Ovarian and placental production of progesterone and oestradiol during pregnancy in reindeer.

We obtained uterine and peripheral venous plasma, and samples of luteal and placental tissues from 2- to 7-year-old, Eurasian mountain reindeer (Rangifer tarandus tarandus) from a free-living, semi-domesticated herd in northern Norway in November 1995, and February and March 1996. In November, ovarian venous blood was also collected from four animals. Plasma samples were assayed for progesterone and oestradiol. The tissue samples were examined by light and electron microscopy, steroid dehydrogenase histochemistry, and northern blot analysis for RNAs for 3beta-hydroxy-steroid dehydrogenase (3beta-HSD) and P450 (side chain cleavage (scc)). Peripheral blood was taken from non-pregnant females in the same herd on the same dates. Peripheral progesterone concentrations in pregnant reindeer (3.4 +/- 0.5 ng/ml, n = 8) clearly exceeded those in non-pregnant animals (0.40 +/- 0.14 ng/ml; P < 0.0004 , n = 10) but oestradiol levels were only marginally higher in pregnant (6.0 +/- 0.7 pg/ml) than in non-pregnant (4.8 +/- 0.5 pg/ml; P = 0.35) reindeer at the stages examined. In pregnant animals, peripheral progesterone and oestradiol concentrations rose slightly between November and March but the differences did not reach significance (progesterone, P = 0.083; oestradiol, P = 0.061). In November, progesterone concentrations in the ovarian vein (79 +/- 15 ng/ml) greatly exceeded (P < 0.03) those in the uterine vein ( 10 +/- 4 ng/ml) which in turn exceeded the levels in the peripheral blood (2.8 +/- 0.4 ng/ml; P < 0.29). Oestradiol concentrations were slightly but significantly (P < 0.05) higher in the ovarian (20 +/- 3 pg/ml) than the uterine vein (13 +/- 1 pg/ml) and, in turn, greater (P < 0.03) than in peripheral blood (4.6 +/- 0.4 pg/ml). All samples of luteal tissue consisted exclusively of normal fully-differentiated cells and stained intensely for 3beta-HSD. Isolated groups of placental cells also stained strongly for 3beta-HSD. RNA for P450 (scc) and 3beta-HSD was abundant in all corpora lutea and lower concentrations of P450 (scc) were present in the placenta. 3beta-HSD RNA in the placenta was below the limit of detection. We conclude that the corpus luteum remains an important source of progesterone throughout pregnancy in reindeer but that the placenta is also steroidogenic.

Trace mineral and vitamin concentrations in the liver and serum of wild muskoxen from Victoria Island.

Selected trace minerals and vitamins were assayed in the liver and serum of 25 wild muskoxen (Ovibos moschatus) from Victoria Island, (Nunavut, Canada) in November, 1995. Mean +/- SE liver concentrations in micromol/kg wet weight were 260+/-16 for copper; 1.04+/-0.06 for selenium; 11.5+/-0.7 for molybdenum and 62.8+/-3.3 for vitamin E. Mean +/- SE serum concentrations in micromol/L were 14.2+/-0.3 for copper; 0.75+/-0.04 for selenium, 1.53+/-0.07 for vitamin A and 5.80+/-0.55 for vitamin E. Comparison of liver and serum concentrations of copper, selenium and vitamin E showed that the concentration in one tissue was a relatively poor indicator of the concentration in the other. The copper-molybdenum interaction often seen in domestic species was not observed. In general, the concentrations of metals and vitamins found in muskoxen were comparable to those in other ungulates although serum vitamin E concentrations were about one-fourth of those expected.

Regulation of steroidogenesis in jc-410, a stable cell line of porcine granulosa origin.

We investigated the regulation of steroidogenesis in a cell line of porcine granulosa origin, JC-410. Cells responded to the protein kinase-A activators, cholera toxin and forskolin, with increased accumulation of intracellular cAMP. Histochemically, cells were shown to contain 3beta-HSD, the enzyme which converts pregnenolone to progesterone. The JC-410 cells produced progesterone and responded to the protein kinase-A activators with an increase in progesterone synthesis. Progesterone levels were also increased by 25-hydroxycholesterol, pregnenolone, estradiol and androstenedione. Follicle-stimulating hormone and luteinizing hormone had no effect on cAMP or progesterone accumulation. Androstenedione was required for the synthesis of estradiol by JC-410 cells. Steady-state levels of mRNA for the steroidogenic enzymes 3beta-HSD and P450scc were increased by treatment with cholera toxin, whereas P450arom was not changed. These cells express the steroidogenic enzymes genes in a similar fashion to primary cultures of porcine granulosa cells. The JC-410 cells may represent a valuable model to study second messenger regulation and the molecular mechanisms involved in steroidogenesis in granulosa cells.

