Environmental stress alters the developmental pattern of delta 5-3 beta-hydroxysteroid dehydrogenase activity in Leydig cells of fetal rats: a quantitative cytochemical study

Quantitative cytochemistry was used to determine the effect of subjecting pregnant rats to environmental stress on the activity of delta 5-3 beta hydroxysteroid dehydrogenase (3 beta-HSD) in Leydig cells of their fetuses. Enzyme activity was measured by microspectrophotometry in individual Leydig cells in cryostat sections of fetal testes on Days 16-21 postconception. Fetuses of stressed mothers lacked the peak of enzyme activity on Days 18 and 19 of gestation that is characteristic of Leydig cells of normal fetuses at this time. In addition, both before and after these 2 days, 3 beta-HSD activity in Leydig cells of stressed fetuses was significantly higher than normal. The altered developmental pattern of 3 beta-HSD activity in the stressed fetuses largely corresponds to the changes in plasma testosterone found previously in male fetuses of mothers exposed to the same regimen of stress. Thus, in the fetal Leydig cell, the activity of 3 beta-HSD, a key steroidogenic enzyme, can be modified by environmental stress, and provides an index of steroidogenic activity of the fetal testes and of the titers of circulating testosterone.     

Endocrine parameters associated with disruption of parturition after bilateral pelvic neurectomy.

Bilateral lesions of the pelvic nerve (BLPN) result in dystocia, but the processes which control this effect are not fully understood. Plasma progesterone, relaxin, and luteinizing hormone (LH) concentrations were measured in blood samples taken in the morning (AM) and evening (PM) of Days 20-23 of gestation from rats with BLPN or sham neurectomy. Ten of 11 sham-operated control animals delivered their entire litters by Day 23 of gestation, but animals with BLPN did not complete parturition by Day 23 when they were sacrificed. Progesterone concentrations were greater in rats with BLPN than in sham-operated rats on Day 20 PM and Day 21 AM, but hormone concentrations declined to minimal values by Day 22 in both groups. Relaxin concentrations were greater in rats with BLPN than in sham-operated rats on Day 21 PM. Thereafter, relaxin concentrations decreased to reach minimum values on Day 23 in both groups. LH concentrations were low throughout the period of study in rats with BLPN; however, a postpartum LH surge was detected in all sham-operated animals. Data from this study indicate that the pelvic nerve does not control parturition by modulating serum relaxin and progesterone concentrations; however, these data suggest that impulses carried by the pelvic nerve influence ovarian secretion of these hormones. In addition, these data indicate that the pelvic nerve transmits stimuli from the cervix to the hypothalamus to facilitate the postpartum LH surge.     

Progesterone and gravidity differentially regulate expression of extracellular matrix components in the pregnant rat myometrium

Myometrial growth and remodeling during pregnancy depends on increased synthesis of interstitial matrix proteins. We hypothesize that the presence of mechanical tension in a specific hormonal environment regulates the expression of extracellular matrix (ECM) components in the uterus. Myometrial tissue was collected from pregnant rats on Gestational Days 0, 12, 15, 17, 19, 21, 22, 23 (labor), and 1 day postpartum and ECM expression was analyzed by Northern blotting. Expression of fibronectin, laminin beta2, and collagen IV mRNA was low during early gestation but increased dramatically on Day 23 during labor. Expression of fibrillar collagens (type I and III) peaked Day 19 and decreased near term. In contrast, elastin mRNA remained elevated from midgestation onward. Injection of progesterone (P4) on Days 20-23 (to maintain elevated plasma P4 levels) delayed the onset of labor, caused dramatic reductions in the levels of fibronectin and laminin mRNA, and prevented the fall of collagen III mRNA levels on Day 23. Treatment of pregnant rats with the progesterone receptor antagonist RU486 on Day 19 induced preterm labor on Day 20 and a premature increase in mRNA levels of collagen IV, fibronectin, and laminin. Analysis of the uterine tissue from unilaterally pregnant rats revealed that most of the changes in ECM gene expression occurred specifically in the gravid horn. Our results show a decrease in expression of fibrillar collagens and a coordinated temporal increase in expression of components of the basement membrane near term associated with decreased P4 and increased mechanical tension. These ECM changes contribute to myometrial growth and remodeling during late pregnancy and the preparation for the synchronized contractions of labor.     

