Differences in the effects of vasopressin and oxytocin on rabbit myometrial activity and a possible mediation of prostaglandins

Uterine responses to vasopressin and oxytocin were monitored in non-pregnant and 3- or 6-8-day-pregnant rabbits by recording the intrauterine pressure. Oxytocin stimulated uterine activity in all groups, but the effect of vasopressin was stimulatory in non-pregnant animals, inhibitory in those 3 days post coitum and weakly stimulatory in those later in pregnancy. Inhibition of prostaglandin (PG) synthesis, by the administration of indomethacin, reduced the spontaneous uterine activity as well as the responses to oxytocin and vasopressin in the non-pregnant rabbits, but had little effect in the pregnant animals. During infusion of PGF-2alpha, PGE-1 or PGE-2 in 6-8-day-pregnant rabbits, the stimulatory response to vasopressin, although slight before the infusion, was inhibited whereas the stimulatory response to oxytocin remained virtually unchanged. The results suggest that vasopressin and oxytocin under certain hormonal conditions, are able to activated the uterine contractions by mechanisms in which the involvement of PG is not obligatory.     

Cholinesterase Activities in Uterus of Normal and Fenchlorphos Treated Blue Foxes (Alopex Lagopus) during Various Reproductive States

The uterine acetylcholinesterase and total cholinesterase (acetylcholinesterase plus butyrylcholinesterase) activities in normal and fenchlorphos treated blue fox vixens were determined during various reproductive states. AChE and Total-ChE of non-medicated vixens in oestrus were about one half of those in anoestrus. In pregnant uteri (luteal phase) the activities were 25 % and 30% compared to anoestrus. In vixens given 100 mg/kg fenchlorphos for 3 weeks during anoestrus, the remaining activity of AChE in uterus were in average 37%. Pregnant and non-pregnant vixens in the luteal phase medicated prior to mating and during time of implantation, displayed AChE activities which were only moderabely reduced (remaining activities 83% and 72% compared to medicated animals in anoestrus: remaining activity 37%). Plasma ChE-activity increased during pregnancy in the controls while enzyme activity was strongly reduced in animals given 100 mg/kg fenchlorphos daily through the whole pregnancy. It was concluded that the previous reported embryotoxic effect of fenchlorphos in the blue fox did not seem to be directed towards the moderate inhibition of the uterine cholinesterases.     

Uterine oxytocin receptors in cyclic and pregnant cows.

Binding of [3H]oxytocin to uterine subcellular preparations ('oxytocin receptor concentrations') was measured in uterine tissue of heifers and multiparous dairy cows at various stages of the oestrous cycle and during early pregnancy. A method for the assay of ovine uterine oxytocin receptors was optimized for use on bovine tissue. Oxytocin receptor concentrations were increased in cyclic animals around the period of luteolysis and oestrus, rising on Day 15 in endometrium and on Day 17 in myometrium while pregnant animals showed no comparable rise. Receptor concentrations then declined on Day 3 after oestrus in myometrium and on Day 5 in endometrium. Some cyclic animals did not show the expected rise in receptors in the late luteal phase; these animals had abnormally high progesterone concentrations for this stage of the cycle. In animals slaughtered on Day 18 after oestrus and/or insemination which had low oxytocin receptor levels, plasma progesterone concentrations were consistently high; while all animals showing the late luteal phase elevation in receptor values had low progesterone concentrations. Oxytocin receptor and progesterone concentrations were negatively correlated (P less than 0.05). These data support the hypothesis that oxytocin receptor level is a key factor in the process of luteolysis in cattle and that in pregnancy there is suppression of uterine oxytocin receptor at the expected time of luteolysis. We suggest that uterine oxytocin receptor levels are partly controlled by circulating steroid hormones and are suppressed during early pregnancy.     