Copper deficiency in a herd of captive muskoxen.

At necropsy, a mature muskox cow was found to have exceedingly low serum and liver copper concentrations of 4.8 = mumol/L and 0.02 mmol/kg, respectively. Serum copper levels were also low in remaining members of the herd but returned to normal after parenteral treatment with calcium copper edetate.

The influence of fatness on the likelihood of early-winter pregnancy in muskoxen (Ovibos moschatus).

Among wild ruminants, muskoxen have an exceptional ability to fatten, but their pregnancy rates are variable and often low. To test whether the likelihood of pregnancy in muskoxen is associated with exceptionally good body condition, we used logistic regression analysis with data from 32 pregnant and 18 nonpregnant muskoxen > or = 1.5 yr of age shot in November (1989 to 1992) on Victoria Island in Arctic Canada. We assayed their serum for insulin-like growth factor-1 (IGF-1). All fatness and mass measures were positively related to the likelihood of pregnancy (P < 0.001), with the strongest associations for estimated total fat mass (80% of outcomes predicted correctly) and kidney fat mass (77%), and weaker models for body mass. Pregnancy was less likely to occur in lactating females than in nonlactating ones (P = 0.03). Although IGF-1 concentrations were higher (P = 0.001) in nonlactating females than in lactating ones (28.7 +/- 1.7 vs. 22.5 ng/ml), no association with pregnancy was detected (P = 0.57). Fatness associated with a 50% probability of pregnancy in muskoxen (22% of ingesta-free body mass or 32 kg fat in females > 3.5 yr old) is much higher than in caribou and somewhat higher than in cattle, and this may partly account for the low calving rates often observed in this species.

Fetal growth in muskoxen determined by transabdominal ultrasonography.

A 5 MHz commercial sector scanner was used to monitor 13 muskox pregnancies and establish normal fetal growth curves. Examinations were carried out between 40 and 197 days of gestation and pregnancy could be detected throughout the period. Early pregnancies were found by scanning lateral to the udder but as pregnancy progressed the fetus was found closer to the dam's umbilicus. Measurements of cranial and abdominal diameters taken at about two week intervals in seven uncomplicated pregnancies in four cows were used to construct fetal growth curves. These can be reliably used in the reproductive management of muskoxen. In addition a series of regressions based on measurements of the fetuses of muskoxen killed in the Arctic are provided. These allow cranial and abdominal diameters to be related to fetal weight and crown-rump length.

Endocrine changes and luteal morphology during pregnancy in muskoxen (Ovibos moschatus).

Progesterone, oestradiol and oestrone were measured in plasma from four captive muskoxen during three consecutive pregnancies (1983-1984, 1984-1985 and 1985-1986). Jugular blood samples were collected weekly (1983) or on an alternating 3:4 day schedule (1984-1986) during the first 12-15 weeks and last 6-10 weeks of pregnancy. Sampling during mid-pregnancy was at intervals of 2 weeks (1983 and 1985) or 1 week (1986). Duration of gestation was about 34 weeks (235 +/- 4 (SD) days (n = 10), range 230-242 days). Progesterone remained at concentrations similar to those found during the luteal phase of the oestrous cycle for the first 10-12 weeks (mean +/- SEM 1.6 +/- 0.1 ng ml-1) after which it rose to a peak (mean 5.5 +/- 0.65 ng ml-1) between weeks 12 and 20. In all ten pregnancies progesterone concentrations declined dramatically between weeks 20 and 22 to luteal phase values where they remained until parturition. The decline was accompanied by an increase in oestradiol and oestrone concentrations which reached mean peak values of 199.23 +/- 87.23 pg ml-1 and 980.48 +/- 203.91 pg ml-1, respectively. Corpora lutea collected from wild muskoxen between 45 and 80 days gestation all showed histological evidence of regression, while corpora lutea from mid-gestation (112-125 days) were in advanced stages of involution. Repeated ovarian ultrasonography of captive muskoxen during the first 100 days of pregnancy confirmed these findings. The unusual, early regression of the corpus luteum of pregnancy indicates that progesterone and oestrogen of mid- and late pregnancy are probably of placental origin.(ABSTRACT TRUNCATED AT 250 WORDS)

Odor of the muskox : A preliminary investigation.