Ontogeny of secretory patterns of LH release and effects of gonadectomy in the chronically catheterized pig fetus and neonate

Two experiments were conducted to determine the ontogeny of secretory patterns of luteinizing hormone (LH) release and effects of gonadectomy on the characteristics of LH secretion in the chronically catheterized pig fetus and neonate. To study secretory patterns in intact animals, blood samples were collected from 44 pig fetuses and their mothers (Days 81, 99, 109 and 113 of gestation) as well as from 25 neonates (Days 4 and 8) every 15 min for 3 h (2 h on Day 81). The results indicate that the fetal adenohypophysis secretes occasional pulses of LH as early as Day 81 of fetal life. Fetal and maternal mean LH levels are low (0.25-0.50 ng/ml) at all gestational ages, with lowest values just before birth (Day 113 post coitum). Four-day-old neonates show a significant increase in pulse frequency (male and female) as well as pulse amplitude (female), relative to fetal values, leading to significant augmentations in mean LH levels. This is associated with reductions in both 17 beta-estradiol and progesterone. By 8 days of age significant sex differences in mean LH levels (males greater than females) appear. Testosterone/5 alpha-dihydrotestosterone levels (males) are low prenatally but are significantly increased after birth, possibly due to the stimulating effects of increasing LH levels. To study the gonadal control of LH secretion, forty-one 105-day-old fetuses and thirty-eight 4-day-old neonates were chronically catheterized and were either gonadectomized or remained as sham or control animals. Forty-eight and 96 h after surgery, blood samples were taken every 15 min for 3 h. No significant changes are detectable at 96 h in mean LH, pulse frequency and amplitude in female or male fetuses or in neonates. While significant reductions in testosterone levels are observed at 96 h in the male fetus and neonate, progesterone concentration is reduced only in the neonate. In the castrated female, on the other hand, neither fetus nor neonate display significant changes in circulating levels of progesterone and 17 beta-estradiol at 96 h. It is concluded that the pituitary of the pig is able to discharge LH with occasional pulses as early as Day 81 of fetal life; however, the pituitary remains suppressed until after birth, probably due to high circulating nongonadal steroids in the fetal compartment.     

Role of luteinizing hormone-releasing factor (LH-RF) and LH on maintenance of early gestation in rats.

The role of luteinizing hormone (LH) and LH-releasing hormone (LH-RH) in the maintenance of early pregnancy in rats was studied. Serum levels of progesterone (P) and LH were measured daily in untreated pregnant rats from Day 4 through parturition. Serum levels of P and LH were determined on Days 11 and 15 of pregnancy in animals treated with antisera to LH (LH-A/S) and to LH-RH (LH-RH-A/S) on Days 8-10. Serum levels of P peaked on Days 7 and 16 in untreated animals, after which they declined sharply just before delivery. Serum LH fluctuated between 30-160 ng/ml during pregnancy but did not exhibit any distinctive peaks. Treatment with .2 ml LH-A/S on Days 8-10 reduced serum P to virtually undetectable levels on Day 11, and only a slight recovery was evident on Day 15. Lower doses of LH-A/S had no effect. Administration of 1.3 ml LH-RH-A/S had no effect on serum levels of P or LH, and did not impede fetal development. The results indicate that LH is essential to the luteotropic complex of early pregnancy in the rat, and also that LH-RH-A/S can maintain to some extent basal levels of P and LH during early pregnancy.     

Soluble T lymphocyte antigen-specific molecules.

