PGF2 alpha and PGE2 binding to rat myometrium during gestation, parturition, and postpartum.

PGFaaand PGE2 binding to rat myometrium gestation, parturition,

during

and postpartum

MIKLOS MOLNAR AND FRANK HERTELENDY Departments of Obstetrics and Gynecology and Physiology, St. Louis University School of Mdicine and St. Mary’s Health Center, St. Louis, Missouri 63104

MOLNAR, MIKL~S, AND FRANK HERTELENDY.PG&, and PGE2 binding to rat myometrium during gestation, parturition, and postpartum. Am. J. Physiol. 258 (Endocrinol. Metab. 21): E740-E747, 1990.-The specific binding of prostaglandins (PG) FzCuand ES was studied in a rat myometrial membraneenriched fraction during the latter part of gestation and parturition, as well as in the postpartal period. Tritiated PGEz and PGF2,, binding was specific, saturable, time dependent, and directly proportional to the amount of membrane protein. Scatchard analysis indicated the presence of high-affinity (IQ) and low-affinity (IQ binding sites for both PGs. The affinity of both binding sites for PGFZ, and the apparent IQ for PGE, remained essentially the same throughout gestation and postpartally and were similar to nonpregnant rats. The apparent Kd, of PGE2, however, increased by IO-fold from day 21 of gestation to 1 day postpartum. Although the maximal binding capacity of the high-affinity (B,,,,) and low-affinity (B,,,,) binding sites of PGF2, showed a nonsignificant increase compared with prepartum values, reaching maximal values 12-24 h postpartum, those of PGE2 showed a significant increase on the third day after delivery. The concentration of prostanoids in uterine venous plasma and amniotic fluid increased significantly with approaching parturition, whereas plasma progesterone decreased, raising the estradiol-progesterone ratio 25fold. After unilateral fetectomy, the binding sites for PGF*, and PGE2 increased significantly compared with the contralatera1 pregnant horns. Administration of the PG synthetase inhibitor, indomethacin, also increased two- to threefold both PGF2,, and PGE2 binding compared with the placebo group, whereas intrauterine administration of PGF2, and PGE2 significantly reduced it. The results demonstrate the existence of high- and low-affinity binding sites for both PGFz, and PGE2 and indicate that the number of PGF*, and PGE2 receptors in the rat myometrium increase around the time of parturition, but the high rate of endogenous production of these prostaglandins may impede the detection of such by conventional techniques. fetectomy;

indomethacin;

uterus; receptor

when progesterone production by the corpus luteum and/or placenta progressively increases, the uterus is relatively quiescent, and its ability to respond to contractile agonists is suppressed (9). With approaching parturition, the sensitivity of the myometrium to such agonists increases, a phenomenon that has been linked in several subprimate mammalian species to falling progesterone levels, rising estrogen levels, and the ensuing increase in the estradiol to progesterone ratio. Even though the mechanism(s) by which female sex DURING GESTATION,

E740

0193-1849/90

$1.50 Copyright

steroids influence uterine contractility is incompletely understood, estrogen has been shown to increase and progesterone to decrease the receptor concentration of myometrial agonists such as oxytocin (21,28) and angiotensin II (33), and of ,&adrenergic receptor agonists (25). Such an increase in myometrial contractility is consistent with an increase in receptor number under the regulatory influence of changing levels of female sex steroids in species where such changes have been documented. However, the extension of this concept to the human female and other primates, which fail to exhibit the falling progesterone-rising estrogen pattern (17)) remains problematic. Prostaglandins (PG) FZcuand E2, on the other hand, can activate the progesterone-dominated, oxytocin-refractory human myometrium and have been successfully used clinically to terminate second trimester pregnancy in the human female when oxytocin is ineffective (6). From a physiological point of view, PGFZ, and PGEZ have been generally viewed as key components of a complex mechanism regulating parturition. The cellular mechanism of action of PGs is assumed to be initiated by specific binding to cell membrane components (20), the characteristics of which comply in general with the requirements for hormone receptors. Such specific binding has been documented in mammalian (26) and avian uterine preparations (2, 31, 32), yet published information about the relationship between myometrial PG receptors and parturition is scanty. The aim of this study was to characterize PGF2, and PGEZ binding to a rat myometrial membrane-enriched preparation during the latter part of gestation, parturition, and the postpartum period. In addition, we also evaluated PG binding after administration in vivo of PGFZ,, PGEZ, and indomethatin, as well as in myometrial preparations from unilaterally fetectomized rat uterine horns. MATERIALS

