Cytosolic glucocorticoid receptors in the porcine lung during development and after hypophysectomy or thyroidectomy.

The ontogeny of fetal lung glucocorticoid receptors and their regulation by the fetal pituitary, adrenal and thyroid gland during lung maturation were investigated. Sites with a specificity typical of glucocorticoid receptors were detectable in lung cytosol, with the order of potency of steroids being dexamethasone greater than cortisol greater than corticosterone greater than 11-deoxycorticosterone greater than progesterone greater than 17 alpha-hydroxyprogesterone greater than oestradiol-17 beta congruent to testosterone congruent to androstenedione congruent to oestrone. The binding affinity for [3H]dexamethasone was high (Kd = 0.23-0.60 nmol/l) and showed an age-related decrease during the perinatal period when cortisol levels were high. After charcoal treatment of the cytosol, however, a decrease in binding affinity was not as clearly evident. The Kd decreased following hypophysectomy of fetuses; thyroidectomy had no significant effect. The concentration of glucocorticoid receptors was high from day 82 to day 100 of gestation (1437 fmol/mg protein) and declined progressively to a lower value at term and following birth (660 fmol/mg protein). Hypophysectomy, but not thyroidectomy, prevented the age-related decline in receptor concentration. Lung glycogen content declined with fetal ageing in association with increases in plasma concentrations of cortisol and thyroxine and with changes in Kd and Bmax, but appeared to be more closely associated with concentrations of thyroxine. Hypophysectomy of fetuses decreased concentrations of both cortisol and thyroxine and prevented the depletion of lung glycogen content. Preliminary results from thyroidectomized fetuses showed decreases in plasma thyroxine and lung glycogen content compared with day-82 fetuses.(ABSTRACT TRUNCATED AT 250 WORDS)

Steroid and prostaglandin concentrations in the plasma of pregnant ewes during infusion of adrenocorticotrophin or dexamethasone to intact or hypophysectomized foetuses.

Catheters were implanted into 16 ewes and their foetuses between days 110 and 124 of gestation. Hypophysectomy was attempted in eight of these foetuses. Continuous infusion of synthetic ACTH (10 microgram/h) or dexamethasone (1mg/24 h) into the foetus, starting between days 124 and 129, induced premature parturition. The concentration of progesterone in the maternal peripheral plasma decreased before parturition in all animals while the level of oestradiol increased in ewes with intact foetuses or in those in which hypophysectomy was incomplete. When hypophysectomy was complete, no increase in the maternal level of oestradiol occurred before delivery. The concentration of 13,14-dihydro-15-oxo-prostaglandin F2alpha increased in the peripheral plasma of ewes with intact or hypophysectomized foetuses infused with ACTH. It is suggested that an intact foetal pituitary gland is required for the rise in the level of oestrogen prepartum, but that this rise is not essential for increased prostaglandin production of parturition.

PIP: The effectiveness of dexamethasone or adrenocorticotrophin (ACTH) in inducing parturition in the hypophysectomized fetus of the ewe was investigated. 16 cross-bred ewes were used. In 8 hypophysectomy was attempted between Days 110-120 of pregnancy. Dexamethasone or ACTH were infused into a fetal vein from Days 124 to 129 of pregnancy until delivery occurred. Progesterone, estrone, and estradiol were measured by radioimmunoassay. Cortisol was measured by competitive protein binding or radioimmunoassay. Prostaglandin (PGFM) was also determined by radioimmunoassay as was ACTH growth hormone and prolactin. The interval from the start of the infusion to delivery in fetuses adminstered dexamethasone was the same whether they were intact or hypophysectomized. The induction-delivery interval was significantly longer in the hypophysectomized fetuses administered ACTH. Hypophysectomized fetuses were less viable than intact ones. Premature delivery following infusion in the intact fetus resulted in a decrease in progesterone in maternal plasma and an increase in estradiol. In maternal plasma of intact fetuses PGFM increased in the last 20-30 hours before delivery when ACTH was infused. Differences between intact and hypophysectomized fetuses in concentrations of progesterone in the maternal plasma and in the cortisol levels were insignificant. When hypophysectomy was complete the estradiol level was without the increase seen in intact fetuses. The results suggest that an intact fetal pituitary gland is required for the rise in the level of estrogen prepartum, but that this rise is unessential for increased PGFM production and parturition.

