Effects of 17 beta-estradiol on myometrial gap junctions and pregnancy in the rat

The effects of estradiol treatment on the development of myometrial gap junctions and premature labour were investigated using timed pregnant rats. In control animals myometrial gap junctions were infrequent between days 17 and 20 of pregnancy, but began to develop on day 21 and were at maximum frequency, size, and membrane area on day 22 during delivery. Gap junctions were completely absent from the myometrium 48 h after delivery. Animals treated with 500 micrograms 17 beta-estradiol/day starting on day 16 of pregnancy developed numerous myometrial gap junctions and delivered their pups prematurely on day 19. Similarly, treatment with 50 micrograms estradiol/day resulted in the development of myometrial gap junctions on day 20 of pregnancy and premature labour. However, treatment with various doses of estradiol up to and including 500 micrograms/day for 3 days beginning 1 day before delivery was not able to maintain the presence of myometrial gap junctions during the postpartum period. These results support the hypothesis that estradiol stimulates the development of myometrial gap junctions and that the presence of gap junctions in the myometrium is a requirement for the occurrence of term, as well as preterm labour. Furthermore, it is evident from this study that the postpartum regression of myometrial gap junctions is not dependent on the decrease in estradiol.     

Effects on maternal and fetal steroid concentrations of induction of parturition in the sheep by inhibition of 3 beta-hydroxysteroid dehydrogenase

Changes in circulating steroid hormones, the incidence of myometrial contractions, and the onset of labour were all monitored after administration of the 3 beta-hydroxysteroid dehydrogenase inhibitor, epostane, to chronically catheterized ewes and fetuses near term. In all animals the drug induced delivery 33-36 h after injection or infusion into the ewe with the birth of live healthy lambs which showed normal subsequent development. Epostane induced immediate, permanent falls in both maternal and fetal plasma progesterone concentrations, accompanied by increased PGF metabolite concentrations in the uterine vein beginning 15 min after treatment. Of the other hormonal changes observed, the most striking was the pronounced drop in both maternal and fetal plasma cortisol. In the fetus this fall was followed by increasing concentrations of circulating ACTH which eventually restored the cortisol levels. By 12-24 h after epostane a substantial overshoot had occurred and at 27-30 h the fetal plasma cortisol concentrations were as high as those seen during normal parturition at term. No significant changes in maternal plasma oestradiol-17 beta could be detected after epostane treatment or during labour. The incidence of slow myometrial contractions increased significantly during the second 3-h period after epostane, although their duration did not change. Contraction patterns typical of first stage labour were seen from 20 to 24 h. These results show that epostane may be used as a safe, predictable inducing agent in sheep if given 6-10 days before term; the lambs showed no signs of prematurity despite their lowered plasma cortisol concentrations which persisted for some hours before labour was induced.     

Labour: when to worry.

To ensure an optimum result in pregnancy it is essential that the physician be alert in the antenatal period to recognize those women and their babies who are at risk during labour. Premature labour, with its attendant risk of respiratory distress syndrome in the newborn, continues to be an important factor in perinatal morbidity and mortality. Early recognition of predisposing factors and the judicious use of myometrial inhibiting agents have helped to reduce the incidence of fetal prematurity in these cases. A long interval between rupture of the membranes and delivery continues to be a danger to both mother and fetus. Delivery is recommended when gestation is beyond 36 weeks or when there are signs of incipient infection, and once labour has begun antibiotics should be used prophylactically. Failure of labour to progress should be recognized and managed aggressively in its early stages. Amniotomy and oxytocin infusion have reduced considerably the incidence of prolonged labour and its risks to both mother and fetus. The role of intrapartum monitoring of the fetal heart rate, measurement of the pH in the fetus''s scalp blood and assessment of amniotic fluid is discussed, as is the monitoring of maternal well-being.     