AND METHODS

Chemicals. Radioactive prostaglandins [5,6,8,11,12,14,15-3H(N)]PGE2 (specific activity 144-200 Ci/mmol) and [5,6,8,9,11,12,14,15-3H(N)]PGF2, (specific activity 150 Ci/mmol), over 99% pure by thin-layer chromatography, were purchased from DuPont NEN (Boston, MA). Unlabeled PGF2, (Enzaprost F) was supplied by Chinoin (Budapest, Hungary) and PGE2, PGDZ, PGE1, and 6keto-PGF,, were provided by Upjohn (Kalamazoo, MI). Indomethacin was a gift from Merck Sharp & Dohme

0 1990 the American Physiological Society

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PGF2,,

AND

PGE2

BINDING

(Rahway, NJ). Whatman glass microfiber filter (GF/B) was obtained from Fisher Scientific (St. Louis, MO). Heparin was purchased from Elkin-Sinn (Cherry Hill, NH) and Nembutal (pentobarbital sodium) from Veterinary Laboratories (Lenexa, KS). Other chemicals used were obtained from Sigma Chemical (St. Louis, MO). Animals. Pregnant and nonpregnant Sprague-Dawley rats, -200 g, were obtained from Sasco (O’Fallon, MO). Labor occurred between the evening of day 21 and the afternoon of day 22 of gestation. Nonpregnant animals were used in diestrous as determined by vaginal smear. The rats were anesthetized with Nembutal, and 3 ml blood were removed from the uterine vein into a syringe, containing 300 IU heparin and 0.03 mg indomethacin, to determine the plasma concentration of prostaglandins, estrogen, and progesterone. Preparation of crude membrane fraction. The uterine crude membrane fraction was prepared according to the procedure described previously (2, 18), with some modifications. Briefly, the rat uteri in various stages of gestation were excised and rinsed with ice-cold saline containing 10 pg/ml indomethacin. The myometrium was dissected from decidua-endometrium and was minced with scissors before homogenization in ice-cold buffer containing 0.25 M sucrose, 1 mM CaC12, 1 mM 2-mercaptoethanol, 10 pg/ml indomethacin, and 10 mM tris( hydroxymethyl)aminomethane (Tris) HCl, pH 7.5 (buffer A). The mince was homogenized on ice by three 15-s bursts using a Polytron PT-10 homogenizer at setting 5. The homogenate was centrifuged for 15 min at 2,000 g. The resulting supernatant was centrifuged for 20 min at 10,000 g, after which the supernatant was incubated for 30 min at 37°C in a water bath, followed by centrifugation at 90,000 g for 1 h. The pellet was suspended in buffer A, using a micro glass homogenizer. Prostaglandin binding assay. Binding of PG was determined by incubation in buffer A of the myometrial preparation (100-200 pg protein/tube), with increasing concentration of [3H]PGF2, (0.7-1.60 nmol/l) or [3H]PGE2 (0.16-50 nmol/l) in the absence or presence of loo-fold concentration of the corresponding nonlabeled prostaglandin. The incubation was carried out at 37°C for 60 min in a Dubnoff shaking water bath in a final volume of 200 ~1. In experiments designed to measure the changes of the maximal binding capacity of PGF2, and PGE2 receptors during pregnancy, aliquots of the suspension were incubated with [3H]PGF2, at concentrations of 7.7 and 160 nmol/l, or with [3H]PGE2 at concentrations of 2.5 and 50 nmol/l in the absence or presence of a loo-fold excess of the corresponding unlabeled prostaglandin to evaluate high- and low-affinity PG binding sites, similar to the method described by other investigators for measuring the maximal binding sites of oxytocin receptors in rat myometrium (1). At the end of incubation, the mixture was diluted with 5-ml ice-cold buffer A and was immediately filtered through a Whatman GF/B membrane filter under vacuum. The filter was washed four times with 5 ml ice-cold buffer A and placed into scintillation vials, and 6 ml of scintillation cocktail [0.55% wt/vol PPO (2,5-diphenyloxazolone), 0.0167% wt/vol POPOP (p-bis[2-(5-phenyloxazolyl)] l