Leuprolide acetate stimulates smooth muscle of the human reproductive tract.

Leuprolide acetate (10(-4) to 10(-5) M) had a stimulatory effect on spontaneous contractions of human myometrium and fallopian tube in vitro. The uterine and tubal tissues were from patients with dysfunctional uterine bleeding that was treated by hysterectomy. Whether the effect of LA on smooth muscle is a component of the mechanism of therapeutic action or an undesirable side effect remains to be determined.

Ultrastructural morphometric characterization of alveolar type II cells of perinatal sheep: a comparison with adults.

The quantitative morphologic changes in alveolar type II cells during the perinatal period were characterized morphometrically in the lungs of fetal lambs at 132, 138, and 147 days gestational age (DGA) and in newborns at 2 days postnatal age (2 DPN). Ultrastructural features were compared with those of type II cells of ewes 365 days old. Lamellar body profile number per type II cell profile was highest at term (147 DGA) and 2 DPN. In adults, the number of lamellar body profiles and volume density of lamellar bodies were equal to those of the 132 DGA fetus. Multivesicular bodies were most common at 138 DGA and in adults. The volume density of cytoplasmic glycogen fell dramatically during the latter part of gestation. The volume density of many cellular organelles increased to the level observed in adults by term (147 DGA). Subcellular composition of type II cells of adult sheep differs from that reported for adult rats chiefly by the volume density of lamellar material within the cytoplasm. Plate-like or globe-like inclusions were present only in the type II cells of adults. Cytoplasmic extensions of the type II cell crossing the basal lamina were most abundant in the 132 and 138 DGA fetal sheep. Cytoplasmic extensions were rare in adults. We conclude that morphologic changes of the alveolar type II cell associated with gestational age follow a species-specific time course. In the sheep, this occurs during the later part of gestation and extends into the neonatal period. Morphologic and morphometric changes appear to correspond with cellular interactions between alveolar type II cells and mesenchymal cells of the interstitium.

Fetal and maternal metabolism of prostaglandin F2 alpha in the guinea pig.

Metabolism of prostaglandin F2 alpha (PGF2 alpha) was studied with both qualitative and quantitative techniques in liver, kidney and lung of fetal and maternal guinea pigs at term. In the pregnant guinea pigs 15-hydroxyprostaglandin dehydrogenase (PGDH) activities were highest in the kidney, followed by the lung and then by the liver. In fetuses PGDH activity was highest in the liver, followed in decreasing order by kidney and lung. The kidneys and lungs of the mothers showed a 6- to 10-fold higher PGDH activity than those of the litter (P less than 0.005), but the mean PGDH activity of liver was lower in the mothers than in the fetuses. It is suggested that the liver may play a major role in prostaglandin metabolism during fetal life, and may fulfil part of the function reserved to the lung during adult life. Maternal and fetal lungs differed greatly in their handling of PGF2 alpha. This was not due to a difference in aeration, for the differences in the metabolism of PGF2 alpha between mother and litter increased when well-aerated neonatal lungs were considered instead of non-aerated fetal lungs. The observed differences in enzyme activity between fetal and neonatal lungs add yet another dimension to the role of prostaglandins in the establishment of neonatal respiration.

Comparison of intrauterine prostaglandin metabolism during pregnancy in man, sheep and guinea pig.

