Summary. Normally cyclic heifers received 2500 i.u. PMSG i.m. at Day 10 of the oestrous cycle and 15 mg prostaglandin (PG) i.m. 48 h later. From 30 h after PG the LH concentration in the peripheral blood was estimated every hour using a rapid RIA method which allowed the LH concentration to be known within 4 h. Monoclonal antibody against PMSG was injected in the jugular vein of 29 heifers at 4·8 h after the maximum of the preovulatory LH peak; 28 heifers were not treated with anti-PMSG (controls). Peripheral blood concentrations of PMSG, LH, progesterone and oestradiol were compared. Ovaries were collected by ovariectomy at fixed times, 22–30 h after the LH peak, and numbers were counted of small (2–10 mm), large (> 10 mm) and ovulated follicles, and of follicles with a stigma.

In anti-PMSG-treated cows, the PMSG concentration fell sharply to non-detectable levels within 2 h of the treatment, indicating that PMSG was neutralized in these cows at the onset of final follicular maturation. In all cows, the concentration of oestradiol showed a significant decrease at about 8 h after the LH peak. After anti-PMSG treatment ovulations took place from 24 until 30 h after the LH peak, whereas in control cows follicles had already ovulated at or before 22 h and ovulations continued until 30 h. At 30 h 90% of the follicles had ovulated in anti-PMSG-treated cows vs 72% in the controls, resulting in 15 and 8 ovulations per cow respectively (P < 0·05). Also, administration of monoclonal antibody against PMSG synchronized final follicular maturation and shortened the period of multiple ovulations.

In conclusion, neutralization of PMSG shortly after the preovulatory LH peak suppresses adverse effects of PMSG on final follicular maturation, leading to an almost 2-fold increase of the ovulation rate.

The influence of luteolysis on myometrial activity in late pregnant cows was studied by measuring electromyographic (EMG) activity and concentrations of progesterone and 13,14-dihydro-15-keto-prostaglandin F_2α (PGFM) in maternal arterial plasma of five cows. Application of two ear implants containing progestagen at day 261 of gestation was followed by an injection of a luteolytic dose of a prostaglandin F_2α (PGF_2α) analogue (PG) on day 264. Calving was initiated by removal of the implants and a single injection of 5 mg flumethason on day 270. All calves were born alive at a mean interval of 36 h. After 4 h of hyperactivity immediately following the injection of PG, myometrial activity was almost completely absent for 20 h; before luteolysis the total duration of EMG activity was 7.21 ± 0.31 min h⁻¹ (mean ± sem), during the 4 h after PG 23.61 ± 3.40 min h⁻¹, and during inhibition 2.70 ± 0.87 min h⁻¹. After the last period, EMG activity recurred and reached values of 14.52 ± 3.07 and 11.17 ± 2.87 min h⁻¹, on days 265 and 269, respectively. Concentrations of progesterone in maternal plasma decreased from 4.17 to 1.14 ng ml⁻¹ within 12 h after PG, whereas concentrations of PGFM in plasma remained low, varying from 15 to 30 pg ml⁻¹. After removal of the implants and injection of flumethason on day 270, EMG activity increased without an inhibitory phase and reached maximum values around the time of expulsion of the calves. PGFM concentrations gradually increased to 40 pg ml⁻¹ at 16 h after removal of the implants and injection of flumethason; there was then a steep increase to mean concentrations of 200 pg ml⁻¹ during expulsion of the calf. These results indicate that a factor, released during luteolysis, exerts an inhibitory effect on the myometrium.