Local and systemic control of myometrial contractile activity during labour in the sheep

The relative contribution of systemic versus local (intrauterine) factors in the activation and stimulation of the sheep myometrium during labour was examined using an in-vivo myometrial explant preparation. Myometrial tissue alone (MYO) or with attached endometrium (ENDO/MYO) was removed from the pregnant uterine horn, sutured to a stainless-steel frame and placed into the omental fat. After 7-10 days the explants developed a pattern of electromyographic activity qualitatively similar to that of the uterine myometrium. Induction of preterm labour by infusion of ACTH (66.6 ng/min for 15 min every 2 h) to the fetus resulted in a reduction in plasma progesterone concentrations and increases in values of oestradiol-17 beta and 13,14-dihydro 15-keto PGF-2 alpha in maternal plasma. The onset of labour, which followed these endocrine changes, was characterized by an increase in EMG burst frequency and reduction in burst duration occurring simultaneously in both the uterine myometrium and in the explants. The response of the uterine and explant myometrium to oxytocin also exhibited a parallel significant increase over the 24-h period leading to delivery. No differences were apparent between the explants containing myometrial tissue alone or those comprising endometrial and myometrial tissue. There was no significant change in uterine or explant EMG activity, or oxytocin responsiveness, after saline administration to the fetus. The pattern of EMG activity changes during spontaneous labour were not distinguishable from those during ACTH-induced labour. As with oxytocin, the responsiveness of the explants to electrical stimulation increased significantly at labour compared to pre-labour. These data suggest that factors within the systemic circulation play a major role in both the onset of labour contractions and the increased response to electrical or hormonal (oxytocin) stimulation during parturition in sheep.     

Active Management of Labour

Active management of labour has been developed to the extent that an assurance is given to every woman who attends this hospital that her first baby will be born within 12 hours. This assurance could lead to a welcome change in present attitudes because the mere prospect of prolonged labour is often a cause of serious concern during a first pregnancy. Labour of strictly limited duration makes it possible to provide every woman with a personal nurse, and it places the problem of pain in a new setting.The results of a prospective study of 1,000 consecutive primigravidae are presented, and the guidelines to a standard policy of management are defined. Difficulty in meeting the commitment to early delivery arose almost exclusively in cases in which the diagnosis of labour was in doubt; only seven women were retained in the delivery unit for 12 hours.     

Progesterone and oestradiol levels and cytoplasmic receptor concentrations in the human myometrium at term, before labour and during labour

Progesterone (P) and oestradiol (E2) cytoplasmic receptor levels in the myometrium of 12 women, who underwent cesarean section at term were determined by means of an exchange assay. Six of the women had an elective cesarean section, and the other six were in active labour when the operation was performed. Both the P and the E2 receptor concentrations were significantly higher in the myometrium of those women who were in labour. The plasma P and E2 levels did not change before labour and during labour. The myometrial E2 concentrations were also similar in both groups. The myometrial P concentration was lower in the labour group, but the difference was not statistically significant. This study is the first which compares the steroid receptor levels at term before labour and during labour in human myometrium, although only the unbound and the cytosol receptor levels were determined. The change of levels in receptor concentrations could be a sign of the decrease of the P dominance in the myometrium during labour.     

Immunohistochemical analysis of connexins 26, 32 and 43 in rat uterus during late pregnancy: lack of connexin 26 in the myometrium

The spatial and temporal patterns of expression of connexin 26, connexin 32 and connexin 43 were investigated in rat uterus at days 17, 19 and 22 of pregnancy and during delivery (23 days) by immunocytochemistry, Gap junctions, which are essential for the development of labour, are known to undergo rapid increase in the rat myometrium at the end of pregnancy. The expression of connexin 43, the major myometrial gap junction protein, was low throughout pregnancy but increased immediately before the onset of labour (day 22). It was found predominantly in the myometrium, although limited staining was also apparent in the stroma. Immunolabelling revealed the presence of connexins 26 and 32 in uterine luminal epithelial cells on days 17 and 19 of pregnancy, with a marked increase in connexin 26 expression at days 19, 22 and 23; however, marked expression of connexin 32 was apparent only at day 23. No immunoreactivity for either connexin 26 or 32 was found in myometrial cells at any stage of pregnancy. We conclude, contrary to other recent reports, that connexin 26 is not a gap junction protein of the rat myometrial smooth muscle cell. © 1998 Chapman & Hall     

The role of prostaglandins for the coordination of myometrial forces during labour