The behavior of captive male muskoxen was observed closely during their characteristic superiority display, the anatomy of the preputial region was studied in two adults and three calves, and preputial washings and preorbital gland secretion were subjected to gas chromatography and mass spectroscopy. During the superiority display, the prepuce was everted to form a pendulous tube tipped with a fringe of matted hair. Owing to the movement of the animal, the urine that dribbled from the preputial opening was liberally applied to the long guard hairs of the belly. The superiority display was almost exclusively confined to dominant males and apparently accounted for their odor. In the quiescent state, the hair seen around the preputial opening was drawn inside and formed an 8 cm-wide band on the lining of the prepuce. The preputial washings contained large amounts of benzoic acid andp-cresol. The infraorbital gland secretion contained cholesterol, benzaldehyde, and a homologous series of saturated γ-lactones ranging from 8 to 12 carbons. The latter compounds and the natural secretion smell similar to the human nose.

Progesterone, oestradiol-17 beta and LH during the oestrous cycle of muskoxen (Ovibos moschatus).

Progesterone, oestradiol-17 beta and LH were measured in plasma from 6 non-pregnant, captive, female muskoxen during the 1984 and 1985 breeding seasons. Jugular blood samples were taken on an alternating 3/4-day schedule in 1984 and daily or at 4-h intervals over oestrus, via indwelling jugular cannulae, for 6 weeks in 1985. Oestrous cycle length was 19.6 +/- 0.96 (s.d.) days (n = 19) and did not vary between the first and subsequent cycles of the season. Progesterone was lowest at oestrus (less than or equal to 0.1 ng/ml), began to rise on Days 4-5, peaked on Days 10-12 (mean = 2.6 ng/ml) and returned to baseline 2-5 days before the next oestrus. A small rise in progesterone before the first cycle of the breeding season was observed on 7 of 12 occasions. Oestradiol-17 beta was significantly higher (P less than 0.001) 1-4 days before, or coincident with, oestrus. The average duration of the LH peak was 24.6 h (n = 7) and coincided with observations of behavioural oestrus. In one animal behavioural oestrus and an LH peak preceded a small progesterone rise at the beginning of the breeding season. The temporal relationship of these three hormones during the muskox oestrous cycle is very similar to that seen in domestic ruminants.

Genetic and temporal variation in serum concentrations and biological activity of horse chorionic gonadotrophin.

The variation in the quantity of circulating chorionic gonadotrophin (CG) and its follicle-stimulating hormone (FSH) and luteinizing hormone (LH) activity in rodent bioassay systems was investigated. A portion of the variability in total CG could be attributed to the stallion that sired the pregnancy and it was possible to select sires and mares to increase CG production. It was further demonstrated that FSH activity per unit of CG was greater at Days 71 and 104 of gestation than at Day 39. LH activity per unit of CG varied with the sire, but no effect of day of gestation could be shown. It was demonstrated that removal of sialic acid increased LH bioactivity and it is proposed that variation in biological activity between animals and during gestation may be a function of differences in carbohydrate content of CG.

Development of horse embryos up to twenty two days after ovulation: observations on fresh specimens.

Forty nine embryos, twenty unfertilized eggs and five other fresh eggs of 'doubtful' status have been recovered from 58 pony mares in 122 flushes up to 22 days after ovulation. The fresh egg or embryo recovery rate was 78% with surgical methods (or at slaughter) and 40-60% with non-surgical methods of recovery. The fertilization rate was about 70%. It has been confirmed that horse embryos normally enter the uterus as blastocysts 5-6 days after ovulation. Three features of early embryo morphology have become clearer upon comparison with unfertilized eggs of similar ages; early embryos are often ellipsoidal in shape; dispersal of most of a thick gel coat seems to be hastened by fertilization; gradual disappearance of refractile granules from the perivitelline space is similar in fertilized and unfertilized eggs. A tense, transparent, acellular capsule (considered to be different from the zona pellucida) is acquired by the spherical blastocysts within the uterus and persists at least until a diameter of 34 mm is attained (at 21 days in the present series). The capsule seems to be analogous, in part, with the 'neozona' described in rabbit blastocyst before attachment, and trophoblastic cells appear to be involved in its formation. Cleavage stages of oviductal embryos and diameters of uterine blastocysts from this series have been described and illustrated and used to extend previous knowledge of early growth patterns in horse embryos.

