher ovulation rate than in immune ewes treated with 600 i.u. (86% v. 67% and 13.4 v. 6.0 respectively). As anti-oestradiol-17B titres increased there was a reduction in the proportion of ewes exhibiting oestrus. The proportion of ewes ovulating decreased as antibody increased in ewes treated with 600 i.u. PMSG but not in those treated with 1200 i.u., suggesting an increased positive feedback of oestradiol with high PMSG doses. Fertilization rates were highest at the lower PMSG dose (68% v. 42%) and increased with increasing titre. Overall, there was no increase in ovulation rate or in yield of embryos over control values from either low (less than 1000) or high (greater than 1000) antibody titres.

44 prepuberal gilts were treated with gonadotropic combinations and investigated for ovarian dynamics. 400 PMS + 200 HCG (Suigonan-Vemie) induced ovulations at the 4th day p.i., 100 FSH + 100 HCG resp. 200 FSH + 200 HCG at the 6th day p.i. At the 8,-11th day p.i. the PMS/HCG-treated animals showed corp. lut, in 100, the FSH/HCG-treated in 50 resp. 87% of the cases. A second injection of 250 HCG 3 days after 200 FSH + 200 HCG increased the number of animals which had ovulated. Zystic ovaries (larger than or equal to 11 mm) developed in all groups. Declaration of gonadotropins in "international units".

Human chorionic gonadotropin (HCG) may be injected to time ovulation and plan oocyte retrieval for clinical in vitro fertilization (IVF). Neither clinical nor experimental data on effects of subtle alterations in timing of the HCG injection on oocyte fertilizability in vitro were available. We induced follicular development in immature hybrid mice with an injection (4 IU) of pregnant mare serum gonadotropin (PMSG). In the first series of experiments, HCG was given 42, 46 or 50 hours later. We collected cumulus-enclosed, oviducal oocytes for IVF using capacitated mouse sperm 13 hours after the last HCG injection. The fertilization incidence (mean, three experiments) fell as the PMSG-HCG interval was reduced (50 hours, 64%; 46 hours, 40%; 42 hours 24%), but this could be explained by a corresponding increase in spontaneous oocyte activation caused by prolonging the HCG-oocyte collection interval. In the second series of experiments, the latter was fixed at 13 hours; the PMSG interval was altered by staggering the initial PMSG injection. No spontaneous activation occurred, but oocytes collected after "early" HCG injection still showed significantly lower fertilization incidences (42 hours, 36%; 46 hours, 46%) compared with the 50-hour injection (66%). The possible clinical implication of this finding is discussed.