After synchronization of oestrus, normally cyclic heifers (n = 31) received 2500 iu pregnant mares' serum gonadotrophin (PMSG) i.m. and had a progesterone-releasing intravaginal device (PRID) without the oestradiol capsule inserted on day 10 of the oestrous cycle and received 15 mg prostaglandin (PG) i.m. 48 h later. PRIDs were removed 96 h after insertion and 16 heifers received 1.0 mg GnRH i.m. while the controls (n = 15) received 10 ml saline i.m. All heifers were injected with anti-PMSG i.v. 10 h later. Peripheral blood concentrations of PMSG, progesterone, oestradiol and LH were compared. Ovaries were collected on death 7 days after the GnRH or saline injection and the number of corpora lutea counted. Heifers were considered to have responded well (> 60 pmol l⁻¹) or poorly (< 60 pmol l⁻¹) to superovulation on the basis of the oestradiol concentration 24 h after PG administration. During PRID treatment, LH concentrations remained at basal values. In the heifers treated with GnRH, a single LH surge occurred 2.3 ± 0.1 h (sd) after the GnRH injection with a maximum concentration of 14.6 ± 2.3 (sem) μg l⁻¹ and a duration of 6–8 h. In 12 of the 15 control heifers, LH concentrations remained low (range 0.10–1.94 μg l⁻¹) during the 72 h following the saline injection; three controls showed a spontaneous LH surge at 18, 23 and 23 h after the saline injection, respectively, with a maximum concentration of 6.0–12.5 μg l⁻¹ and a duration of 10–12 h. The oestradiol concentration increased continuously during PRID treatment until the injection of GnRH or saline, when it was four times higher in the heifers that responded well than in the heifers that responded poorly. It decreased sharply 6 h after GnRH indicating that the follicles still responded normally to a preovulatory LH signal, whereas in control heifers a similar decrease took place 4 h later following anti-PMSG treatment. In the GnRH-treated heifers, the heifers responding well showed a significantly higher number of corpora lutea than did the animals showing a poor response, 16.4 ± 2.2 (n = 9) and 5.4 ± 1.4 (n = 7), respectively. Five of the 12 control heifers without an immediate LH surge showed a single corpus luteum, and seven heifers did not have a corpus luteum. For the three controls with an LH surge, 33.0 ± 8.5 corpora lutea were observed. In conclusion, the preovulatory LH signal can be effectively postponed in PMSG/PG-superovulated heifers using a PRID. However, the PRID treatment has to be followed by GnRH to obtain the LH surge at a defined time of preovulatory follicular development. Follicular function with regard to oestradiol secretion and the potential to ovulate remains unchanged.

Summary. Preovulatory bovine follices (n = 73) were collected at different times after the onset of oestrus until shortly before ovulation, which occurred at 24 ± 1 ·4 h after the peak concentration of LH in the peripheral blood. Non-atretic antral follicles (n = 9) of 15–19 mm were also collected from cows during the luteal phase of the oestrous cycle. Follicular fluid concentrations of dehydroepiandrosterone, androstenedione and oestrone, and of LH, FSH and prolactin were compared in 2-h periods relative to the LH plasma peak. Before the LH surge the concentrations of the steroids were much higher than in non-atretic luteal-phase follicles of similar size. From 0 to 6 h after the LH peak the steroid concentrations decreased sharply to remain low until ovulation; only that of androstenedione increased again after 14 h to remain constant. The ratio between the concentrations of androstenedione and dehydroepiandrosterone remained constant until 14 h after the LH peak; at 14 h it increased about 4-fold and remained high until ovulation. The ratio between the oestrone and androstenedione concentration increased gradually to a 10-fold higher value until at 14 h an abrupt decrease was observed. These changes indicate that after the LH peak androgen production is directly inhibited and, at a slower rate, the aromatizing activity. Androstenedione appeared to be the major aromatase substrate.

Before the plasma LH peak the follicular fluid concentration of FSH was higher than in luteal-phase follicles; the concentrations of LH and prolactin were not different from those in luteal-phase follicles. About 4 h after the LH plasma peak the LH concentration in follicular fluid reached a maximum, which was one seventh of that in peripheral blood; it remained elevated until 20 h. The FSH concentration was higher after the LH plasma peak than before; just before ovulation it was still 2-fold higher. The concentration of prolactin fluctuated throughout the preovulatory development from the onset of oestrus until ovulation. It is suggested that oestradiol biosynthesis in bovine preovulatory follicles is terminated by an inhibitory action of the preovulatory LH peak on androgen production.