Systematic studies using a superfusion technique for recording myometrial contractility in vitro have been conducted in our department to explore whether prostaglandins (PG) have a differential action on the different segments of the pregnant uterus and also whether the qualitative and quantitative response undergoes a change during spontaneous labour. Myometrial specimens were excised from the fundal area and from the lower uterine segment at elective caesarean section in the 39th week of pregnancy before commencement of labour and at acute caesarean section during ongoing labour. Before labour PGF2 alpha was without or had a very weak effect on upper segment preparations but was stimulatory on lower segment specimens. PGE2 and PGI2 generally induced a biphasic dose-dependent response (stimulation followed by inhibition). During spontaneous labour PGF2 alpha and PGE2 always stimulated upper segment preparations while the contractile activity of specimens from the lower segment was inhibited by PGE2, PGF2 alpha was generally without effect. PGI2 had the same biphasic action before as during labour. With all reservations for the validity of in vitro experiments, the results favour the hypothesis that initiation of labour in the human involves a qualitative shift in the myometrial reactivity to prostaglandins. These alterations may involve suppression of expulsive forces and perhaps some tightening of the lower uterine segment during pregnancy. Following initiation of labour there is a marked increase in the excitatory action of both PGE2 and PGF2 alpha in the fundal area while the lower uterine segment reacts in a way that favours dilatation.     

The effect of labour on uterine blood flow in the pregnant ewe.

The effect of labour on cardiac output and uterine blood flow was measured in pregnant ewes at a mean gestation of 124 days using radioactive microspheres labelled with 169Yb and 85Sr. Labour was induced by a continuous infusion of ACTH into the foetal circulation. Cardiac ouput measured before ACTH infusion in seven ewes was 5234 +/- 175-9 ml./min (mean +/- S.E.) and total uterine blood flow was 732 +/- 57-9 ml./min (mean +/- S.E.). Measurements during labour in six ewes showed a significant increase in cardiac output to 6175 +/- 149-6 ml./min (P less than 0-005) but no significant change in uterine blood flow. However, the partition of blood flow was altered; thus myometrial flow increased by 67% from 114 +/- 15-4 ml./min to 190 +/- 13-2 ml./min (P less than 0-005) while placental blood flow decreased, although not significantly, from 618 +/- 55-9 ml./min to 575 +/- 40-7 ml./min. Similar changes were observed in one ewe in spontaneous labour at term and in another ewe receiving an infusion of 4 mg oestradiol 17beta over a 24 hr period. It is concluded that labour is not associated with any major alternation in total uterine blood flow although myometrial blood flow is increased. It is not known whether this is due to the rise in circulating oestrogens which occurs prior to parturition in the ewe, or to other factors such as the work of uterine muscle during labour.     

Different patterns of myometrial activity and 24-h rhythms in myometrial contractility in the gravid baboon during the second half of pregnancy.

Two clearly distinct epochs of myometrial contractility were observed in 13 pregnant baboons when recorded either as intraamniotic pressure (IAP) or myometrial electromyogram (EMG). Contractures, epochs lasting longer than 3 min, were the characteristic form of myometrial activity throughout pregnancy. Contractures generated only small increases in IAP. Short-lived contractions, generating larger increases in IAP, were characteristic of labor and delivery. Power spectral analysis of IAP and myometrial EMG activity proved to be an effective means whereby periods when the myometrium was in the contractures or contractions mode could be easily distinguished. Concomitantly recorded maternal intraabdominal temperature showed significant 24-h variations. When myometrial activity switched from low-amplitude, long-lasting regular contractures of pregnancy to contractions, the switch always occurred around the onset of darkness. Five baboons went into spontaneous labor, 3 prematurely and 2 at term. In these animals the switch from contractures to contractions occurred for several nights before delivery. The recurrence and timing of the switch from contractures to contractions for several nights before delivery were similar to the pattern we and others have observed in the pregnant rhesus monkey. The presence of 24-h periodicity in the patterns of specific types of myometrial activity in another nonhuman primate lends support to the view that similar 24-h patterns of myometrial activity may occur in pregnant women.     

Synthesis and in vitro characterization of oxytocin receptor targeted PEGylated immunoliposomes for drug delivery to the uterus

