Influence of the embryo on the distribution of maternal immunoglobulins in the mouse uterus

The effect of the embryo on the distribution of IgA, IgG and IgM was studied by an immunoperoxidase technique on mouse uterine sections, (1) during the first part of pregnancy and pseudopregnancy, and (2) in delayed implantation combined with different progesterone-oestradiol treatments designed to extend the delay or induce implantation, and in nonpregnant ovariectomized mice similarly treated. The number of glandular lumina containing IgA increased particularly from the implantation period, but in pseudopregnancy this number decreased from the morning of Day 4, and afterwards continued to decline. In delayed implantation, the number of glandular lumina containing IgA also rose considerably when implantation was induced by oestradiol, whereas under the same progesterone-oestradiol treatment, nonpregnant ovariectomized animals displayed no such increase. Significant staining for IgG in the stroma was observed on Day 4 of pregnancy and pseudopregnancy but prolonged staining for IgG was observed only during pregnancy. In addition, significant numbers of IgA-plasma cells in the stroma were observed mostly in uteri containing embryos. These results indicate that embryos might affect the process by which ovarian hormones regulate IgA and IgG distribution.     

Partial characterization of a luteal factor that induces implantation in the ferret

This study was designed to test the hypothesis that ferret corpora lutea (CL) secrete a compound that acts in conjunction with progesterone to induce blastocyst implantation and to identify the chemical nature of this compound. CL and the residual ovarian tissue, obtained predominantly on the ninth day of pseudopregnancy, were extracted with 0.05 M phosphate-buffered saline. The extracts were injected into pregnant ferrets that had been ovariectomized on Day 6 of pregnancy and had received Silastic implants containing progesterone. Aqueous luteal extracts, but not those of the residual ovarian tissue, induced implantation in test animals. Fractionation of the luteal extracts by passage through a series of filters with molecular weight (MW) cutoffs ranging from 500 to 50,000 consistently revealed that the biologically active fraction was retained on the filter with the highest MW cutoff employed. Moreover, blastocyst implantation failed to occur in ovariectomized, progesterone-treated ferrets after one-half of a luteal preparation (MW greater than 50,000) was incubated with a broad-spectrum protease. These data are consistent with the hypothesis that CL of the ferret secrete a protein during the preimplantation period that is essential for blastocyst implantation.     

Traumatic deciduoma and delayed nidation in castrated pregnant rat after ACTH injection]

After administration of ACTH it is possible to obtain traumatic deciduomata in rats ovariectomized during early pregnancy and delayed implantation in rats ovariectomized day 4 of pregnancy and injected dayly with oestradiol from day 8 or 9. The substance responsible seems to be Progesterone or Progesterone like corticoid.     

Failure of platelet-activating factor (PAF-acether) to induce decidualization in mice and failure of antagonists of PAF to inhibit implantation

The possible role of platelet-activating factor (PAF) in the uterine responses associated with implantation was investigated. Attempts to trigger a decidual cell response in the uteri of hormonally sensitized, ovariectomized mice by instilling PAF-acether (1-1000 ng) intraluminally were unsuccessful. The effect of PAF antagonists on implantation was investigated in females ovariectomized on Day 3 of pregnancy and treated with progesterone. Implantation was induced in these females by injection of 10 ng oestradiol-17 beta on Day 8. Hourly intraperitoneal injections of three PAF antagonists (WEB 2086, CV 3988 and BN 52021 at doses of 1.2-1.4 mg/kg) given over a 24-h period starting 1 h before the injection of oestradiol-17 beta had no significant effect on the occurrence of implantation sites. Intraluminal injection of WEB 2086 (15 micrograms) or BN 52021 (5 micrograms) either 3 h before or 6 h after the nidatory oestradiol also had no significant inhibitory effect on implantation. SRI 63-441 given once daily over the first 4 days of pregnancy at a dose of 40 micrograms/30 g body weight had no inhibitory effect on the establishment of pregnancy. These results are not consistent with a critical role for PAF in implantation in mice.     

Effects of medroxyprogesterone acetate on gestation in mink

Mink ovariectomized 14 days after the first of two matings received injections of 2 mg MPA, the same MPA treatment + an oestradiol-17 beta implant or no replacement therapy. Some mink were ovariectomized after implantation and given a single dose of 2 mg MPA or no replacement therapy. MPA persisted in the serum at detectable levels for 13 or more days in all mink treated. MPA and MPA + oestradiol induced uterine growth but neither treatment was capable of inducing embryo implantation. More embryos were retained in mink treated with MPA alone and these appeared to be viable. Implanted embryos persisted for a longer period in animals that were ovariectomized and treated with MPA. MPA neither supported pregnancy nor permitted parturition. Serum LH was elevated by 1 week after ovariectomy and elevations persisted for a further 20 or more days. While MPA alone had no apparent negative feedback effects on LH, animals that received MPA + oestradiol did not display any elevation of LH, suggesting that oestradiol or a combination of MPA and oestradiol has a potent negative feedback in mink.     