Summary. Sequential protein synthesis and protein phosphorylation patterns were generated by radiolabelling bovine cumulus–oocyte complexes after various periods of culture with [³⁵S]methionine and [³²P]orthophosphate respectively. The radiolabelled oocytes were assessed for their nuclear status and used individually for gel electrophoresis. Marked changes in the protein synthesis patterns were observed exclusively after germinal vesicle breakdown (GVBD), whereas oocytes which remained in the germinal vesicle stage showed a consistent protein synthesis pattern. The changes were observed after 8 and 16 h of culture, shortly after GVBD and before first polar body extrusion. From 3 h of culture, dominant phosphoprotein bands with apparent molecular weights of 24 000 and two between 50 000 and 60 000 were observed. The latter bands displayed slight molecular weight changes, which were not closely time related. After GVBD, the phosphoprotein band with M _r 19 000 was no longer observed. This study demonstrates that specific changes in protein synthesis and protein phosphorylation are programmed during bovine oocyte maturation.

Keywords: cow; oocyte; maturation; protein synthesis; protein phosphorylation

Dose–response relationships between GnRH and LH, and between GnRH and testosterone, were investigated in six male dogs by intravenous administration of a GnRH analogue at different doses. Each dose of GnRH analogue induced an immediate rise in the plasma concentration of LH and then a rise in plasma testosterone concentration. Irrespective of the dose used, the rise in testosterone began 10 min after the GnRH injection. Administration of GnRH at doses of 0.01, 0.1, 1, 10 and 100 μg kg⁻¹ resulted in maximum LH concentrations in plasma (mean ± sem; n = 6) of 22 ± 7, 27 ± 6, 40 ± 7, 57 ± 13 and 56 ± 10 μg l⁻¹, respectively. These doses induced maximum concentrations of testosterone in plasma (mean ± sem; n = 6) of 16 ± 4, 20 ± 4, 22 ± 3, 22 ± 4 and 24 ± 7 nmol l⁻¹, respectively. The lag time between peak concentrations of LH and testosterone varied from 35 to 55 min. The calculated maximum response of testosterone to LH, secreted by the anterior pituitary after GnRH injection, was 1.8 times higher than to GnRH. It was concluded that intravenous administration of GnRH induced marked and dose-dependent increases in plasma concentrations of LH and testosterone, and that there does not appear to be a direct effect of GnRH on Leydig cells in male dogs.

Summary. Six heifers were injected i.m. with 2500 i.u. PMSG followed by 15 mg prostaglandin 48 h later. Serial blood samples were collected through a catheter in the caudal vena cava every 10 min for 8 h on Day 10 (7 h after PMSG administration), during luteal regression (7 h after prostaglandin administration) and on the day thereafter. Four normally cyclic heifers served as a control group. Concentrations of progesterone, androstenedione, oestradiol, LH, FSH, and PMSG in the vena cava samples were measured and the frequency and amplitudes of episodic pulses of all hormones were estimated except for PMSG. Ovaries were collected by ovariectomy at 50 h after onset of luteal regression to determine the number of preovulatory follicles (non-atretic follicles ⩾ 10 mm).

Stimulation of follicular growth by administration of PMSG resulted in the following effects on the secretion of steroids and endogenous gonadotrophins. (1) There were no alterations in progesterone concentration and the amplitude and frequency of episodic pulses. Mean (±s.e.m.) concentrations were 54·1 ± 5·8, 19·1 ± 3·1 and 3·4 ± 0·9 nmol/l on Day 10 (L), during luteal regression (LR) and on the day thereafter (F) respectively. (2) There were no alterations in the episodic secretion patterns of androstenedione. Mean concentrations were 0·20 ± 0·02, 0·15 ± 0·02 and 0·11 ± 0·02 nmol/l for the L, LR and F periods respectively. (3) There was an increase in oestradiol concentration from 17·1 ± 3·0 pmol/l during the L period to 233·7 ± 86·4 pmol/l during the F period. Pulse amplitude was enhanced compared to corresponding periods in control animals whereas pulse frequency remained the same. The oestradiol concentration was significantly correlated with the number of preovulatory follicles (r = 0·82, P < 0·05). (4) There was a suppression of the frequency of episodic LH pulses (/8 h) during the LR (3·2 ± 0·7) and F (4·3 ± 0·4) periods compared to corresponding periods in control heifers (9·5 ± 0·9 and 7·0 ± 1·5 respectively). The preovulatory LH peak occurred earlier in 4 of 6 treated heifers. (5) There was a suppression of FSH concentrations, pulse amplitude and frequency during the LR and F (17·4 ± 0·9 mg/l, 4·7 ± 0·8 μg/l and 7·5 ± 0·4 pulses/8 h) periods compared to the corresponding F-period values (35·6 ± 6·2 mg/l, 9·8 ± 1·6 μg/l and 9·3 ± 0·3 pulses/8 h) in control heifers.