Transmission electron microscopy of horse embryos 3-16 days after ovulation.

The 23 embryos were obtained by flushing the reproductive tract. Though the general cytology was observed, most attention was given to the formation of the embryonic capsule. It first appeared as a thin uniform layer on the inner surface of the zona pellucida of embryos recovered from the uterus on Day 6. By Day 8 the capsule was about 1 micron thick and the zona pellucida had been shed. In fixed embryos of 11 days and over the capsule was 3 microns thick and had a finely stippled but otherwise homogeneous appearance.

Hydroxysteroid dehydrogenases in the ovine endometrium and trophoblast.

A histochemically demonstrable 3 beta-hydroxysteroid dehydrogenase appears in the uterine epithelium of the pregnant ewe on Day 12 and persists until after parturition. It is either very weak or absent in the non-pregnant ewe. A similar enzyme can just be seen in the trophoblast on Day 18, increases in strength until Day 33 and persists to term. The appearance of the endometrial enzyme coincides with events associated with maternal recognition of pregnancy.

The location of eggs retained in the oviducts of mares.

The oviducts of 24 mares were examined to determine the site of retention of unfertilized eggs. The ampullary-isthmic junction regions of 42 of the 48 oviducts were serially sectioned and examined histologically. The remaining parts of the oviducts were flushed and the flushings searched microscopically. Of 45 eggs located, 40 were in the sectioned segments of 24 oviducts and only 5 were in the flushings. All but one of the sectioned segments contained prominent masses of material obstructing the lumen, but these were apparently not the direct cause of egg retention since eggs were found on both sides of them.

Embryo transport through the mare's oviduct depends upon cleavage and is independent of the ipsilateral corpus luteum.

Two experiments were conducted using 14 mares. In Exp. 1, mares were inseminated with semen treated with TEPA, which, in other species, has been shown to lead to an arrest in ovum cleavage at 2--4 cells. The oviducts and/or uterus were then flushed 7--10 days after ovulation in 6 mares (Group A) or 2--6 days after ovulation in 5 mares (Group B). Fresh eggs were found in the oviduct flushes of 5 Group A and 5 Group B mares: 9 of the 10 eggs appeared to have cleaved, but none had developed beyond 16-cells. Seven eggs contained spermatozoa and 3 of 4 eggs from each group showed evidence of fertilization when examined ultrastructurally. Group A mares had thus retained fertilized eggs in the oviduct beyond the time at which they would normally have entered the uterus (6 days), indicating that development beyond at least the 2- to 4-cell stage is necessary for normal transport. In Exp. 2, 5 attempts were made to recover the embryo within 4 days of ovulation and transfer it to the contralateral oviduct. A single pregnancy resulted, indicating that a unilateral interaction with the corpus luteum was not necessary for the transport of the embryo to the uterus.

Oestrogens and androgens in blastocoelic fluid and cultures of cells from equine conceptuses of 10-22 days gestation.

Six samples of blastocoele fluid recovered between 10 and 22 days gestation were tested in human clinical radioimmunoassay systems measuring total oestrogens and total androgens. The results were erratic but in 5 cases measurements for oestrogen equivalent to between 1000 and 70,000 pg/ml and for androgen between 1000 and 85,000 pg/ml were recorded. Cells from two blastocysts were cultured in medium 199 with and without horse serum. When the used media were assayed, values equivalent to at least 8000 pg oestrogen/ml were obtained on 7 of 11 occasions. In 9 of 11 samples the androgen concentration exceeded 700 pg/ml. In control media the oestrogen and androgen concentrations did not exceed 120 and 360 pg/ml respectively.

Embryological studies on the dartmoor pony during the first third of gestation.

A sequence of ten known-age embryos recovered from Dartmoor Ponies during the first 4 months of gestation is described. Changes in size, shape and vascularity of the conceptus are recorded, as well as the growth and decline of the yolk-sac and the succeeding establishment of the allantochorion. The progress of equine somatogenesis is compared with that of the pig which it lags on average of about 4 days. Some quantitative data are presented.