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- Author: R. B. Heap x
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SUMMARY
A technique is described for the recovery of uterine washings from four species, the rat, rabbit, sheep and cow. The washings were recovered from intact animals at oestrus and during the luteal phase of the reproductive cycle. The amounts of several chemical constituents were measured in the washings (sodium, potassium, chloride, phosphorus, iron, copper, aluminium, magnesium, calcium, cobalt, manganese, carbohydrate, lipid) and the nitrogen distribution was determined. The method, which involved flushing the uterine horn with an isotonic solution, was evaluated in terms of the recovery of a given volume of flushing solution, the effect of uterine ligation, post-mortem changes in the levels of chemical constituents in the uterine washings, and changes in the amounts of the different constituents in utero during the flushing procedure. There was a significant difference in the amount of several of the constituents in uterine washings recovered at two stages of the reproductive cycle. The values were greater at oestrus than at dioestrus in the rat (Na, K, Cl, N, carbohydrate), but greater in the luteal phase than at oestrus in the rabbit, sheep and, to some extent, in the cow (K, P, N, carbohydrate). The application of the method and the interpretation of the results are discussed.
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SUMMARY
Steroid hormone concentrations in plasma have been measured in blood samples taken from conscious sows with ear vein catheters. In late pregnancy, the plasma progesterone concentration ranged from 6 to 12 ng/ml and it decreased in all animals before the onset of parturition. Total unconjugated oestrogens increased to high values of up to about 3 ng/ml in late pregnancy and then declined after the onset of parturition. Oestrone was the predominant unconjugated oestrogen measured. Plasma corticosteroid (mainly cortisol) concentration was about 33 ng/ml and showed no consistent change at the time of parturition.
During lactational anoestrum the plasma concentration of progesterone and total unconjugated oestrogens was very low, while that of corticosteroids was 21 ng/ml. When the piglets were weaned at 26–31 days, sows came into oestrus 4–12 days later, and this was preceded, or accompanied by, an increase in plasma oestrogens. In the luteal phase, plasma progesterone concentrations rose to 20–35 ng/ml. A sow whose piglets were removed at birth, showed signs of oestrus (vulval enlargement and a lordosis response), but a lack of receptivity to the boar associated with no detectable changes in the plasma oestrogen concentration; however, ovulation probably occurred since plasma progesterone values increased in a manner comparable to that found after the formation of normal corpora lutea in other sows. After a second non-receptive cycle, the sow was mated and became pregnant at the third post-weaning oestrus.
At parturition the concentration of progesterone and total unconjugated oestrogens was greater in placental venous plasma than in maternal jugular plasma, which indicates placental synthesis of these hormones. A greater concentration of plasma corticosteroids in foetal blood than in placental venous or maternal jugular plasma suggests foetal synthesis in late pregnancy.
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SUMMARY
Progesterone was isolated from systemic and uterine venous plasma and from placental tissue of intact and ovariectomized pregnant guinea-pigs. It was measured quantitatively by fluorescence assay.
Concentrations of progesterone in intact animals rose from a mean value of 64 ng./ml. plasma 11–16 days post coitum (p.c.) to reach their highest values (265 ng./ml.) 30–45 days p.c. and decreased to 140 ng./ml. immediately before parturition. In pregnant guinea-pigs, ovariectomized 28 days p.c. or later, the progesterone levels were lower (75 ng./ml. at 34–35 days p.c., 146 ng./ml. at 40–45 and 50–55 days p.c.) except at the end of pregnancy (165 ng./ml.). During the last 2 weeks of pregnancy the difference in the progesterone levels of the two groups was not statistically significant.
In ovariectomized pregnant guinea-pigs, the progesterone level was high in the uterine vein but low in placental tissue. The mean wet weight of placentae was, however, significantly correlated with systemic progesterone values.
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SUMMARY
Sites of production and uptake of progesterone were compared in acute experiments in two sheep and two goats 119–126 days pregnant. In the two goats progesterone was produced mainly by the ovaries (up to 10 mg./day, placenta 0 mg./day) whereas in the two sheep the placenta made the largest contribution (up to 14mg./day, ovaries about 2 mg./day). Adrenal production was less than 2% of the ovarian output except in one goat (20%). In four out of five foetuses studied, umbilical arterial concentrations of progesterone were higher than umbilical venous ones. Δ5-3β-Hydroxysteroid dehydrogenase was demonstrated histochemically in the foetal adrenals but not the ovaries.
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SUMMARY
Plasma progesterone concentration was measured in arterial, jugular, mammary venous and ovarian venous blood of 20 goats, together with measurements of mammary and ovarian blood flow.
The level in arterial plasma in conscious normal and in castrated males was less than 2·6 ng./ml.; in anoestrous and oestrous females it was 3·4 ± 0·8 (s.e.) ng./ml. In the luteal phase of the ovarian cycle the level rose to 10·7 ± 2·2 ng./ml. and during pregnancy it remained above 10 ng./ml. until near term. In a goat that aborted at 15 weeks of pregnancy no progesterone was detected 3 days before or at the time of abortion.
When an active corpus luteum was present in the ovaries, the mammary venous level of progesterone was lower than the arterial concentration in 38 out of 44 instances, the mean difference being 23 ± 4% of the arterial level. In two goats the concentration in jugular vein blood was 80 and 85% of the arterial level but the mammary venous levels were lower still (57 and 75 % of the arterial concentration, respectively). In three goats sampled repeatedly during the whole reproductive cycle, simultaneous measurement of mammary blood flow showed that during pregnancy the udder was taking up to 2–30 ng. progesterone/min./10 g. tissue from the plasma. In a fourth, much older goat (in early pregnancy, but not lactating), the uptake was consistently lower. In all four animals the arteriovenous difference was linearly related to the arterial plasma concentration (r = 0·84).