Growth hormone (GH) was measured in the sera of control, hypothyroid (thyroidectomized [Tx]) and GH-treated Tx rats and their fetuses on Days 19, 20, 21, and 22 of gestation and in their progenies on postnatal Days 1, 5, 30, and 75. Maternal endogenous serum GH increased dramatically between the 19th and 20th days of gestation and remained elevated through the 22nd day in control rats, but was depressed significantly in Tx and GH-treated Tx rats during this period. GH was not always detected in the sera of 19-day-old fetuses. On Day 20, GH was depressed in fetuses of Tx mothers as compared with those form controls or GH-treated Tx mothers. GH was elevated in sera of fetuses from GH-treated Tx rats over fetuses of control and Tx only rats on the 22nd day of gestation. In postnatal rats, those from GH-treated mothers continued to show elevated serum GH on Day 1 as compared with those from Tx only mothers. On postnatal Days 5 and 30, progenies of Tx mothers had significantly elevated GH as compared with progenies of control mothers. At 75 days of age, the GH levels of these progenies had normalized. We have shown previously that the hormonal secretions of the pituitary-thyroid axis are badly disrupted in the progenies of Tx and GH-treated Tx mothers and that even as adults these animals have tissue (brain and liver) deficits of active thyroid hormones. Although the onset of GH secretion is mildly delayed in fetuses of Tx but not GH-treated Tx mothers, the serum GH levels of both groups of progenies are elevated during most of the neonatal period through the time of puberty. It is, therefore, concluded that GH in the absence of adequate levels of thyroid hormones is ineffective in preventing many of the learning and memory deficits induced in the progenies of Tx mothers.     

Metabolism of testosterone in vivo in the pregnant rat

The metabolism of testosterone (T) was examined during the second half of pregnancy in the rat to determine whether utilization of T for estradiol (E2) synthesis occurs via conversion of T to androstenedione (A). On Days 11, 16, and 21 of gestation (term = Day 23), rats (n = 7-9/group) were anesthetized and a constant infusion of [3H]T was initiated. At 60 min, blood was obtained from a jugular vein and the ovaries (Days 11, 16, and 21), and placentae and uterine tissue (Day 16 only) were removed. In a second study performed in rats on Day 16 of gestation (n = 8-10/group), the ovaries and/or gravid uterus were removed 15 min after initiation of [3H]T infusion, and blood was taken from a jugular vein 60 min later. Radiolabeled T and A were purified from serum and tissues by paper chromatography. In a third group of rats (n = 6), jugular vein samples were obtained sequentially on Days 11, 16 and 21 of gestation and serum concentrations of T were measured by radioimmunoassay. The metabolic clearance rate of T was constant during the study period (overall mean = 31 1/day). In contrast, the serum concentration of T (pg/ml) on Day 16 of gestation (863 +/- 108) exceeded (p less than 0.02) that on Day 11 (445 +/- 74); the latter was similar to that measured on Day 21 (592 +/- 109). Thus, the estimated production rate of T was greatest on Day 16 of gestation.(ABSTRACT TRUNCATED AT 250 WORDS)     

Determination of the source of immunoreactive relaxin in the cat

We have recently reported the secretory profile of relaxin throughout gestation in the cat. Because the appearance of relaxin begins at about Day 20 (Day O = ovulation) and because implantation begins shortly before this at Days 13-14, we hypothesized that relaxin was of feto-placental origin. To test this hypothesis, we used 4 experimental groups: 1) Control (laparotomy-only at Day 23 or 42, n = 4); 2) Early Ovariectomy (Ovx, bilateral ovariectomy between Days 23 and 26, n = 4); 3) Late Ovx (bilateral ovariectomy between Days 40 and 44, n = 4); 4) Tissue Removal (removal of feto-placental units, uterus, and one ovary on Days 16, 21, 28 and 35, n = 1 per day). Pregnancies were maintained in both Ovx groups by progesterone administration. Relaxin secretory patterns in Ovx groups were similar to the Control data. Relaxin was detectable in plasma beginning at about Day 20, with maximum concentrations reached by Day 30. Relaxin concentrations were highest (immunoactivity per mg tissue) in homogenates of placental tissues as compared to luteal, fetal, or uterine tissues. Altogether, these data indicate that the feto-placental unit is the source of relaxin in the cat.     