Metabolism of prostaglandin F2 alpha (PGF2 alpha) was studied in uterine tissues from pregnant women (n = 10), sheep (n = 6) and guinea pigs (n = 6). Two maternal tissues, myometrium and decidua or maternal placenta, and two fetal tissues, fetal placenta and membranes, were studied in each species. PGF2 alpha was metabolized via the well-known pathway into 15-keto-PGF2 alpha and 13,14-dihydro-15-keto-PGF2 alpha, while 13,14-dihydro-PGF2 alpha was tentatively identified in some tissues. The presence of 15-hydroxyprostaglandin dehydrogenase (PGDH) and 13,14-prostaglandin reductase was demonstrated in all tissues from each of the three species studied, but the quantitative intrauterine distribution of these enzymes differed from one species to another. In man, PGDH activities were highest in fetal membranes followed by fetal placenta and then by decidua and myometrium. In sheep, PGDH activities were highest in the maternal placenta followed in decreasing order by myometrium, fetal placenta and membranes. In the guinea pig, PGDH activity was highest in the fetal membranes followed in decreasing order by maternal placenta, fetal placenta and myometrium. Quantitative assay of PGDH showed that PGDH activity in the pregnant uterus is higher in women than in sheep and guinea pig. The results are discussed in relation to the involvement of prostaglandins in parturition.

Randomised clinical trial to determine optimum initiation time of norgestrel-progestin only contraception in Eldoret Teaching Hospital, Kenya.

In a randomised controlled trial to determine the optimum time of initiation of Ovrette, a progestin only oral contraceptive among postpartum women, who fully or nearly fully breast-fed their infants in the first six months, no difference was found between group 1 (initiating at six weeks postpartum) and group 2 (initiating the pill at return of menses or 6 months postpartum). There were no pregnancies in either group during the 18 month follow-up. There were no significant differences in the continuation rates between the two group.

PIP: The optimal time for initiation of progestin-only oral contraception (POC) among fully or nearly fully breast-feeding women was investigated in a comparative study conducted among 200 women delivering at Kenya's Eldoret Teaching Hospital in a 2-year period during 1992-94. 100 women were randomly assigned to initiate POC use at 6 weeks' postpartum (group 1), while the remaining 100 women deferred POC initiation until the onset of menses or 6 postpartum months (whichever event occurred first) (group 2). At 12 months postpartum, 53.57% of mothers in group 1 and 56.67% in group 2 were still taking Ovrette; at 18 months, these rates were 46.10% and 43.59%, respectively. The mean duration of use was 7.6 months in group 1 and 7.5 months in group 2. There were 178 episodes of adverse experiences with Ovrette, 16.9% of which involved menstrual disturbances. Adverse experiences were more likely to be perceived as serious and related to Ovrette in group 1 than in group 2. There were no pregnancies recorded, suggesting that initiation of Ovrette use at 6 months postpartum or when menses reappears is a feasible strategy. However, postpartum women should be provided with counseling on the efficacy of the lactational amenorrhea method.

Respiratory movements in the fetal guinea pig in utero.

Intrauterine respiratory movements in the fetal guinea pig were detected from about 0.6 of term by the use of chronically implanted tracheal and amniotic catheters. Three distinct patterns were seen: 1) "slow" breathing recurring at a rate of 1-8/min; 2) "rapid irregular" breathing in which episodes last a few seconds to 40 min and the frequency of breathing is high (greater than 0.5 HZ) (the respiratory movements are associated with negative tracheal pressures of of 0.5-20 Torr); and 3) "rapid regular" breathing which recurs in bursts lasting 0.6-6.8 min with a maximum incidence of 3/h. Inspiratory efforts in the range of 10-20 Torr were uniform during a single episode. All three types of respiratory activity were continued into first stage labor, both during and between contractions, but were absent during second stage labor.

Regulation of prolactin secretion in the pregnant pig: effect of fetal hypophysectomy and fetal death in small litters.

Fetal effects on prolactin secretion in late pregnancy and at term were studied in gilts with small litters of intact, hypophysectomized, partially hypophysectomized or dead fetuses. Litter size was reduced at surgery at Days 30-40 of gestation and fetal hypophysectomy or sham procedure attempted on Days 90-95 of gestation using an electrocoagulating probe. Plasma prolactin concentrations rose markedly before parturition in three gilts with normal litters (P = 0.05) and to lower peak concentrations in two gilts with a single fetus. Plasma prolactin did not change from basal concentrations in 4 gilts with small litters (1-2 fetuses) of hypophysectomized fetuses or, in 2 gilts with dead fetuses. Basal plasma prolactin concentrations were significantly lower in gilts carrying hypophysectomized or dead fetuses, compared to normal gilts (P = 0.01). A surge of prolactin secretion preceded delivery in 2 gilts with fetuses with incomplete hypophysectomy and in 1 gilt with 1 of 3 hypophysectomized fetuses. Abnormal patterns of prolactin secretion were seen in 1 gilt with 3 of 5 hypophysectomized fetuses and in 1 gilt with 1 partially hypophysectomized fetus. The results suggest that the fetus may be associated with a mechanism regulating plasma prolactin concentration at term. It is possible that this association may be by way of the fetal pituitary endocrine axes, and/or a function of conceptual mass.