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benzene) in a 2:l mixture of toluene-Triton X-100] was added to each vial. The vials were capped, shaken, and allowed to stay overnight at room temperature. The radioactivity of samples was determined in a Beckman LS-7000 liquid scintillation counter. The assays were performed in duplicate. Specific binding was calculated as the difference between total and nonspecific binding values and is expressed in terms of DNA content of tissue homogenate. (In two experiments designed to characterize specific binding of pooled membrane preparations, DNA determinations were not performed, and binding data are related to protein content.) The experiments were repeated at least four times with membranes isolated from different animals. Saturation curves of specific binding values were analyzed using an Apple IIe computer and the ALLFIT program (10). Fetectomy. One uterine horn from each of 15 pregnant rats was fetectomized via laparatomy on the 14th day of gestation. Both fetus and placental tissues were removed. The animals were divided into five groups of three rats per group. The first group was killed on the 20th day of gestation, the second group at delivery, the third group 12 h after delivery, the fourth group 1 day after delivery, and the fifth group 5 days after delivery. The uterine horns were excised and processed individually as described above. Treatment with indomethacin. Sixteen rats on the 15th day of gestation were divided into two groups of eight rats per group. One of the groups received via a gastric tube 2 ml of 0.5 M sucrose solution (control). The other eight rats were treated with indomethacin (2 mg/kg body wt) and suspended in the same volume of sucrose solution twice a day for four days. On the morning of the 5th day the animals were anesthetized, their uteri removed, and PG binding sites were estimated using the same procedures as described above. Assays. Protein concentrations of the starting homogenate and the crude membrane preparations were determined by the Coomassie brilliant blue G-250 binding method (4), using bovine serum albumin fraction V as the standard. DNA content of the myometrial homogenate was determined by the method of Burton (5). Concentrations of PGFZ,, PGE2, 13,14-dihydro-15-ketoPGF2, (PGF,), 6-keto-PGF1, (the stable metabolite of prostacyclin), thromboxane Bz (TxB2), progesterone, and estrogen were measured by radioimmunoassay procedures (14). StatisticaL analysis. Data were subjected to one-way analysis of variance and post hoc Newman-Keul’s test. When otherwise indicated, Student’s t test or paired t test were used. Differences of P < 0.05 level were considered significant. Data points represent means t SE of duplicate determinations from a number of experiments as indicated in the legends to Figs. 7-10. Correlation between [3H]PG uptake and membrane protein was computed by linear regression analysis. RESULTS

The specificity of PGF2, and PGEZ binding is illustrated in Fig. 1. A concentration of 1 X 10D8 M unlabeled PGF2, displaced 50% of [3H]PGF2,. About a tenfold


E742

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AND

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BINDING

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Prostaglandin Log,, (mol/O FIG. 1. Specificity of PGFzcl and PGE, binding to rat uterine membrane preparations. Pooled rat myometrial tissue from postpartum rats was processed within 12 h from parturition as described in MATERIALS AND METHODS. [“HIPGF,,, or [“H]PGEz (0.2 &i) was incubated with membrane fraction in presence of increasing amounts of various prostaglandins. Binding measured in presence of 1 pM PGFztI or 1 FM PGEz was regarded as nonspecific binding and was subtracted from each value. Incubation was done in 0.2 ml volume at 37°C for 60 min. Each assay tube contained 0.2 mg membrane protein.

higher concentration of PGEl or PGEZ was required to accomplish the same, whereas the efficiency of PGD2 and 6-keto-PGF1, to compete for PGFZ, binding sites was only l-2% of that of the homologous ligand. Similarly, 1 X 10D8 M PGEz or PGE, displaced 50% of [3H]PGE2, whereas the ability of PGF2, and PGD2 to compete was -5% and that of 6-keto-PGF,,, was 0.05). This was followed by a progressive increase, reaching significant levels (P c 0.01) 3 days postpartum (92.0 t 5.6 fmol/mg DNA). By 5 days postpartum, levels returned to about nonpregnant values (35.2 t 4.3 fmol/ DNA). The number of low-affinity binding sites in-