Effects of 9-ene-tetrahydrocannabinol on uterine estrogenicity in the mouse.

Several early (Phase I) and late (Phase II) estrogenic effects of 9-ene-tetrahydrocannabinol (THC) were examined in the adult mouse uterus. An injection of THC (2.5 or 10 mg/kg body wt) in ovariectomized mice neither stimulated uterine water imbibition or accumulation of [125I]bovine serum albumin (Phase I responses) at 6 h, nor antagonized these Phase I responses elicited by estradiol-17 beta (E2). With respect to Phase II responses, although single injections of THC (2.5, 5.0 and 10 mg/kg body wt) alone were ineffective in influencing uterine weight at 24 h or incorporation of [3H]thymidine at 18 h, this drug interfered with these responses elicited by E2 in a dose-dependent manner. In contrast, an injection of THC in progesterone (P4)-primed ovariectomized mice modestly enhanced (61%) uterine incorporation of [3H]thymidine. However, E2-stimulated uterine thymidine incorporation in P4-primed ovariectomized mice was antagonized by THC treatment. Effects of THC on blastocyst implantation were examined. Single or multiple injections of various doses of THC neither induced implantation in P4-primed delayed implanting mice, nor interfered with E2-induced implantation. Furthermore, daily injections of THC (10 mg/kg body wt) during the peri-implantation period had no apparent adverse effects on implantation, or on experimentally induced decidualization (deciduomata). The data suggest that THC is neither pro- nor antiestrogenic with respect to Phase I responses. However as regards Phase II responses, THC is modestly pro-estrogenic in the P4-treated uterus, but is anti-estrogenic in the presence of E2. These estrogen agonistic/antagonistic effects of THC on uterine Phase II responses do not adversely affect the process of implantation and decidualization.     

The effects of ovarian steroids in controlling rat uterine prostaglandin F2-alpha during the peri-implantation period

Rats with delayed implantation, induced by ovariectomy or hypophysectomy, as well as those with normal pregnancy were used to examine the changes in uterine prostaglandin F2 alpha (PGF2 alpha) associated with implantation. In normal pregnant rats, while maximal uterine production of PGF2 alpha was found at 09:00, maximal catabolic enzyme activity (CEA) was seen at 17:00 of day 4. Uterine content of PGF2 alpha was high at 17:00 of day 4, but decreased by 80% within the next 24 h. There was no change in PGF2 alpha production during the first 6 h after injection of estradiol to hypophysectomized animals. There was, however, a dramatic decrease in production within the next 6 h. In contrast, CEA was not different in animals treated with estrogen than in those receiving only progesterone. In ovariectomized animals, uterine PGF2 alpha production also was lowered by estrogen but in these animals CEA was significantly elevated 18 h after injection of estradiol. Estrogen caused a greater increase in PGF2 alpha content in the hypophysectomized, compared to the ovariectomized, rats. The results are consistent with the view that ovarian steroids play an important role in controlling the changes in uterine PGF2 alpha around the time of implantation in rat.     

Effects of oestradiol-17 beta and progesterone on the number of plasma cells in uteri of ovariectomized mice

Immunoglobulins A and G were localized by immunoperoxidase labelling in uteri of ovariectomized mice treated with oestradiol-17 beta and progesterone. The administration of oestradiol or progesterone alone to ovariectomized mice for 3 days increased the number of IgA plasma cells from about 1 to 14 per histological section. When the two hormones were administered simultaneously for 3 days the number of plasma cells per section was equal to or greater than with either hormone alone. Treatment with oestradiol followed by progesterone in a sequence that prepares the uterus for implantation resulted in about 31 IgA plasma cells per section. Counts of IgG plasma cells showed similar trends but the numbers were smaller. The results indicate that progesterone increases rather than decreases the number of plasma cells in the mouse uterus. This is consistent with observations on intact mice during oestrus and pregnancy and suggests that the marked increase in endometrial plasma cells at the time of implantation in mice is a response to progesterone acting on an oestrogen-primed uterus.     

Reproduction in the ferret. I. The effect of ovariectomy on the course of gestation.