In ovarian venous blood, collected under spinal anaesthesia, the plasma concentration was not significantly different from that in peripheral blood during anoestrus and oestrus, but when active corpora lutea were present the concentration was 620–1600 ng./ml. Simultaneous measurement of ovarian venous outflow suggested that 9–10·5 mg. progesterone/day was being produced by the ovaries, and about 20 % of this was taken up by the udder.
Peripheral blood progesterone levels were unaffected by chasing the animal but were raised by spinal anaesthesia and were raised still further after laparotomy.
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The nitrogen distribution of uterine washings differs from that of blood plasma (Ringler, 1956; Junge & Blandau, 1958; Heap & Lamming, 1962). The acid-soluble nitrogen, soluble in an equal volume of 11% trichloracetic acid, represents a considerable proportion of the total nitrogen of the uterine washing in the rat at oestrus, in the rabbit during the luteal phase and in the cow during the latter half of the luteal phase but not to the same extent in the sheep (Heap, 1962; Heap & Lamming, 1962). The occurrence of the acid-soluble component was associated with a translucent appearance of the trichloracetic acid (TCA) extract of uterine washings. It was also extracted with lipid solvents. The levels of acid-soluble nitrogen and of total carbohydrate in uterine washings of the rat and rabbit showed a positive correlation (Heap, 1962; Heap & Lamming, 1962).
Seromucoid in the TCA extract of uterine washings was precipitated
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SUMMARY
Uterine washings were recovered from intact rats and rabbits at different stages of the reproductive cycle and from spayed animals, some of which were treated with ovarian hormones. The washings were analysed for several chemical constituents.
In the rat at oestrus there was a marked increase in both the amount of uterine fluid and in several chemical constituents of the uterine washing (sodium, potassium, nitrogen, carbohydrate). Similar significant increases were shown in spayed rats treated with oestradiol. The uterine washing from the rabbit contained significantly greater amounts of potassium, phosphorus, nitrogen and carbohydrate during the luteal phase (pseudopregnancy) than at oestrus. These increases were confirmed in spayed groups treated with progesterone alone or with oestradiol + progesterone and also with oestradiol replacement therapy which was probably caused by excessive dosage. These findings showed that ovarian hormones affected the composition of uterine washings and their significance is discussed.
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Yolk sac and endometrial tissue were obtained from tammar wallabies between 11 and 25 days after the removal of pouch young. Tissues were examined histologically and steroid-metabolizing enzymes were identified by incubation for 3 h at 37 °C in Medium 199 containing labelled steroid precursors. Yolk sac membrane (YSM) incubated with labelled pregnenolone produced a small amount of progesterone and pregnanediols; 80·5 ± 8·4 (s.e.m.) % of the original substrate remained unmetabolized. Labelled androstenedione was metabolized to 5α-androstane-3,17-dione and androsterone, and only 5·8 ± 3·8% of the original substrate remained at the end of incubation. Incorporation of androstenedione or dehydroepiandrosterone (DHA) into phenolic compounds was low (0·5 ± 0·1%). There was no evidence for the enzymes, arylsulphatase or sulphotransferase, in YSM. Endometrial tissue from the same animals metabolized pregnenolone, DHA and androstenedione, converted progesterone to androstenedione, and produced aqueous-soluble steroid conjugates. The results demonstrated that YSM contains enzymes associated predominantly with steroid catabolism and with incipient progesterone synthesis. The findings are discussed in relation to the histological appearance of the tissues and compared with placental steroid synthesis in eutherian mammals.
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ABSTRACT
Enzymic dispersion and density gradient separation were used for the isolation of enriched populations (60–90%) of cells from the corpus luteum, placenta and peripheral blood of pregnant sheep and goats. Analysis of the steroids produced from radioactive pregnenolone demonstrated that placental binucleate cells can produce progesterone and 5β-pregnanediol whereas white blood cells were relatively inactive. Thus, sheep binucleate cells converted pregnenolone predominantly to progesterone as did sheep luteal cells. However, goat binucleate cells produced 5β-pregnanediol as the major metabolite, which is consistent with its production in vivo during pregnancy. Production of progesterone (sheep) or 5β-pregnanediol (goat) by binucleate cells was shown to be proportional to the number and viability of the cells. In contrast with the binucleate cells there was no evidence that trophectodermal uninucleate cells play a significant role in placental progesterone or 5β-pregnanediol synthesis in either species.
Journal of Endocrinology (1991) 129, 283–289
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SUMMARY
C19-16-unsaturated steroids have been extracted from the urine and spermatic vein plasma of a mature boar and identified by thin-layer chromatography, gas—liquid chromatography and combined gas—liquid chromatography—mass spectrometry. 5α-Androst-16-en-3β-ol (approximately 250 μg./1.) was identified in the urinary glucuronide fraction. This compound and the 3α-epimer occurred in the spermatic vein plasma predominantly as sulphates but a small quantity of the 3α-isomer was extractable with ether prior to hydrolysis of steroid conjugates. Traces of 5,16-androstadien-3β-ol have been tentatively identified in the plasma sulphate fraction; 5α-androst-16-en-3-one occurred as free steroid. No 16-unsaturated steroids were found in the plasma glucuronide nor in urinary sulphate fractions. The latter contained an unidentified compound of similar polarity to the C19-16-unsaturated steroids.
Neutral 17-oxosteroids were measured in extracts obtained from both the urine and spermatic vein plasma. Of the dehydroepiandrosterone (DHA) in the urine 60% occurred as sulphate and 40% as glucuronide with only traces as free steroid. Androsterone and aetiocholanolone occurred only as glucuronides. In the spermatic vein plasma, DHA occurred predominantly as sulphate with small amounts as glucuronide and free steroid.