Abstract

Targeted delivery of therapeutics to the uterus is an important goal in the treatment of obstetric complications, such as preterm labour, postpartum hemorrhage, and dysfunctional labour. Current treatment for these obstetric complications is challenging, as there are limited effective and safe therapeutic options available. We have developed a targeted drug delivery system for the uterus by conjugating anti-oxytocin receptor (OTR) antibodies to the surface of PEGylated liposomes (OTR-PEG-ILs). The functionality of the OTR-PEG-ILs has previously been evaluated on human and murine myometrial tissues as well as in vivo in a murine model of preterm labour. The aim of this study was to report the pharmaceutical synthesis and characterization of the OTR-PEG-ILs and investigate their specific cellular interaction with OTR-expressing myometrial cells in vitro. Immunoliposomes composed of 1,2-distearoyl-sn-glycero-2-phosphocholine (DSPC) and cholesterol were prepared using an optimized method for the coupling of low concentrations of antibody to liposomes. The liposomes were characterized for particle size, antibody conjugation, drug encapsulation, liposome stability, specificity of binding, cellular internalization, mechanistic pathway of cellular uptake, and cellular toxicity. Cellular association studies demonstrated specific binding of OTR-PEG-ILs to OTRs and significant cellular uptake following binding. Evaluation of the mechanistic pathway of cellular uptake indicated that they undergo internalization through both clathrin- and caveolin-mediated mechanisms. Furthermore, cellular toxicity studies have shown no significant effect of OTR-PEG-ILs or the endocytotic inhibitors on cell viability. This study further supports oxytocin receptors as a novel pharmaceutical target for drug delivery to the uterus.     

An opioid antagonist stimulates myometrial activity in early postpartum cows

The influence of the opioid antagonist naltrexone on myometrial activity of cows was tested during the early postpartum period, when uterine contractions are a highly sensitive indicator of oxytocin actions. Registering intrauterine pressure oscillations in five first calf heifers revealed that myometrial activity was enhanced after naltrexone was administered (0.02 mg/kg BW). However, plasma concentrations of oxytocin were not consistently elevated. In addition, naltrexone did not reliably potentiate the suckling-induced peak of oxytocin release in plasma. In all cases, naltrexone injections caused increases in the cows' respiratory rates and distinct behavioral changes during the first days postpartum, but not later. We conclude that: 1) In the early postpartum period, an inhibitory opioidergic control of myometrial activity may exist in cattle. 2) This control does not seem to be caused by a change in circulating oxytocin levels. 3) Opioids may exert an inhibitory influence on respiratory functions in cattle.     

Active management of labour: current knowledge and research issues.

OBJECTIVES--To review the evidence that the package of labour interventions collectively called "active management"--namely, strict diagnostic criteria for labour, early amniotomy, early use of oxytocin, and continuous professional support--reduce rates of caesarean sections and operative vaginal delivery in first labours. DESIGN--Review of observational data, supplemented by evidence from four separate overviews of relevant randomised trials previously published as part of the Cochrane Collaboration pregnancy and childbirth database. RESULTS--Observational data do not permit a clear conclusion. There have been no randomised trials of the total package of active management or of the use of strict diagnostic criteria alone, but trials of early amniotomy, early oxytocin, and these interventions combined do not suggest that these interventions are effective in reducing rates of caesarean sections or operative vaginal deliveries. In contrast, the provision of continuous professional support in labour seems to reduce both types of operative delivery, although the effect on caesarean sections is confined to those settings where non-professional companions are not normally present in labour. CONCLUSIONS--Delivery units should endeavour to provide continuous professional support in labour, but routine use of amniotomy and early oxytocin is not recommended.     

Temporal structuring of delivery in the absence of a photoperiod: preparturient myometrial activity of the rhesus monkey is related to maternal body temperature and depends on the maternal circadian system.

No convincing evidence exists that the shift from myometrial contractures to contractions, which determines the synchronized 24-h rhythm in the dynamics of the primate uterus, may be attributed to an endogenous circadian rhythm. We therefore wished to ascertain whether a 24-h periodic shift would also occur in the myometrial activity of animals kept under constant conditions. We studied five pregnant rhesus monkeys, kept in continuous darkness from 56-77 days gestational age until delivery at 117-167 days gestational age. During the last week before delivery we determined the individual phase, level, and amplitude of circadian changes in maternal body temperature and 24-h myometrial activity patterns in the form of contractions. In all five monkeys, a rhythm with a period of 24-h characterized the temporal incidence of preparturient contraction activity. A consistent phase lag of 6-7 h from the temperature crest was observed in four out of the five animals. The circadian phase of all individual rhythms was idiosyncratic among animals. We conclude that endogenous rhythms in body temperature and preparturient myometrial activity are truly circadian. In addition, these rhythms are either interdependent or subject to the same maternal timekeeping mechanism, supporting the hypothesis that the exact time of the day at which birth occurs in the rhesus monkey depends on the maternal circadian system.     