The role of the ovaries in the maintenance of pregnancy was studied in the ferret. Ovariectomy at the time of implantation showed that some embryos survived for 7 days after the operation but all were destroyed after 10 days, although the trophoblast continued to grow at a much faster rate than normal. Ovariectomy performed after implantation showed that no fetal development occurred when the ovaries were removed at Day 21 post coitum, but that fetuses developed for an appreciable length of time in animals ovariectomized on Days 23 to 27 post coitum. Ovariectomy in late gestation resulted in speedy expulsion of the fetuses. An increase in the placenta:fetus ratio did not alter the response to ovariectomy in late gestation. The uteri in all ovariectomized animals showed progestational endometria when examined shortly after explusion of the fetuses.     

Implantation, deciduoma formation and live births in mast cell-deficient mice (W/Wv)

The intraluminal injection of oil produced deciduoma formation in ovariectomized, mast cell-normal (+/+) and mast cell-deficient (W/Wv) mice that were treated with exogenous steroids. Oil injection and trauma (e.g. sutures) also produced a deciduoma in ovariectomized +/+ and W/Wv mice that had received a single control (+/+) ovary transplanted under the kidney capsule. After transfers of donor blastocysts, implantation and live births were obtained in +/+ and W/Wv mice containing a single ovary transplant. Our results demonstrate that uterine mast cells are not required for the production of a decidual cell response, implantation, gestation or the birth of live offspring in mice.     

Catechol estradiol induced implantation in the mouse

Induction of implantation is among the most sensitive responses to estrogens. The ability of catechol estradiols, 4-hydroxy-estradiol-17 beta (4-OH-E2) and 2-hydroxy-estradiol-17 beta (2-OH-E2), to induce implantation in ovariectomized pregnant mice was compared to that of estradiol-17 beta. Delayed implantation was maintained by the daily administration of 2 mg of progesterone. A single injection of 3 ng of estradiol-17 beta, 50 ng of 4-OH-E2, or 2,000 ng of 2-OH-32 consistently induce a full complement of implantation sites in all animals. Before the estrogenicity of the latter steroid can be established the lack of contaminating estrogens must be proved.     

Initiation of embryo implantation and maintenance of early pregnancy in the rat by chlordecone (Kepone).

The effect of chlordecone (Kepone), an insecticide/fungicide with reproductive toxicity, on the early stages of pregnancy in the rat was studied. Intraperitoneal injection of chlordecone into adult virgin female Holtzman strain rats before mating, in doses as high as 80 mg/kg, did not prevent fertilization, early development of the embryo to the blastocyst stage, transport of the embryo through the oviduct, or its implantation into the uterus. However, a single dose of 60 or 80 mg/kg, but not 20 or 40 mg/kg, before mating significantly reduced the concentration of progesterone in the serum of rats undergoing normal embryo implantation 5 days later. A dose of 80 mg/kg of chlordecone reduced progesterone levels in the serum by more than 50% within 48 hr in ovariectomized rats with Silastic tubing implants containing crystalline progesterone. This dose of chlordecone induced deciduomata formation in progesterone-primed ovariectomized rats to the same extent as 1 microgram of estradiol benzoate. The minimal effective single dose of chlordecone to initiate implantation of blastocysts in the uteri of hypophysectomized progesterone-primed rats, and to maintain embryo development for at least 5 days, was 50 mg/kg. Daily doses of 20 mg/kg for 3 or 5 days were effective at initiating implantation but did not maintain pregnancy. The latter treatment, however, did not prevent initiation of implantation or embryo development induced by subsequent administration of estrone. The results are consistent with the view that chlordecone is a weak estrogen that has both nongenomic and genomic estrogenic actions.     

Implantation in ovariectomized mice treated with dibutyryl adenosine 3',5'-monophosphate (dibutyryl cyclic AMP).

Experiments are described that demonstrate that uterine intraluminal injection of a 1-25 mM-solution of dibutyryl cyclic AMP (dcAMP) in phosphate buffered saline (PBS) induced implantation in ovariectomized pregnant mice. Pretreatment with progesterone was essential for this effect. When PBS was injected alone, it did not induce implantation in mice treated with progesterone. Bilateral adrenalectomy had no effect on the ability of dcAMP to substitute for oestradiol, showing that the effect was not due to dcAMP-induced oestrogen synthesis in the adrenal cortex. It is suggested that the dcAMP may act at the level of the uterus, the embryo, or both.     

Effect of an estrogen antagonist on development of blastocysts and implantation in the hamster

CI 628 citrate, an estrogen antagonist, was given as a single intraluminal injection (5 micrograms) on day 3 to ovariectomized, progesterone-treated hamsters. This significantly reduced embryo cleavage rate, transformation of morula into blastocyst, and completely inhibited implantation. The effects of the drug could be reversed by estradiol-17B (1 microgram) but not estradiol-17 alpha (1 microgram) injected intraluminally with CI 628 citrate. Our finding suggests a role of estrogen present in hamster preimplantation embryo in the triggering of embryonic differentiation and implantation of the blastocyst.     