Effects of prostaglandins on peripheral progesterone and uterine diamine oxidase in the pregnant hamster

Serum progesterone and uterine levels of diamine oxidase (DAO) activity were determined during pregnancy in hamsters. Progesterone was elevated on Day 1 of pregnancy, had a transient peak on Day 5, remained relatively constant on Days 6–10, and then increased on Days 13 and 14. Uterine DAO activity could not be detected until Day 7 of pregnancy, approximately 1 $\text{1}\text{2}$ days after the initiation of implantation. DAO activity was associated with placental tissue, and more than 90% of the activity was localized in the maternal placenta. The temporal relationship between changes in serum concentrations of progesterone and uterine levels of DAO activity following PG administration also was studied. Serum progesterone was significantly depressed by 6 hr after treatment with PGs on Day 7 of pregnancy. However, uterine levels of DAO activity at 6 hr in the treated animals were not different from those in control animals. In contrast, both the serum progesterone concentrations and uterine levels of DAO activity were significantly lower at 24 hr after PG treatment. The effects of PG treatment on uterine DAO activity were completely blocked by concomitant administration of progesterone. However, concomitant administration of Provera® only blocked the effect of one PG analog that was tested (9-deoxo-9-methylene-16,16-dimethyl0-PGE₂). The data indicate that changes in uterine DAO activity following treatment with the PGs used here are primarily a consequence of a decrease in peripheral progesterone (i.e. a luteolytic effect of the PG).     

Acid phosphatase and leucine aminopeptidase activity in the uterine flushings of non-pregnant and pregnant gilts

The activities of uteroferrin, measured as acid phosphatase (AP), and an aminoacylpeptidase (AA) were measured in uterine flushings collected from gilts on Days 6, 8, 10, 12, 14, 15, 16 and 18 of the oestrous cycle and pregnancy (N = 37). Changes in AP (P less than 0.05) were associated with day for both specific and total AP in non-pregnant and pregnant gilts. For pregnant and non-pregnant gilts, AP activity was greatest between Days 14 and 16 and then decreased to Day 18. The AA specific activity increased (P less than 0.01) between Days 10 and 12 of the oestrous cycle and pregnancy, but neither effects of pregnancy nor day by pregnancy status interaction were detected. The AA total activity was greater for pregnant gilts (P less than 0.01). These data suggest an inhibitory effect of oestrogens of blastocyst origin on synthesis and/or secretion of uteroferrin, but not AA.     

Cholinergic responsiveness of intestinal muscle in the pregnant guinea pig

Clinical observations and limited animal experiments have suggested that gastrointestinal motility is suppressed during pregnancy. We therefore compared isometric contractions of colon and ileal circular muscle in response to carbachol (10(-8) to 10(-4) M). Data was analyzed by comparing mean maximal tension, dose-response curves, and EC50 values for tissue from the two groups of animals. Circular muscle from proximal colon, distal colon, and ileum in pregnant animals developed less tension in response to carbachol than did tissue from non-pregnant controls. Dose-response curves in the pregnant groups were depressed, when compared with non-pregnant groups, at concentrations of 10(-6) M and greater. Sensitivity of the muscle to cholinergic stimulation, as measured by EC50 values, was similar in the ileum and proximal colon but increased slightly (p less than 0.05), by a factor of approximately 2, for distal colonic muscle from pregnant animals. Assuming that circular muscle contractions are primarily responsible for mixing and propulsion in the gut, this reduction in responsiveness to excitatory cholinergic stimulation is consistent with the concept of pregnancy-related suppression of gastrointestinal motility.     

Ca2+-activated proteinase in the rat. Quantification by immunoassay in the uterus during pregnancy and involution, and in other tissues.

Rat uteri were taken at various stages of pregnancy and involution post partum, and several other tissues were taken from pregnant and non-pregnant animals. Portions of each tissue were homogenized in the presence of proteinase inhibitors, and the amounts of the high-Ca2+-requiring Ca2+-activated proteinase in the supernatants were measured by a two-site immunoradiometric assay using 125I-immunoglobulin G. The proteinase was shown, by protein blotting, to be immunologically identical in all tissues. The amounts in the various tissues, expressed in units of proteinase activity/g wet wt., were: lung, 95; kidney and small intestine, 42; liver, 20; brain, heart and skeletal muscle, 13. Uterine wet weight increased at the end of pregnancy by about 8-fold, but the amounts of proteinase per uterus increased by about 22-fold; alternatively, expressed in units of proteinase activity/g wet wt., the mean uterine values were: non-pregnant, 28.6; term-pregnant, 77.0. As the wet weight of the uterus fell rapidly during involution, the amounts of proteinase activity remained relatively high. The data suggest that the Ca2+-activated proteinase may have some role in tissue resorption during uterine involution, but the high proteinase activity present before parturition must be regulated in ways which are not yet clear.     