Identification of bactenecin-1 in cervicovaginal fluid by two-dimensional electrophoresis in an ovine model of preterm labour

Preterm labour is a major problem in obstetrics. Timely intervention with available treatments is hampered by the lack of a reliable test of imminent preterm delivery. Current methods of diagnosis are based on the detection of breakdown products of foetal membranes or structural changes to the cervix when preterm labour is well established. The aim of this study was to screen the cervicovaginal fluid (CVF) proteome to identify labour-associated proteins that could be used as markers of imminent preterm delivery. Labour was induced in sheep at 135 days of gestation (term 147 days) by foetal infusion of dexamethasone (1 mg/24 h). CVF samples were collected before and 28 h after the start of infusion as well as at delivery (58.7 +/- 1.9 after the start of infusion, n = 5). One protein that was upregulated eight-fold, was bactenecin-1, a member of the cathelicidin family of antimicrobial proteins. This antimicrobial protein warrants further investigation as a marker of preterm labour, particularly during the period after the initiation of labour but before there is marked cervical connective tissue breakdown.     

Loss of myometrial oxytocin receptors during oxytocin-induced and oxytocin-augmented labour

Oxytocin is used widely for the induction and augmentation of labour, but there is little information about the dynamics of oxytocin receptors in human myometrium during parturition, and the possible effect of oxytocin infusion. This information is important because G protein-coupled receptors, such as the oxytocin receptor, undergo desensitization after prolonged or repeated stimulation. The concentration of myometrial oxytocin receptors and the steady state of its mRNA were measured in patients undergoing Caesarean sections before or during spontaneous or induced labour. The concentration of receptors before labour was 477 (175-641) fmol mg(-1) protein (median, quartile range), and decreased to 140 (72-206; P < 0.05) and 118 (69-75; P < 0.01) fmol mg(-1) protein during prolonged oxytocin-augmented and oxytocin-induced labour, respectively. The corresponding oxytocin receptor mRNA concentrations decreased by 60- and 300-fold, respectively. The decrease in receptor binding and mRNA in women receiving oxytocin infusion indicates that homologous receptor desensitization occurs in vivo.     

Expression of scaffolding, signalling and contractile-filament proteins in human myometria: effects of pregnancy and labour

Successful parturition requires the co-ordination of numerous myometrial signalling events to allow for timely and efficient uterine contractions. Late pregnancy and labour onset in humans may be associated with changes in the expression of myometrial proteins implicated in such uterine contractile signal integration. Accordingly, in myometria from non-pregnant women and pregnant women, not in labour or in labour, we examined the content of putative plasmalemmal scaffolding proteins (caveolin-1 and -2) and compared these to the proportions of signal transducing rho-associated kinases (ROKalpha and beta) and contractile filament-associated proteins alpha-actin, myosin regulatory light chain (MLC(20)) and h-caldesmon. There was no effect of pregnancy or labour on the proportion of caveolin, ROK betaor alpha-actin. However, pregnancy was associated with a decrease in ROKalpha and MLC(20) such that ROK alpha: alpha-actin and MLC(20): alpha-actin ratios were reduced compared to myometria of non-pregnant women. In contrast, h-caldesmon was up-regulated in pregnancy resulting in an elevated h-caldesmon: alpha-actin ratio. There were, however, no further significant changes in ROK alpha, MLC(20) or h-caldesmon expression with spontaneous or oxytocin-induced labour. These data suggest that the mechanism(s) integrating myometrial signalling events with the onset of human labour does not involve differential alterations of the cellular expressions of caveolins, ROK, alpha-actin, MLC(20) or h-caldesmon.     

Prophylactic administration of the beta-blocker carazolol to influence the duration of parturition in sows

A change in the ratio of myometrial alpha- and beta-receptors during pregnancy in the pig causes beta-receptors to dominate at parturition. Hence, epinephrine, when discharged because of stress, causes myometrial relaxation. Consequently, parturition is interrupted and delivery delayed. In a double-blind field trial with 1066 sows, prophylactic treatment with 0.5 mg/50 kg b.w. of the beta-blocking agent carazolol (Suacron^®)⁺ at the onset of labour resulted in a shortened duration of parturition (p < 0.05), a reduction in the stillbirth rate and in obsterrical complications as well. The incidence of puerperal disorders was reduced in first-litter gilts from 20.5% (placebo group) to 9.9% (p < 0.01).