Direct evidence for the involvement of prostaglandin F2 alpha in the first step of estrone-induced blastocyst implantation in the spayed rat.

The effect of prostaglandin F2 alpha (PGF2 alpha) on blastocyst implantation in spayed rats has been studied. In preliminary experiments, the first implantation sites were observed 8 - 12 hours after a single injection of estrone in ovariectomized and progesterone-conditioned rats. Intraluminal instillation of PGF2 alpha into the right uterine horn 8 - 10 h after the estrone injection increased the number of implantation sites. Even treatment with PGF2 alpha without previous estrone injection induced the first step of blastocyst implantation as shown by uterine dye site reaction (Niagara-blue test). The results are discussed with regard to the possible role of PGF2 alpha in the regulation of the blastocyst implantation processes in the rat.     

Qualitative patterns of protein synthesis in the preimplantation mouse embryo. II. During release from facultative delayed implantation.

Qualitative patterns of newly synthesized protein were examined in mouse embryos released from facultative delayed implantation either by removal from the uterus with subsequent culture in vitro, or by the administration of progesterone and estradiol 17β (in vivo) to the ovariectomized mother. It was found that the qualitative pattern of protein synthesis during the first 25 hr following release from delay is the same as the pattern obtained from normal blastocysts immediately prior to implantation. These results are discussed in relation to possible mechanisms of activation of delayed blastocysts.     

Oxytocic activity of plasma and posterior pituitary lobe after ovariectomy and implantation of stilboestrol or progesterone tablets in female rats

The purpose of this work was to compare the oxytocic activity of plasma and posterior pituitary lobe extract in rats after sham operation, ovariectomy and after subcutaneous implantation of stilboestrol or progesterone tablets in ovariectomized rats. On the 5th day after ovariectomy or implantation of hormones, sample of 2 ml of blood were obtained under urethane anaesthesia from the cephalic end of the right external jugular vein, and the animals were killed by decapitation. The posterior pituitary lobe was removed and homogenized in 0.9% NaCl solution acidified with glacial acetic acid. The oxytocic activity of plasma and extracts of the posterior pituitary lobe was determined by the method of Van Dongen and Hays on fragments of lactating rat mammary tissue. On the 5th day after ovariectomy or implantation of stilboestrol the oxytocic activity was found to be significantly increased in the plasma and posterior pituitary lobe, and after progesterone implantation it was decreased in the posterior pituitary lobe.     

Blastocyst implantation in ovariectomized, adrenalectomized hamsters treated with inhibitors of steroidogenesis during the pre-implantation period

The possible involvement of blastocyst estrogen production in the initiation of implantation, as indicated by the presence of areas of increased endometrial vascular permeability on Day 5 of pregnancy, was examined in hamsters. The animals were ovariectomized and adrenalectomized on Day 3 to remove maternal sources of estrogen, and pregnancy was maintained with medroxyprogesterone acetate. Aminoglutethimide phosphate (AGP) and cyano-ketone, inhibitors of steroidogenesis, administered from Days 3 to 5 of pregnancy, did not affect the proportion of hamsters in which implantation was initiated. However, the AGP treatment was associated with a lower proportion of embryos, recovered on Day 4, which were blastocysts, fewer implantation sites on Day 5, and smaller implantation swellings on Day 9. AGP treatment had no significant effect on the uterine concentrations of prostaglandins (PGs) of the E series, which were higher in the implantation sites than elsewhere in the uterus on Day 5. These results suggest that neither maternal nor blastocyst estrogen production is essential for the initiation of implantation in the hamster. In addition, the data suggest that the localized elevated PGE concentrations at implantation sites are induced by a blastocyst signal which is independent of blastocyst steroidogenesis.     

LH discharge induced by medial preoptic implantation of estrone or dopamine in the ovariectomized estradiol primed rat.

Effect of implantation of estrogens or catecholamines into the medial preoptic area through chronically implanted double cannula on the release of LH was examined in the ovariectomized estradiol-primed rat. Implantation of estrone at 12:00 on the second or third day after subcutaneous injection of estradiol benzoate (20 mug) increased the serum concentration of LH at 18:00 compared with that in the non-implanted controls, whereas implantation of estradiol benzoate on the third day did not affect it. Dopamine implantation at 12:00 on the third day also induced a significant increase of LH concentration at 18:00, and, in contrast, norepinephrine implantation decreased the concentration at 18:00. It may be said that the medial preoptic area is responsive to estrone and can induce LH release, whereas it is not to estradiol. Furthermore, dopamine was effective for the activation of the medial preoptic area in relation to the inducement of LH release.