Ultrasound-guided transfundal uterine sperm recovery from Macaca fascicularis

Previous studies from this center have indicated that the cynomolgus macaque (Macaca fascicularis) may serve as a model for human sperm interaction with the cervix and uterus. In some macaque species, transcervical aspiration of the uterine contents carries a significant risk of disturbing the cervical milieu due to the serpentine nature of the cervix. The only alternatives have been surgical procedures such as laparotomy or laparoscopy. In this paper, we report our experience with a new technique for ultrasound-guided sampling of spermatozoa in the macaque uterus. Twenty adult female cynomolgus macaques were monitored for menses (first day of menses = day l), and one mating per cycle was allowed on day 10, 11, or 12. In one group of ten animals, cervical mucus was sampled at 3 or 18 hr postcoitus (pc) and ultrasound-guided uterine aspiration was performed at 24 hours pc. In a second group of ten monkeys, uterine aspiration was at six hr pc and sperm numbers and motility were counted in the uterine fluid. Uterine fluid was obtained from fourteen of twenty monkeys. Pregnancy occurred in ten of the twenty experimental cycles. Ultrasound-guided uterine aspiration appears to be a reliable method for the evaluation of sperm transport in female macaques. The correlations between uterine sperm recovery and cervical mucus sperm populations arc discussed. The high conception rate in treatment cycles indicates that this procedure can be performed without apparent risk to pregnancy.     

Lower prevalence of chromosome aberrations and SCEs in self-poisoned pregnant women

A cytogenetic investigation was conducted in 18 self-poisoned pregnant and 16 self-poisoned non-pregnant women and in 31 pregnant and non-pregnant controls. Blood samples for analysis of chromosomal aberrations and SCEs were collected from women who were at different early stages of pregnancy. The difference between self-poisoned women and controls was very highly significant in the case of chromatid-type and unstable chromosome-type aberrations and highly significant in the case of SCEs. Further, the frequency of chromatid aberrations in pregnant women relative to non-pregnant ones was significantly lower suggesting a possible protective effect of pregnancy.     

Circadian variations of plasma renin activity (PRA), aldosterone and electrolyte concentrations in plasma in pregnant and non-pregnant goats

The aim of this study was to estimate and analyse circadian variations of the renin-angiotensin-aldosterone system (RAA) activity in blood of goats and the influence of late pregnancy on the circadian variations of RAA system. The study was carried out on a group of 17 non-pregnant and 9 pregnant goats. The animals were kept in uniform environmental conditions, (9 h light/15 h darkness). Blood samples were collected seven times over a period of 24 h, every 4 h. Plasma renin activity (PRA), plasma aldosterone (PA), sodium, potassium and chloride concentrations were determined. PRA and PA of both groups changed during 24 h, with the highest values in the dark phase and with higher RAA system activity (especially during the night) in the pregnant goats. In the non-pregnant goats, no circadian changes in PRA and PA were observed. The circadian changes in PRA and PA found in pregnant goats had acrophases at 06:27 h and 01:13 h, respectively. Plasma electrolyte concentrations in both groups of goats also changed during 24 h. These results suggest that circadian changes of potassium concentration in plasma of goats during late pregnancy may be one of the main factors affecting the RAA system.     

The Acute Phase Protein Ceruloplasmin as a Non-Invasive Marker of Pseudopregnancy, Pregnancy, and Pregnancy Loss in the Giant Panda

After ovulation, non-pregnant female giant pandas experience pseudopregnancy. During pseudopregnancy, non-pregnant females exhibit physiological and behavioral changes similar to pregnancy. Monitoring hormonal patterns that are usually different in pregnant mammals are not effective at determining pregnancy status in many animals that undergo pseudopregnancy, including the giant panda. Therefore, a physiological test to distinguish between pregnancy and pseudopregnancy in pandas has eluded scientists for decades. We examined other potential markers of pregnancy and found that activity of the acute phase protein ceruloplasmin increases in urine of giant pandas in response to pregnancy. Results indicate that in term pregnancies, levels of active urinary ceruloplasmin were elevated the first week of pregnancy and remain elevated until 20–24 days prior to parturition, while no increase was observed during the luteal phase in known pseudopregnancies. Active ceruloplasmin also increased during ultrasound-confirmed lost pregnancies; however, the pattern was different compared to term pregnancies, particularly during the late luteal phase. In four out of the five additional reproductive cycles included in the current study where females were bred but no birth occurred, active ceruloplasmin in urine increased during the luteal phase. Similar to the known lost pregnancies, the temporal pattern of change in urinary ceruloplasmin during the luteal phase deviated from the term pregnancies suggesting that these cycles may have also been lost pregnancies. Among giant pandas in captivity, it has been presumed that there is a high rate of pregnancy loss and our results are the first to provide evidence supporting this notion.     