E744

PGF*,

AND

PGEz

BINDING

creased, though not significantly, at the time of delivery from 157.7 t 35.3 to 407.1 t 55.1 fmol/mg DNA and to 827.6 Z!I 21.4 f mo l/ mg DNA 3 days postpartum (P < O.Ol), returning to nonpregnant levels by 5 days postpartum. Plasma concentrations of PGF2, in the uterine venous blood increased progressively more than tenfold from day 16 of gestation to parturition, from 0.18 t 0.04 (n = 6) to 2.6 t 0.46 rig/ml (n = 11). An even more marked increase was observed in the levels of the PGFZ, metabolite, PGF,, from 0.06 t 0.02 to 2.05 t 0.45 rig/ml. TxB2 concentrations stayed constant during this period, rising only twofold at parturition to 0.64 t 0.19 rig/ml (n = 9). Similarly, a significant 3.5-fold increase (from 0.09 t 0.01 to 0.33 t 0.1) in 6-keto-PGF1, levels was observed during parturition. By 3 days postpartum (when PG binding in vitro was at or near its peak), uterine plasma prostanoid levels were virtually nondetectable. Prostanoid levels in the amniotic fluid collected between days 16 and 21 of gestation showed a similar pattern. PGF2, levels increased from 0.32 t 0.04 on day 16 to 4.73 t 0.74 rig/ml (n = 6) by day 21 of gestation. Concentrations of PGE2 rose from a low of 0.81 t 0.06 on day 16 to a maximum of 6.07 t 1.40 rig/ml on day 21 of gestation. Interestingly, both TxB2 and 6-keto-PGF1, also increased almost lo-fold during the same period, from 0.48 t 0.13 to 4.11 t 1.16 and from 0.08 t 0.01 to 0.76 t 0.14 rig/ml, respectively. Concomitantly with these increments in uterine prostanoid production with approaching parturition, plasma levels of steroid hormones in uterine venous blood exhibited their expected changes. Progesterone levels dropped precipitously from 268.9 t 33.0 at day 16 to 23.7 t 1.3 rig/ml by day 21, while estradiol increased from 116.7 t 10.4 to 285.3 t 59.0 pg/ml, raising estradiol to progesterone ratios (pg/ ng) about 25-fold. In view of the observed increase in uterine production of PCs and the concomitant decrease in the binding capacity of myometrial preparations in vitro, we attempted to alter endogenous levels of PGs and then measure PGFZ,, and PGEZ binding. In the first of such experiments, 1 pg PGFZ, or PGEZ was injected in one uterine horn, the contralateral horn serving as control. Thirty minutes later the animals were hysterectomized, and the specific uptake of [3H]PGF2, and [3H]PGE2 by the freshly prepared membrane-enriched fraction of myometrial homogenate was determined. Administration of exogenous PGF2, or PGEZ significantly suppressed subsequent myometrial binding in vitro, suggestive of reduced availability of unoccupied binding sites (Fig. 7). Next, we removed the fetoplacental unit from one uterine horn of each of another group of rats, with the pregnant horn serving as control. In myometrial membranes prepared from fetectomized horns, the specific binding of [H]PGFz, to both high- and low-affinity binding sites increased two- to threefold during labor compared with the contralateral pregnant horn (Fig. 8). This was followed by a rapid postpartal fall of the highcapacity binding site in the fetectomized uterus coincident with a rise in the binding capacity of the spontaneously delivered control uterus and the falling endoge-

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LOWSITES

HIGHAFFINITY

BINDING

LOWSITES

FIG. 7. Effect of intrauterine administration of PGFz, and PGE2 (1 ,ug/rat) on subsequent in vitro uptake of [3H]PGF2, and [3H]PGE2. Rats (n = 4/group) were laparotomized under Nembutal anesthesia within 24 h postpartum, and PGs were injected (1 ml) into one uterine horn, the contralateral horn receiving 1% ethanol in saline (1 ml). Thirty minutes later, animals received an overdose of Nembutal, and both uterine horns were excised and processed as described in MATERIALS AND METHODS. Open bars, specific uptake of [3H]PGFz, and [3H]PGE2 in crude myometrial membrane fraction of vehicle-treated horns; hatched bars, same in PG-treated uterine horns. * P < 0.05; ** P c 0.025; *** P 0.01 by paired t test.