Changes in serum immunoreactive inhibin and follicle-stimulating hormone during gonadal development in male and female rats.

We have measured serum and ovarian immunoreactive inhibin alpha (irI alpha) and serum FSH in fetal and neonatal rats from 20 days of gestation until 40 days of age. For animals aged 10 days or older, serum measurements were made on intact and gonadectomized animals. Serum irI alpha was detectable in intact male and female rats at all ages studied. In females, irI alpha levels were low until Day 5 and then increased steadily to peak at Day 25. Thereafter they declined until Day 35 to reach levels typical of adult females. There was a significant decrease in irI alpha levels 24 h after ovariectomy at all ages. Serum FSH levels in females were low until Day 7, then increased rapidly to plateau from Days 10-15. The levels then declined until Day 25 and were generally unchanged after that time. There was a significant increase in FSH 24 h after ovariectomy in rats aged 20 days and older, and in younger rats by 48 h after ovariectomy. In male rats, serum irI alpha levels were significantly higher than females until Day 7. The levels increased at Day 7 and then remained relatively constant until Day 20, after which they declined to reach typical adult male levels. Serum irI alpha levels were significantly lower in males than females from Days 25-40. There was a significant decrease in serum irI alpha 24 h after castration at all ages studied. Serum FSH levels in males were low until Day 20, increased at Day 25, and thereafter remained relatively unchanged.(ABSTRACT TRUNCATED AT 250 WORDS)     

Effects of environmental stress or ACTH treatment during pregnancy on maternal and fetal plasma androstenedione in the rat

Prenatal stress applied during the last trimester of pregnancy has been shown to alter fetal development and influence adult sexual behavior. Since androstenedione (Δ⁴) has the potential to participate in differentiation processes, this study was designed to assess the effect of prenatal stress on maternal and fetal Δ⁴ titers. Restraint/illumination/heat (environmental stress) or ACTH injections were used to stress pregnant rat dams beginning on Day 14 of pregnancy. Blood samples and organ weights were obtained from nonpregnant animals, pregnant rats on Days 5, 10, 15, 18, and 20 of pregnancy, and fetuses on Days 18 and 20 of gestation. Maternal and male and female fetal Δ⁴ titers were determined by radioimmunoassay. ACTH and environmental stress significantly reduced fetal body weight and male anogenital distance. Environmental stress also significantly reduced the size of 20-day fetal adrenals and testes. Each treatment caused significant short-term (1 hr after treatment) and long-term (16 hr after treatment) elevation of maternal plasma Δ⁴ on Days 15 and 18 of gestation, but only short-term elevation of Δ⁴ titers on Day 20. ACTH treatment did not cause long-term elevation of fetal Δ⁴ although both ACTH treatment and environmental stress generated a significant short-term increase in fetal Δ⁴ titers. Environmental stress produced long-term elevation of fetal Δ⁴ in 18-day fetuses of both sexes and in 20-day female fetuses. It is concluded that maternal stress and exogenous ACTH significantly elevate maternal and fetal Δ⁴ titers during the prenatal period postulated to be critical in sexual differentiation of the rat brain.     

Progesterone prevents the pregnancy-related decline in protein kinase A association with rat myometrial plasma membrane and A-kinase anchoring protein