Effect of haloperidol, suckling, oxytocin and hand milking on plasma relaxin and prolactin concentrations in cyclic and lactating pigs.

Prolactin secretion was stimulated in 5 cyclic gilts during the luteal phase (Day 10-13) with 5 mg haloperidol given i.v. Stimulation of prolactin secretion was also attempted by inducing milk let-down by suckling (4 sows), or by the injection of 1 mg oxytocin i.v. followed by hand milking (3 sows). Plasma prolactin concentrations increased significantly 1-2 h after haloperidol injection, and in 3 of 4 sows during suckling (P = 0.001); plasma relaxin concentrations did not change significantly at these times. No change was observed in plasma prolactin or relaxin concentrations at 15 min or 1-2 h after oxytocin injection and hand milking. Plasma relaxin concentrations ranged from below the sensitivity of the assay (100 pg/ml) to 450 pg/ml in lactating sows and from 100 to 2000 pg/ml in cyclic gilts. The results suggest that in cyclic gilts treated in the luteal phase with a dopaminergic receptor blocker, and in lactating sows during suckling, elevations in plasma prolactin concentrations were not accompanied, during the same period, by detectable changes in relaxin concentrations.

Induction of premature parturition in sheep: adrenocorticotrophin and corticosteroid changes during infusion of Synacthen into the foetus.

Synacthen (10 microgram/h) infused into foetal sheep at about day 125 of gestation caused premature delivery within 4 days. During the first 24 h of the infusion the secretion of adrenocorticotrophin was depressed and the assayable hormone concentration was within 500-1000 pg/ml. Subsequently, large fluctuations occurred in the adrenocorticotrophin concentration in the plasma of foetal sheep which were consistent with the stimulation of adrenocorticotrophin secretion. No transfer of Synachthen to non-infused twin foetuses was observed. The plasma corticosteroid concentration in the infused foetus also showed a small rise during the first 24 h of the infusion followed by large fluctuations in concentrations that coincided with the changes in adrenocorticotrophin. No consistent changes in the plasma corticosteroid concentration in the non-infused twin foetus were observed. These results are discussed in relation to the hormone changes occurring during normal delivery.

Relaxin inhibits myometrial activity in the ovariectomized non-pregnant ewe.

Myometrial activity was abolished abruptly but reversibly in 4 out of 5 ewes by the intravenous injection of 1 mg (500 GPU) porcine relaxin. Recovery began only after about 90 min and was not complete until 3-4 h after the injection. During the relaxin-induced inhibition the myometrium responded to oxytocin administered intravenously in doses of 250 mU. One ewe received intrauterine infusions of 2.5 and 5.0 microgram PGF-2 alpha per min during the period of relaxin inhibition: the former dose evoked a slight, and the latter a marked, response from the myometrium. The rate of rise of intrauterine pressure and the mean amplitude of pressures cycles were significantly depressed at 1, 1.5 and 2 h after the relaxin injection.

Hypophysectomy and porcine fetal lung development.