nous PGF2, levels. With respect to PGE2 binding, significant differences between fetectomized and control uteri were observed shortly after parturition was completed, particularly in the case of the low-affinity receptors (Fig. 9) . Finally, to evaluate the effect of inhibition of endogenous PG production on PG binding in vitro, pregnant rats were treated with indomethacin. In myometrial membranes prepared from indomethacin-treated rats, binding of both PGF2, and PGEz to either low- or highaffinity binding sites increased by two- to threefold (Fig. 10) . DISCUSSION

Although the intimate mechanism by which PGs trigger uterine contractions is not well understood, the interaction with specific “receptors” in the plasmalemma of myometrial smooth muscle cells is generally considered to represent the initial step (20). Previous studies using differential and discontinuous sucrose gradient centrifugation of rat (18) and hen (29) myometrial homogenates have shown a parallel increase in PGF2, binding and the degree of enrichment of the various fractions with plasma membrane as judged by a corresponding increase in the activity of marker enzymes. Similar findings were reported for nonpregnant human myometrial tissue (8). These and other studies support the notion that uterine cell membranes contain specific PG receptors. It should be noted, however, that binding of PGs to intracellular organelles, including the sarcoplasmic reticulum, of bovine myometrium (7) has also been demonstrated. In this paper we have presented evidence that a crude membrane fraction of rat myometrial homogenate contains at least two kinds of binding sites for each of the two primary PGs, PGFB, and PGEZ. These PGs have been widely implicated in the mechanism of parturition in terms of promoting uterine contractility as well as


PGF2,

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FIG. 8. Effect of fetectomy on high (A)- and low (@-affinity binding of PGF2, in rat myometrial membrane preparation. Open bars, maximal binding capacity of PGFZCI receptors in pregnant uterine horns. Hatched bars, binding capacity of fetectomized horns, from which fetoplacental units were removed. Results are means t SE of four different experiments done in duplicate. Statistical analysis of data was performed using paired t test. * P c 0.05; ** P C 0.01 when compared with pregnant horns.

Labor Days of pregnancy

Post-partum

12n

Days of pregnancy

J

partum

FIG. 9. Effect of fetectomy on high (A)- and low (@-affinity binding of PGE2 in rat myometrial membrane preparation. Open bars, maximal binding capacity of PGEz receptors in pregnant uterine horns. Hatched bars, binding capacity of fetectomized horns. Results are means t SE of four different experiments done in duplicate. * P < 0.05; ** P < 0.01 by paired t test.

Daysof pregnancy

Post-partum

20 Labor Days of pregnancy

t* A

60 2 50 z

3

Iu

LOWHIGHLOWHIGHAFFINITY BINDING SITES AFFINITY BINDING SITES FIG. 10. Effect of indomethacin treatment on maximal binding ca-

pacities of high- and low-affinity binding sites for PGF2, (A) and PGE2 (B). Open bars, control rats; hatched bars, indomethacin (2 mg/kg body wt twice a day for 4 days)-treated rats. Four pairs of horns from each group were processed separately to measure PGF2, binding, and remaining two groups of four were treated identically to assess PGE, binding. Values are expressed as means t SE (n = 4). * P < 0.05; ** P < 0.01 by Student’s t test.

cervical ripening in humans (16, 19), lending support to a functional role for these binding sites. With the exception of the high-affinity PGE2 binding site, whose affinity for its ligandwas about IO-fold higher before parturition than after delivery, the apparent &s of PGF2, and the low-affinity I&., of PGEZ remained virtually constant during late pregnancy and after parturition and were not