Increase in urinary thromboxane excretion during pregnancy and labor

Urinary TXB2 excretion was measured during pregnancy and labor using high pressure liquid chromatography and radioimmunoassay. From the first trimester onwards TXB2 levels in urine of pregnant women (n = 60) were significantly (p less than 0.001) higher than in non-pregnant women (n = 12) and they increased, albeit not significantly, with advancing gestation. Labor was associated with a two-fold increase in urinary TXB2 excretion. Levels in established labor were significantly higher than at any other time in pregnancy (p less than 0.001), but the levels in incipient labor showed considerable overlap with these in late pregnancy. Thus urinary TXB2, while not necessarily originating from the pregnant uterus, appears to reflect the uterine activity of labor and may be the expression of a general stimulation of prostanoid production during parturition.     

Blood concentrations of gonadotrophins, prolactin and gonadal steroids in males and in non-pregnant and pregnant female African elephants (Loxodonta africana)

No seasonal variation in any of the hormones measured was apparent in males or females. Testosterone levels in males increased around puberty (10-11 years) and remained significantly higher in adult than prepubertal males. This was not accompanied by any significant change in levels of LH, FSH or prolactin. In non-pregnant females there was no apparent difference in levels of LH, FSH or prolactin with age. There was a significant increase in progesterone around puberty (12 years) but there was considerable overlap in values between prepubertal and adult females. During pregnancy, progesterone levels were significantly higher than in non-pregnant females with maximum levels occurring at mid-pregnancy (9-12 months). However, there was considerable overlap in values between non-pregnancy and pregnancy. Concentrations of LH and FSH decreased significantly during mid-pregnancy while prolactin levels increased dramatically during pregnancy; after 7 months of gestation until term levels were always at least 8 ng/ml greater than in any non-pregnant female. It is suggested that this consistent increase in plasma/serum levels of prolactin can be used to diagnose pregnancy in the elephant.     

Intermediate filaments in smooth muscle from pregnant and non-pregnant human uterus.

The intermediate filament proteins desmin and vimentin from pregnant and non-pregnant uterine muscle and smooth-muscle cells in culture were analysed using SDS/PAGE. The desmin content in uterine muscle increases dramatically during pregnancy, whereas vimentin remains unchanged or changes very little. When muscle cells are kept in culture, a considerable increase in vimentin content is observed as compared with vimentin in freshly isolated non-pregnant uterine tissue. Our results strengthen the view that vimentin and desmin filaments have independent function and turnover, and point to a predominantly structural role for desmin filaments.     

Collagenase activity in cervical tissue of the non-pregnant and pregnant human cervix

Collagen degradation has been suggested to play an important role in the process of cervical ripening at term. After giving informed consent tissue samples were taken from the posterior lip of 10 non-pregnant women of child-bearing age, 10 women having a termination of pregnancy at 9-12 weeks' gestation and 12 women who had had normal spontaneous deliveries at term. Collagenase activity was measured by a specific technique using native, triple helical collagen. Proteolytic activity was determined with commercially available 14C-methaemoglobin as substrate. For identification of collagen fragments SDS-polyacrylamid-gel-electrophoresis was performed using a modified Laemmli-technique. Collagenase and protease activity were found in all extracts of the different groups. There was a marked increase in enzymatic activities in the postpartum tissue samples suggesting that significant collagen breakdown had taken place. The absence of typical collagen fragments in the SDS-electrophoresis seemed to be due to a very rapid further degradation of the fragments by proteases into small peptides which then readily left the cervix. The present findings confirm the essential role of proteolytic enzymes in cervical dilatation at term.     