The presence of cAMP-dependent protein kinase (PKA) in the plasma membrane compartment and its association with an A-kinase anchoring protein (AKAP150) is implicated in mediating cAMP regulatory events in the rat myometrium. The association of PKA with purified myometrial plasma membrane declined gradually between Day 16 and Day 21 of gestation, with a decrease of 53% +/- 11% of the catalytic subunit and of 61% +/- 7% of the regulatory subunit at Day 21 compared with Day 19. To determine the role of progesterone in this association, pregnancy was prolonged by administration of progesterone or shortened by administration of the antiprogestin RU486. Progesterone treatment maintained PKA association with plasma membrane at Day 21 at 123% +/- 23% (catalytic subunit) and 92% +/- 4% (regulatory subunit) of Day 19 levels. In contrast, protein phosphatase 1, protein phosphatase 2B, phospholipase Cbeta(3), and AKAP150 concentrations in the plasma membrane did not change over this interval or with progesterone treatment. Changes in PKA coimmunoprecipitated with membrane-associated AKAP150 paralleled those in total plasma membrane on Days 19 and 21 and on Day 21 following progesterone treatment. In contrast, plasma membrane PKA catalytic and regulatory subunits decreased by 20 h after RU486 injection on Day 15 of pregnancy to levels resembling those on Day 21. These data indicate that progesterone prevents the decline in PKA associated with myometrial plasma membrane and with AKAP150 in the pregnant rat. The decrease in membrane-bound PKA between Days 19 and 21 and after RU486 treatment precedes the onset of parturition in both experimental paradigms. The loss of plasma membrane PKA may be critical for the decrease in the inhibitory effect of cAMP on oxytocin-induced phosphatidylinositide turnover that occurs near the end of pregnancy and may contribute to enhanced myometrial contractile responsiveness near term.     

Evidence for hormonal imbalance after methadone treatment in pregnant and pseudopregnant rats

The effects of methadone (METH) on the plasma estriol level and hormonal target tissues' cyclic nucleotides (cAMP and cGMP) were investigated in pregnant and pseudopregnant rats. In the pregnant animals, METH (5 mg/kg/day), given once daily from Days 6 to 15 of gestation, significantly reduced the maternal body weight gain in association with an increase in the number of dams bearing resorptions (56%) and a significant reduction in fetal body weight (33%). An inhibition of the plasma estriol level by METH was observed on Day 9 of gestation. Stimulation of the sympatho-adrenal axis and hypothalamo-pituitary axis by acute METH administration was observed and correlated with a significant increase in the levels of cyclic nucleotides in the uterus and adrenal glands of pregnant rats. However, tolerance to METH effects on cyclic nucleotide levels developed by Day 15 of gestation. METH also depressed the fetal cyclic nucleotide levels on Days 12 and 15 of gestation. These findings suggest that METH had pronounced effects on hormonal secretion during pregnancy, and hormonal transport to or hormonal production by the fetuses. In contrast, METH did not exhibit any adverse effects on the hormonal and cyclic nucleotide levels of pseudopregnant rats with deciduoma formation; a model for the maternal compartment. These latter findings may reflect METH's adverse effects on the fetal compartment, and suggest the use of pseudopregnancy as a model to distinguish adverse drug effects between these compartments.     

Measurement of messenger RNA for luteinizing hormone beta-subunit and alpha-subunit during gestation and the postpartum period in ewes

Pituitary content of luteinizing hormone (LH) and mRNAs for LH beta-subunit (LH beta), alpha-subunit, prolactin, and growth hormone were measured in ewes on Days 50 and 140 of gestation and on Days 2, 13, 22, and 35 postpartum. Content of LH in dissociated anterior pituitary cells declined (P less than 0.05) between Days 50 and 140 of gestation and remained low at 2 days postpartum. By 22 days postpartum, pituitary concentrations of LH were comparable to concentrations in normally cycling ewes. During gestation concentrations of mRNA for LH beta and alpha-subunit paralleled changes in cellular content of LH, reaching minimal levels on Day 140. By Day 2 postpartum, pituitary concentrations of mRNAs for LH beta and alpha-subunit began to increase; they reached maximum levels by Day 13 postpartum. There appeared to be a gradual linear increase in mRNA for prolactin through gestation and the postpartum period. No changes in mRNA for growth hormone were noted during the prepartum or postpartum periods. These data suggest that the decline in pituitary concentrations of LH during gestation is due to a decrease in cellular mRNA for LH beta and alpha-subunit. The increase in mRNA for LH beta and alpha-subunit appears to precede an increase in cellular content of LH in the postpartum ewe by several days.     

Ontogeny of the response of the hypothalamo-pituitary-adrenal axis to maternal immobilization stress in rats.