12h

S Post- partum

significantly different from nonpregnant myometrial preparations. No differences in the binding affinities of PGF2, or PGE1 were observed in the human myometrium during the menstrual cycle and before and during labor (12) Concentration of PGF2, and PGE2 receptors did not change significantly until after parturition, when the specific uptake of the tritiated ligands increased. This finding is consistent with reports on PGFZ, receptors in human myometrium during pregnancy and labor (11,lZ) and would indicate that the striking changes in steroid hormone levels have little influence on PGF receptor concentration, contrary to what has been reported for oxytocin receptors (1, 28). This would also suggest that the putative role of these PGs in the initiation of parturition is mainly a function of increased production by various uterine compartments. However, because of a marked increase in uterine PG production, we considered the possibility that part of the receptors were masked by occupancy of endogenously produced PGs. This notion is supported by subsequent experiments, which showed that raising endogenous levels by direct injection of PGF2, and PGEz into one of the uterine horns 30 min before excision of both treated and control horns caused a 40-50% suppression of [3H]PG uptake by crude membrane fraction. Moreover, chronic treatment of pregnant rats with indomethacin, a potent inhibitor of PG synthase, at a dose that has been shown to inhibit PG production in vivo (24) caused a significant two- to


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threefold increase in the binding of both PGF2, and PGE,. An “upregulation” of PGE receptors in rat liver membranes has also been reported after 4day treatment with indomethacin or acetyl salicylic acid (24). It is well established that human fetal membranes are a rich source of PGs (3). Moreover, uterine stretch has also been shown to increase PG release (15). We eliminated both of these factors by unilateral fetectomy and compared PGF*,, and PGE, binding in myometrial preparations from the experimental uterine horns to that found in the contralateral pregnant ones. Using such an experimental approach, we have been able to show a twoto threefold increase in [“H]PGF2, binding and in the concentration of the high-affinity PGEZ receptors by the fetectomized myometrium at the time of parturition. These experiments seem to suggest that the concentration of PGF2,, receptors (and to a lesser extent PGEz receptors) increase around the time of parturition, but the estimation of such binding is partially masked by high endogenous uterine levels of these ligands. Although uterine venous blood levels of PGE2 were not measured (due to accidental loss of the PGE, fraction during chromatography), those of the amniotic fluid reached values by day 21 higher than any of the other prostanoids studied. In addition, the affinity of the high-affinity binding (Kd,) of PGEZ in the pregnant myometrium was found to be 5-10 times higher than the corresponding PGFg,, binding site. Thus a relatively small increase in PGEZ with approaching parturition may have a significant physiological impact. The observed changes in steroid hormone and PGFZ, levels in uterine venous plasma during late gestation and parturition are consistent with those reported by other investigators (22). It should be noted that a precipitous fall in progesterone levels, starting on day 18, preceded the rise in PGF2,, and PGF, in plasma, as well as these and PGE2 in amniotic fluid, whereas plasma estradiol levels peaked at day 21 simultaneously with PG levels. The role of steroid hormones in uterine PG synthesis has been extensively studied, with results favoring a stimulatory effect by estradiol (23). It would appear, however, that the inhibitory effect of progesterone may be the dominant influence. Indeed, it has been shown that administration of progesterone to ovariectomized rats decreased deacylation of phospholipids and the release of the prostanoid precursor arachidonic acid in uterine minces, as well as inhibiting the stimulant effect of estradiol on phosphatidylinositol turnover (30, 31). A rapid fall in progesterone production would be expected to remove this inhibition, allowing the effect of estrogen to be manifest, resulting in increased release of arachidonate and the synthesis of prostanoids. This study also provides evidence for a significant increase in thromboxane and prostacyclin production, particularly as reflected in amniotic fluid levels, shortly before parturition. Although these prostanoids may also have a hitherto undefined role in parturition, the observed increase may have resulted simply from the enhanced provision of the precursor arachidonate. Whether or not steroid hormones are also involved in the regulation of PG receptors, as has been demonstrated for