The effect of hypophysectomy on the development of the lung parenchyma and maturation of pulmonary alveolar type II cells was examined in the fetal pig. In fetuses from four different gilts, hypophysectomy was performed on gestational day 69 or 70. Littermates from the same gilts served as controls. Fetuses were delivered by caesarean section at term (113 +/- 1 days of gestation), and the lungs were fixed by intratracheal instillation. Plasma cortisol and thyroxine concentrations in the umbilical artery were markedly lower for hypophysectomized fetuses compared with control fetuses. Body weight was similar for both groups of fetuses. Total lung volume was 53% smaller in hypophysectomized fetuses compared with control fetuses. However, alveolar septal tissue and capillary luminal volumes were similar in both groups. Total lung alveolar surface area was twice as great in control animals compared with hypophysectomized animals. The volumes of epithelium, interstitium, and endothelium of centriacinar alveolar septa per unit surface area of epithelial basal lamina were 3.7, 4.8, and 2.4 times greater in hypophysectomized fetuses compared with control fetuses. Alveolar type II cell composition also differed significantly between groups. The volume fraction of glycogen in type II cells was 51% for hypophysectomized fetuses and 12% for control fetuses, while lamellar body volume fraction was 8% in hypophysectomized fetuses and 23% in control fetuses. The frequency of alveolar type II cell contact with mesenchymal interstitial cells via foot processes was 5 times greater in the lungs of control animals compared with hypophysectomized animals. These findings demonstrate significant effects of hypophysectomy on the morphogenetic and cytodifferentiation activities of all major tissue compartments of the pulmonary gas exchange area during the final trimester of fetal development.

Adrenocorticotrophin and corticosteroid changes during dexamethasone infusion to intact and synacthen infusion to hypophysectomized foetuses.

Synacthen has been infused at 10 microgram/h into 4 hypophysectomized foetal sheep. This caused a rise in adrenocorticotrophin to a mean value of 512 pg/ml which remained fairly constant and did not show the large fluctuations seen on infusion into intact foetuses. The Synacthen half-life in the circulation had a mean value of 0.27 min. Hypophysectomy did not necessarily delay the foetal corticosteroid response to Synacthen infusion. The corticosteroid concentration achieved had a mean value of about 50 ng/ml which is substantially below that for intact foetuses and in one case it was maintained at only 20 ng/ml. Despite this delivery occurred within 5 days. Dexamethasone infusion reduced the plasma adrenocorticotrophin concentration by about 70% and the plasma cortisol concentration by about 60%.

Characterization of long-term monolayer cultures of human granulosa cells from follicles of different size and exposed in vivo to clomiphene citrate and hCG.

To determine the effects of prolonged hCG treatment in vitro upon granulosa cells from follicles of various sizes previously exposed to clomiphene citrate and hCG in vivo, progesterone and relaxin concentrations of spent media were correlated with light microscopic and ultrastructural characteristics. Intact, freshly dispersed cells were characterized by numerous lipid droplets, elliptical mitochondria with tubular or lamellar cristae, moderate rough-surfaced endoplasmic reticulum (RER), sparse smooth-surfaced endoplasmic reticulum (SER), and few Golgi. After 10-24 days in culture, 2 morphologically distinct cell types, 'granulosa-type' and 'luteal-type', were noted at the light microscopic level. Ultrastructurally, lipid droplets decreased in number, mitochondria became pleomorphic, RER became more prominent and dilated, and Golgi became more widely dispersed. Tubular SER became abundant and annular nexuses became more numerous after hCG treatment in vitro. Granulosa cells generated from all follicles responded to hCG treatment with significantly increased progesterone secretion after 4 days in culture. Relaxin was not detectable in any sample of medium. This study shows that human granulosa cells from 15-25-mm follicles retain their differentiated function of progesterone secretion in long-term culture and recover responsiveness to hCG in vitro, as demonstrated by enhanced progesterone secretion and development of prominent SER and increased annular nexuses.

Maintenance of pregnancy and levels of progesterone and relaxin in the serum of gilts following a stepwise reduction in the number of corpora lutea.