In this study we investigated the response of the rat fetal hypothalamo-pituitary-adrenal (HPA) axis to an acute maternal stress in late gestation. On day 20 of gestation, pregnant rats were exposed to forced immobilization stress for up to 60 min. In mothers, a significant increase in plasma ACTH and corticosterone(B) was observed at 20 and 60 min. The ACTH content in the maternal pituitary decreased significantly at 60 min. Fetal blood pH was decreased by the maternal stress, showing a hypoxic condition of the fetus. Fetal plasma ACTH increased transiently at 20 min. Fetal plasma B increased at 20 and 60 min. ACTH in the fetal pituitary and the placenta did not show marked changes due to the maternal stress. Pregnant rats on day 18-21 of gestation were subjected to a 20 min maternal stress. In the basal condition without stress, fetal plasma ACTH and B showed parallel ontogenic patterns, having a peak value on day 19 of gestation. Fetal plasma ACTH as well as plasma B were increased significantly by the maternal stress at all points evaluated. These results indicate that fetal hypoxia is important in stress transmission to the fetal HPA axis in this type of maternal stress, and the fetal HPA axis responds to the stress as early as day 18 of gestation.     

Influence of progesterone concentrations on secretory functions of trophoblast and pituitary during the first trimester of pregnancy in dairy cattle

The essential role played by progesterone in the maintenance of pregnancy is unequivocal; however, the effects of progesterone on the secretory patterns of placental and pituitary molecules during the gestation period are not well defined. The objective of this study was to describe pregnancy-associated glycoprotein (PAG) concentrations (measured by RIA-497 and RIA-Pool) in pregnant females with progesterone concentrations lower (low-P4 group, n=20) or higher (high-P4 group, n=17) than the mean of 8.74 ng/mL on Day 21 (AI=Day 0). Luteinizing hormone (LH) and prolactin concentrations were also measured in both groups. Throughout the study period, blood samples were collected on Days 0, 21, 45, 60, and 80 from 37 females that were confirmed to be pregnant. PAG concentrations measured by both RIA-497 and RIA-Pool tended to be higher in high-P4 group than in low-P4 group from Day 30 until Day 80. On Day 80, plasma PAG concentrations that were measured using RIA-497 were observed to be higher (P<0.05) in the high-P4 group than in the low-P4 group (10.2+/-8.7 ng/mL versus 6.9+/-3.8 ng/mL). Concentrations of LH on Day 60 and prolactin on Day 80 were observed to be significantly lower (P<0.05) in the high-P4 group. There was a tendency for the concentrations of LH (Days 45 and 80) and prolactin (Days 30, 45, and 60) to be lower in cows in the high-P4 group than in the low-P4 group. Our results suggest the existence of a relationship among the concentration levels of progesterone, PAG, LH, and prolactin during early pregnancy.     

Insulin-like growth factors and their binding proteins define specific phases of myometrial differentiation during pregnancy in the rat

While the insulin-like growth factor (IGF) system is known to regulate uterine function during the estrous cycle, there are limited data on its role in myometrial growth and development during pregnancy. To address this issue, we defined the expression of the Igf hormones (1 and 2), their binding proteins (Igfbp 1-6), and Igf1r receptor genes in pregnant, laboring, and postpartum rat myometrium by real-time PCR. IGF family genes were differentially expressed throughout gestation. Igf1 and Igfbp1 mRNA levels were upregulated during proliferative phase (Days 6-12) of rat gestation. Igfbp3 gene expression also was elevated in proliferating smooth muscle cells (SMCs) and was highest at the time of transition between proliferative and synthetic phases (Days 12-15). Igfbp6 gene expression profile paralleled plasma progesterone (P4) concentrations, peaking during the synthetic phase (Days 17-19) and decreasing thereafter. Administration of P4 at late pregnancy (starting from Day 20) to maintain elevated plasma P4 concentrations blocked the onset of labor and prevented the fall in Igfbp6 mRNA levels. In contrast, the treatment of pregnant rats with the P4 receptor antagonist RU486 on Day 19 induced preterm labor and the premature decrease of Igfbp6 gene expression. Igfbp2 gene expression was transiently upregulated during the contractile phase of gestation (Days 21-23) solely in the gravid horn of unilaterally pregnant rats, but it was not affected in P4- or RU486-treated animals, supporting a role for mechanical stretch imposed by the growing fetuses. Igfbp5 gene was induced during postpartum involution. Our results suggest the importance of the IGF system in phenotypic and functional changes of myometrial SMCs throughout gestation in preparation for labor.     