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uterine oxytocin (21, 28), angiotensin (27), and adrenergic receptors (29, cannot be ascertained from these experiments. It has been reported, however, that treatment of ovariectomized hamsters with estradiol dosedependently decreased, whereas progesterone increased the specific binding of PGE, and PGFZ, in uterine tissue (33). These steroid-induced changes in PG binding site concentration could be attributed to changing endogenous levels of PGF2, and PGEZ, assuming a slowly dissociating stable PG-receptor complex. On the other hand, a direct effect of estrogen and progesterone on PG receptor concentration cannot be ruled out. In fact, our results have shown that PG receptor concentration increased significantly with approaching parturition when rats were treated with indomethacin at dose levels that were expected to have inhibited endogenous PG synthesis. These observations, taken together with the significant increments of PG binding capacity of postpartum myometrial preparations when endogenous levels of PG fall, support the idea that PGF2, and PGE, exert their action in parturition by an increase in uterine PG production, as well as PG binding sites. In summary, we have demonstrated the existence of two specific binding sites each for PGEZ and PGFZa in rat myometrial preparations. There was no apparent change in the concentration of these binding sites before and during spontaneous parturition. However, experiments that altered endogenous levels of prostaglandins suggest that the number of binding sites may have increased, but these were partially masked by high concentrations of endogenous PG in the term pregnant uterus. This work was supported by National Institute of Child Health and Human Development Grant HD-09763, National Science Foundation Grant INT-8421360, and a grant from the Kerenyi Prenatal Research Fund, New York. Dr. Molnar is the recipient of a Fellowship from the Lalor Foundation. Address for reprint requests: F. Hertelendy, Dept. of Obstetrics and Gynecology, St. Louis University Medical Center, 3635 Vista Ave. at Grand Blvd., P. 0. Box 15250, St. Louis, MO 63110-0250. Received

2 August

1989; accepted

in final

form

2 January

1990.

REFERENCES 1. ALEXANDROVA, M., AND M. S. SOLOFF. Oxytocin receptors and parturition. I. Control of oxytocin receptor concentration in the myometrium at term. Endocrinology 106: 730-735, 1980. 2. ASBOTH, G., H. TODD, M. TOTH, AND F. HERTELENDY. PGE, binding, synthesis, and distribution in hen oviduct. Am. J. Physiol. 248 (Endocrinol. Metab. 11): E80-E88, 1985. 3. BLEASDALE, J. E., AND J. M. JOHNSTON. Prostaglandins and human parturition: regulation of arachidonic acid mobilization. Rev. Perinat. Med. 5: 151-191, 1985. M. M. A rapid and sensitive method for the quantita4. BRADFORD, tion of microgram quantities of protein utilizing the principle of protein-dye binding. Anal. Biochem. 72: 248-254, 1976. 5. BURTON, K. Determination of DNA concentration with diphenylamine. Methods Enzymol. 12: 163-166, 1968. 6. CALDER, A. The clinical use of prostaglandins for early and late abortion. In: Eicosanoids and Reproduction, edited by K. Hillier. Lancaster, UK: MTP, 1987, p. 184-194. M. E., AND J. D. MILLER. Prostaglandin Ez receptor in 7. CARSTEN, the myometrium: distribution in subcellular fractions. Arch. Biochem. Biophys. 212: 700-704, 1981. 8. CRANKSHAW, D. J., J. CRANKSHAW, L. A. BRANDA, AND E. E. DANIEL. Receptors for E type prostaglandins in the plasma membrane of nonpregnant human myometrium. Arch. Biochem. Bio1phys. 198: 70-77, 1979.