It is concluded that the increased number of preovulatory follicles after PMSG administration severely suppresses endogenous FSH secretion by means of enhanced concentrations of oestradiol and probably also of inhibin.

Keywords: superovulation; PMSG; steroids; gonadotrophins; cow

Normally cyclic heifers (n = 34) received 2500 iu pregnant mares' serum gonadotrophin (PMSG) i.m. at day 10 of oestrus, and 15 mg prostaglandin (PG) i.m. at day 12. Thereafter, a monoclonal antibody against PMSG was administered i.v. before (n = 24), at (n = 6) or shortly after (n = 4) the preovulatory LH surge. Peripheral blood concentrations of LH and oestradiol were compared; follicular development was monitored by daily ultrasound scanning; and the numbers of preovulatory-sized follicles and ovulations were counted 96 h after injection of PG following death. Anti-PMSG treatment before the LH surge inhibited the LH surge in 16 heifers (67%). In these heifers, the initial increase in oestradiol concentration upon PMSG stimulation to 167.5 ± 35.0 pmol l⁻¹ was terminated immediately after anti-PMSG treatment and decreased rapidly to basal values, while the number of preovulatory-sized follicles remained constant until 68 h after PG injection; on average 0.4 ± 0.1 ovulations were counted. In the remaining eight heifers, five animals showed an immediate, but temporary, 20–60% drop in oestradiol concentration after anti-PMSG treatment. In all eight heifers 25% of the preovulatory-sized follicles ovulated. Treatment with anti-PMSG at or shortly after the LH surge did not affect the pattern of oestradiol concentration, but a significantly higher ovulation rate was observed in the animals treated shortly after the LH surge: 20.3 ± 2.6 versus 6.3 ± 2.3 in animals treated at the LH surge, which corresponded to 76% and 24% of the preovulatory-sized follicles monitored shortly before the period of multiple ovulations. Thus, neutralization of PMSG at any time before the maximum concentration of the preovulatory LH surge severely suppresses the functionality of the majority or all of the stimulated follicles, which is dependent on the time of injection of anti-PMSG. Although the follicles retain their size, they lose the potential to ovulate. It is concluded that interfollicular asynchrony of development is present at the time the LH surge occurs.

Summary. Five dogs were hypophysectomized on Day 4 and 9 on Day 18. Prolactin and LH stimulation tests showed that hypophysectomy was complete in 6 dogs only. In these dogs, the progesterone concentration was measured in the peripheral blood; it decreased sharply immediately after surgery. It regained normal values in 3 of the 4 dogs hypophysectomized on Day 4, and remained low in the 2 dogs hypophysectomized on Day 18. This indicates that, in the dog, luteal function is autonomous during a certain period. The luteal period of the 3 dogs hypophysectomized on Day 4 was shorter than that of control animals, although the time of onset of luteal regression appeared to be similar. This indicates that pituitary luteotrophic support is required during the second part of the oestrous cycle of the dog.

Summary. Pregnant gilts (3/group) were given no treatment, 10 mg bromocriptine twice daily by mouth, from Day 111 of pregnancy to 1 day post partum, 25 mg progesterone s.c. at 6-h intervals from Days 111 to 116 inclusive or 400 mg indomethacin by mouth at 6-h intervals from Day 111 to 116 inclusive. Before spontaneous delivery maternal plasma prolactin and relaxin concentrations started to rise almost simultaneously between 57 and 47 h before the first piglet and both hormones reached peak values when the plasma progesterone concentration had started to decline rapidly (~21–23 h). Suppression of prolactin levels by bromocriptine prevented the onset of lactation completely but had no obvious influence on changes of the other hormone concentrations and the course of parturition. Progesterone treatment delayed the onset of expulsion of the piglets but did not delay the simultaneous increase in prolactin and relaxin concentrations. These changes in hormone levels were prevented by indomethacin treatment but occurred essentially unchanged when the treatment was ended. The results support the concept that parturition in the pig is preceded by a biphasic increase of plasma prostaglandin levels.