The number of corpora lutea (CL) was reduced in pregnant gilts in a stepwise fashion. Eleven pregnant gilts were unilaterally ovariectomized at Day 30, leaving from 6 to 12 CL on the remaining ovary. At Day 40, the number of CL was again reduced by about half in each gilt and this was repeated on Day 50 until 1 to 3 CL remained. Blood was obtained to determine the level of progesterone in plasma on each day of surgery and 24 h later. Four gilts aborted; one had 1 CL and three had 2 CL. One gilt which had 1 CL resorbed the litter. The six gilts that maintained pregnancy from Day 50 to parturition had 1, 3, 2, 2, 2 and 2 CL, respectively. Gilts pregnant at Day 60 were also bled on that day and at 8-h intervals beginning 2 to 3 days prior to expected parturition at Day 114. At laparotomy on Day 50, gilts had from 3 to 20, nonluteinized follicles ranging from 12 to 20 mm in diameter. Hypertrophy of CL was not detected, nor were accessory CL formed. The level of progesterone in serum dropped significantly 24 h after surgery and rose to levels intermediate to pre- and postsurgery levels 10 days later. Parturition was uneventful and levels of relaxin and progesterone appeared normal.

Lung development in the nephrectomized ovine fetus.

The effect of fetal nephrectomy on lung development was studied in sheep. Fetal kidneys were removed early in the canalicular stage of lung development (95 to 99 days of pregnancy) and lung structure examined during the alveolar stage (125 to 134 days of gestation). Progesterone and estradiol 17 beta concentrations in maternal and fetal plasma were normal for gestational age, thus indicating that the ewes were not close to labour at the time the fetuses were removed. Mean body weight was significantly reduced in nephrectomized fetuses (P less than 0.05). Overall growth of the fetal lung was not markedly affected by fetal nephrectomy. However, compared to controls, alveolar airspaces were smaller in the cranial lobes of nephrectomized fetuses (P less than 0.05) and made up a smaller percentage of the parenchyma for the whole lung. There were fewer lamellar bodies per type 2 alveolar cell in nephrectomized fetuses (P less than 0.05), and more cells were without lamellar bodies (P less than 0.01). Umbilical venous thyroxin (T4) concentrations were lower in nephrectomized fetuses compared to control values (P less than 0.005). Plasma thyroxin concentration in nephrectomized fetuses correlated directly with thyroid weight (P less than 0.02). There was no significant difference in cortisol concentrations in maternal or fetal plasma or fetal tracheal fluid between the two groups. It is thus possible that the delayed lung development observed in nephrectomized fetuses was related to reduced thyroid activity.

A morphometric study of lung maturation in the adrenocorticotrophin-infused fetal lamb.

Adrenocorticotrophin (ACTH) effects on lung development were studied in sheep. ACTH was infused into 10 fetal lambs at 129 days of gestation at a rate of 29, 110 or 250 micrograms/day for 72 h, a time course which was independent of the mechanical and hormonal forces associated with labor. Lungs were compared with those of normal fetuses at term (day 147). ACTH accelerates maturation of components of lung structure toward term values. Different components of cytodifferentiation of the type II alveolar cell showed different levels of sensitivity. The degree of acceleration, not the nature of the changes, was influenced by dose.

The effects of hydrocortisone on lung structure in fetal lambs.

The effect of cortisol infusion on fetal lung development was studied in lambs. Changes were compared with those of control groups of saline-infused fetuses of the same age (day 132) and normal late gestation fetuses (142 +/- 4.6 days). Cortisol was infused into five fetal lambs at 129 days of gestation at a rate of 17.0 mg/day. Four fetuses were delivered by hysterotomy at the onset of labour-like uterine activity (58 +/- 3 h). In cortisol-infused fetuses the concentration of cortisol in fetal plasma and tracheal fluid rose to levels similar to those in normal fetuses during the last week of gestation. Progesterone concentration in maternal plasma declined at about 48 h after the start of treatment. Cortisol-infused lambs showed increases in fixed lung volume, specific lung volume, absolute volume of both parenchyma and non-parenchyma and the proportion of the parenchyma which was potential airspace and a decrease in the proportion of parenchyma. For cortisol-infused lambs Type II cell size and the abundance of lamellar bodies, and the volume fraction of cell occupied by the nucleus were similar to the 142 day group, whereas Golgi apparatus and RER were closer to age matched saline-infused (day 132) controls. Glycogen content was midway between the two control groups. We conclude that infusion of cortisol for about 60 h at physiological levels, beginning at 0.85 of gestation, accelerates many, but not all aspects of pulmonary parenchymal maturation, expressed in terms either of morphogenesis of the gas exchange area or differentiation of Type II alveolar cells.