Androgen and estradiol levels in plasma and amniotic fluid of late gestational male and female hamsters: uterine position effects

Using radioimmunoassay we have measured the plasma and amniotic fluid levels of androgen and estradiol in male and female hamster fetuses nearing parturition. On Days 14 and 15 of gestation (day of birth = Day 16), plasma levels of androgen are higher in males than females while estradiol levels are equal. Amniotic fluid levels of these hormones, while lower than plasma, reflect the difference in androgen and the similarity in estradiol between sexes. Uterine position analysis on Day 14 suggests that female siblings located caudally suppress amniotic fluid androgen and elevate estradiol levels of male siblings. Comparison of Day 18 gestation male and female rat amniotic fluid androgen to Day 14 hamsters reveals that male rats are bathed in high levels of androgen. Female rats have lower levels which are not different from those of male hamsters. Female hamsters are exposed to little androgen. Relevance to behavioral sexual differentiation and the display of adult behavior is discussed.     

Synchronization of oestus and timed insemination of mares.

Oestrus was synchronized in 116 mares by means of an i.m. injection of prostaglandin F-2 alpha (Day 0) and of fluprostenol (a PG analogue) on Day 16. Mares were then randomly divided into three groups. Group A mares (N = 30) were given 2500 i.u. hCG I.M. ON Day 20 and artificially inseminated on Day 21 without detection of oestrus. Group B mares (N = 32) were given 2500 i.u. hCG i.m. on Day 20 and inseminated on Days 21 and 23, also without oestrus detection. Group C mares (N = 54) were teased on Days 18, 19, 21, 23 and 25 and inseminated on Days 19, 21, 23 and 25 while they were in oestrus. Semen was collected by artificial vagina from 3 stallions. One-third of the mares in each group were assigned to each stallion at random. The gel-free fraction was divided equally among the mares, and used within 1 h of collection. Pregnancy rates at about 60 days of gestation were not significantly different. A high rate of synchronization of oestrus (80%) was attained within 48 h of treatment with fluprostenol.     

Myometrial adenylyl cyclase protein decreases on the last day of pregnancy in the rat

To determine whether gestation-related changes in responsiveness of the rat uterus to beta-adrenergic agonists are mediated at the level of adenylyl cyclase, we measured myometrial adenylyl cyclase activity and protein quantities during pregnancy and labor. In rat myometrial membranes, basal adenylyl cyclase activity increased from the nonpregnant state to mid (Days 12-14) and then late (Days 18-20) gestation and then decreased intrapartum (Day 22). Stimulated adenylyl cyclase activity, at the level of the beta-adrenergic receptor (isoproterenol, 10(-4) M), the G protein (GTP, 10(-5) M), or the adenylyl cyclase enzyme (MnCl(2), 20 mM), was similarly altered during gestation. Total adenylyl cyclase protein was quantified by [(3)H]forskolin binding assay in myometrial membranes from nonpregnant and pregnant (Day 14, Day 20, Day 21, and intrapartum Day 22) rats. Adenylyl cyclase protein increased progressively from nonpregnant rats to pregnant rats at mid (Day 14) and late (Day 20) gestation, but it decreased abruptly to nonpregnant levels on Day 21, the day before parturition, and remained at similar levels on Day 22 (intrapartum). The gestation-related increase in expression of myometrial adenylyl cyclase protein may facilitate uterine quiescence during pregnancy, and the abrupt decrease of adenylyl cyclase protein on the last day of pregnancy may be a contributing mechanism for the initiation of labor.