Relationship between release of plasminogen activator and estrogen by blastocysts and secretion of plasmin inhibitor by uterine endometrium in the pregnant pig

Pig blastocysts isolated between Days 10 and 16 of pregnancy release the protease, plasminogen activator (PA), into the medium in a time-dependent manner when cultured in vitro. Production is biphasic. The initial phase (Days 10-12) coincides with the early elongation stages, while release during the second phase (Days 14-16) occurs during a time at which the DNA content of the blastocysts is increasing markedly. Uterine flushings from these pregnant animals contain the zymogen substrate for PA, plasminogen, presumably as a serum transudate. Plasminogen is present in highest amounts at Day 12. The blastocyst, therefore, has the potential ability to generate the broadly specific protease, plasmin, within the uterine lumen. However, during this same period, the endometrium secretes an inhibitor of plasmin into the uterine lumen. In pregnant animals the amount of plasmin inhibitory activity rose 7-fold between Day 10.5, when the blastocysts were spherical, and Day 12, when they had become filamentous. At Day 12 each uterine horn contained about 3 to 4 mg of plasmin inhibitor. A similar release of inhibitor can be initiated in nonpregnant gilts given a single, intramuscular injection of estradiol valerate on Day 11 of the estrous cycle. It is suggested that the initiation of estrogen production by the elongating blastocyst triggers the release of plasmin inhibitor by the maternal endometrium and that the inhibitor serves to prevent a proteolytic cascade of reactions initiated by blastocyst PA, which might otherwise damage the uterine epithelium.     

Changes in the innervation and catecholamine concentrations in the myometrium of pregnant and non-pregnant sheep

The innervation of the myometrium in virgin, pregnant and post-partum ewes was examined using the SPG histofluorescence method and a high-performance liquid chromatographic catecholamine assay. In non-pregnant ewes, fluorescent axons were visible in all regions of the uterus. At 50 days of pregnancy the innervation was unchanged, but at 100 days axons were scarce over the whole uterus with the exception of the tubal extremities of the uterine horns. Noradrenaline concentrations were also significantly lower in late pregnancy, and significant variations occurred in different regions of the uterus, with the greatest concentrations present in the tubal extremities. At 13-16 weeks post partum, the density of innervation was variable, although the noradrenaline concentration was only slightly less than in virgin animals. Dopamine was also present in substantial quantities, but the mean concentrations remained unchanged during pregnancy.     

An investigation of the uterine luminal environment of non-pregnant and pregnant pony mares.

Uterine flushings were collected from 30 non-pregnant Pony mares on Days 8, 12, 14, 16, 18 or 20 after oculation. Mares were allowed a recovery period of one oestrous cycle and were mated at the next oestrus. They were then ovario-hysterectomized on days which corresponded to the day of the oestrous cycle to which they were assigned. Uterine flushings were analysed for total recoverable protein and acid phosphatase activity. Least squares analysis indicated a status X day interaction for total protein (P less than 0.10) and acid phosphatase activity (P less than 0.005) in which the latter was higher in uterine flushings during pregnancy. Peripheral plasma oestrone and oestradiol concentrations were measured by radioimmunoassay and results indicated that plasma oestrone concentrations in pregnant and non-pregnant mares were not different, and oestradiol was lower (P less then 0.005) in the peripheral plasma during pregnancy. conceptus membranes were incubated in vitro for 120 min in a chemically defined medium. Incubation medium was then assayed to assess oestrone and oestradiol production capacilities at Days 8, 12, 14, 16, 18 and 20 of pregnancy. Conceptus membrane production of oestradios (pg/5 ml/h) increased (P less than 0.05) from Day 8 (243 pg/5 ml) to Day 20 (108 763 pg/5 ml). A similar trend, but of lower magnitude, existed for oestrone production.     

The relaxant property of local prostaglandin E-2 on the non-pregnant uterus - a cyclic triphasic response.

The response of the non-pregnant human uterus to intravenous (i.v.) injections and intra-uterine instillation of various doses of prostaglandin E-2 (PGE-2) was evaluated at the different phases of the menstrual cycle in 13 fertile regularly menstruating women who were neither lactating nor using any hormonal therapy. Uterine contractility was recorded by the microballoon technique in at least three sessions(proliferative, mid-cycle and secretory) in a single cycle with endometrial biopsy performed immediately following the last session to ascertain that the particular cycle was an ovulatory one. Single i.v. injections of PGE-I had a consistent stimulatory effect on the contractility throughout the cycle with a tendency towards a decreased uterine response at mid-cycle and luteal phase as compared to the proliferative part of the cycle. Intra-uterine instillation of the compound induced a peculiar and interesting type of response. In the proliferative and secretory phases of the cycle the response was one of stimulation; being more pronounced in the former period. However, around ovulation time, the local administration induced an evident uterine relaxation in most cases without any instance of stimulation. The possible implication of this triphasic response behaviour on the non-pregnant uterus within certain physiological events and pathological conditions is discussed.