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actomyosin to parturition. In: Search 9. CSAPO, A. I. From uterine and Discovery: A Tribute to Albert Szent-Gyorgyi, edited by B. Kaminer. New York: Academic, 1977, p. 117-159. 10. DE LEAN, A., P. J. MUNSON, AND D. RODBARD. Simultaneous analysis of families of sigmoidal curves: application to bioassay, radioligand assay, and physiological dose-response curves. Am. J. Physiol. 235 (Endocrinol. Metab. Gastrointest. Physiol. 4): E97E102, 1978. 11. FUKAI, H., D. DEN, H. SAKAMOTO, H. KODAIRA, F. UCHIDA, AND S. TAKAGI. Study of oxytocin receptor: II. Oxytocin and prostaglandin FPtr receptors in human myometria and amnion-decidua complex during pregnancy and labor. Endocrinol. Jpn. 31: 565570,1984. 12. GIANNOPOULOS, G., K. JACKSON, J. KREDENTSER, AND D. TULCHINSKY. Prostaglandin E and FZtr receptors in human myometrium during the menstrual cycle and in pregnancy and labor. Am. J. Obstet. Gynecol. 153: 904-910, 1985. 13. GIMENO, A. L., AND M. F. GIMENO. Prostaglandins and the regulation of motility in non-pregnant isolated uterus. Trends Pharmacol. Sci. 5: 28-30, 1984. (Editors). Methods of Hormone 14. JAFFE, B. M., AND H. R. BEHRMAN Radioimmunoassay (2nd ed.). New York: Academic, 1987. 15. KLOECK, F. K., AND H. JUNG. In vitro release of prostaglandins from the human myometrium under the influence of stretching. Am. J. Obstet. Gynecol. 115: 1066-1069, 1973. 16. LIGGINS, G. C. Endocrinology of parturition. In: Fetal Endocrinology of Parturition, edited by M. J. Novy and J. A. Resko. New York: Academic, 1981, p. 211-237. 17. LIGGINS, G. C., C. S. FORSTER, S. A. GRIEVES, AND A. L. SCHWARTZ. Control of parturition in man. Biol. Reprod. 16: 39-56, 1977. 18. LINTNER, F., M. TOTH, AND F. HERTELENDY. Copurification of prostaglandin FZcr receptors with rat uterine plasma membranes. Experientia Base1 39: 1102-1103, 1983. 19. MCDONALD, P. C., J. C. PORTER, B. E. SCHWARTZ, AND J. M. JOHNSTON. Initiation of parturition in the human female. Semin. Perinatol. 2: 273-286, 1978. 20. MORRELL, R. M. Prostaglandin receptors and their functional interactions. In: Cell Membrane Receptors for Viruses, Antigens and Antibodies, Polypeptide Hormones and Small Molecules, edited by R. F. Beers, Jr., and E. G. Bassett. New York: Raven, 1976, p. 467-510.

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Menstrual induction with PGF2 alpha and PGE2.

Characterization of G proteins in rat myometrium. A differential modulation of Gi2 alpha and Gi3 alpha during gestation.

PGF2 alpha stimulates release of PGE2 and PGI2 in the isolated perfused rat heart.

Gap junctions: their presence and necessity in myometrium during parturition.

The placenta, PGE2 and parturition.

[On the effect of PGE2 and PGF2 alpha on circular and longitudinal myometrium strips of different corpus uteri tissues of the guinea pig in vitro (author's transl)].

Vasoactive effects of intra- and extravascular serotonin, PGE2 and PGF2 alpha in human umbilical arteries.

Changes in electrical properties of rat myometrium during gestation and following hormonal treatments.

Matrix Metalloproteinase Expression in the Rat Myometrium During Pregnancy, Term Labor, and Postpartum.

Intrarenal prostaglandins: effect of sodium and indomethacin on PGE2 and PGF2 alpha in rabbits.

Self-differentiation of human fetal lung organ culture: the role of prostaglandins PGE2 and PGF2 alpha.

An ultrastructural study of the testis and epididymis of the rat after treatment with prostaglandins E2 and F2 alpha (PGE2 and PGF2 alpha).

[Perioperative behavior of prostaglandin E2 (PGE2) and 13,14-dihydro-15-keto-PGF2 alpha (DHK-PGF2 alpha) in the serum in bronchial cancer].

Eicosanoids PGF2 and PGE2 alpha enhance the attachment of rabbit tracheal epithelial cells to a collagen substrate.

Vaginal administration of PGE2 for induction of labor stimulates endogenous PGF2 alpha production.

Luteal expression of thyroid hormone receptors during gestation and postpartum in the rat.

Cytosolic free calcium increased by prostaglandin F2 alpha (PGF2 alpha), gonadotropin-releasing hormone, and angiotensin II in rat granulosa cells and PGF2 alpha in human granulosa cells.

A comparison of PGE2 and PGF2 alpha vaginal gel for ripening the cervix before induction of labour.

PGF2 alpha, PGE2, and sex steroids from the abdominal gland of the male crested newt Triturus carnifex (Laur.).

Fine structure of foetal rat pancreatic alpha and beta cells at term and during prolonged gestation.

Immunohistochemical localization of PGF2 alpha receptor in the rat ovary.

Glycogen levels in the rat myometrium at the end of pregnancy and immediately postpartum.

Proliferative effect of PGD2 on osteoblast-like cells; independent activation of pertussis toxin-sensitive GTP-binding protein from PGE2 or PGF2 alpha.

Corpus luteum susceptibility to prostaglandin F2 alpha (PGF2 alpha) luteolysis in hysterectomized prepuberal and mature gilts.