Tumour necrosis factor-α is the signal induced by mating to shutdown a 2-methoxyestradiol nongenomic action necessary to accelerate oviductal egg transport in the rat.

Mating shut down a 2-methoxyestradiol (2ME) nongenomic action necessary to accelerate egg transport in the rat oviduct. Herein, we investigated whether tumour necrosis factor-α (TNF-α) participates in this mating effect. In unmated and mated rats, we determined the concentration of TNF-α in the oviductal fluid and the level of the mRNA for Tnf-a (Tnf) and their receptors Tnfrsf1a and Tnfrsf1b in the oviduct tissues. The distribution of the TNFRSF1A and TNFRSF1B proteins in the oviduct of unmated and mated was also assessed. Finally, we examined whether 2ME accelerates oviductal egg transport in unmated rats that were previously treated with a rat recombinant TNF-α alone or concomitant with a selective inhibitor of the NF-κB activity. Mating increased TNF-α in the oviductal fluid, but Tnf transcript was not detected in the oviduct. The mRNA for TNF-α receptors as well as their distribution was not affected by mating, although they were mainly localized in the endosalpinx. Administration of TNF-α into the oviduct of unmated rats prevented the effect of 2ME on egg transport. However, the NF-κB activity inhibitor did not revert this effect of TNF-α. These results indicate that mating increased TNF-α in the oviductal fluid, although this not associated with changes in the expression and localization of TNF-α receptors in the oviductal cells. Furthermore, TNF-α mimicked the effect of mating on the 2ME-induced egg transport acceleration, independently of the activation of NF-κB in the oviduct. We concluded that TNF-α is the signal induced by mating to shut down a 2ME nongenomic action in the rat oviduct.

Aberrant cannabinoid signaling impairs oviductal transport of embryos.

Ectopic pregnancy is a major reproductive health issue. Although other underlying causes remain largely unknown, one cause of ectopic pregnancy is embryo retention in the fallopian tube. Here we show that genetic or pharmacologic silencing of cannabinoid receptor CB1 causes retention of a large number of embryos in the mouse oviduct, eventually leading to pregnancy failure. This is reversed by isoproterenol, a beta-adrenergic receptor agonist. Impaired oviductal embryo transport is also observed in wild-type mice treated with methanandamide. Collectively, the results suggest that aberrant cannabinoid signaling impedes coordinated oviductal smooth muscle contraction and relaxation crucial to normal oviductal embryo transport. Colocalization of CB1 and beta2-adrenergic receptors in the oviduct muscularis implies that a basal endocannabinoid tone in collaboration with adrenergic receptors coordinates oviductal motility for normal journey of embryos into the uterus. Besides uncovering a new regulatory mechanism, this study could be clinically relevant to ectopic pregnancy.

Effect of oestradiol and progesterone on the instant and directional velocity of microsphere movements in the rat oviduct: gap junctions mediate the kinetic effect of oestradiol.

The oviducal transport of eggs to the uterus normally takes 72-96 h in the rat, but this is reduced to less than 20 h after a single injection of oestradiol (E2). This accelerated transport is associated with an increased frequency of pendular movements in the isthmic segment of the oviduct, with increased levels of the gap junction (GJ) component Connexin (Cx) 43, and is antagonised by progesterone (P). In the present study, we investigated the effect of these hormones on the instant and directional velocity of pendular movements and the role of the GJ and its Cx43 component in the kinetic response of the oviduct to E2 and P. Using microspheres as egg surrogates, microsphere instant velocity (MIV) was measured following treatment with E2, P or P + E2, which accelerate or delay egg transport. Microspheres were delivered into the oviduct of rats on Day 1 of pregnancy and their movement within the isthmic segment was recorded. Oestrogen increased MIV with faster movement towards the uterus. After P or P + E2, MIV was similar to that in the control group. Two GJ uncouplers, namely 18 alpha- and 18 beta-glycyrrhetinic acid, blocked the effect of E2 on MIV. Connexin 43 mRNA levels increased over that seen in control with all treatments. In conclusion, the effects of E2 on MIV resulted in faster movements that produced accelerated egg transport towards the uterus. Gap junctions are probably involved as smooth muscle synchronisers in this kinetic effect of E2, but the opposing effects of E2 and P are not exerted at the level of Cx43 transcription.

Egg transport in ovariectomized rabbits as affected by different combinations of oestrogen and progesterone.

A total of 1548 eggs was transferred to the oviducts of rabbits ovariectomized 45 days to 7 months earlier. The pattern of egg transport was disrupted, and the majority of eggs were in the vagina 60-72 h after transfer. To determine the role of the ovarian hormones on egg transport, the effects of various combinations of oestrogen and progesterone were studied. None of the hormonal treatments produced the normal transport pattern and large proportions of transferred eggs were retained in the oviducts. It is concluded that as long as progesterone is the dominant hormone, eggs move very slowly through the isthmus, and that a surge of oestrogen is required to modify the action of progesterone and to speed the movement of eggs to the uterus.

Effects of 4-hydroxyandrostenedione and exogenous testosterone on blood concentrations of oestradiol and oviducal embryo transport in the rat.

The effect of decreasing oestrogen secretion on the oviducal migration of embryos was investigated in pregnant rats. The reduction of oestradiol production was achieved by administration of the aromatase inhibitor 4-hydroxy-4-androstene-3,17-dione (4-OH-A) at various times after coitus. When 4-OH-A was administered from days 2 to 5, nearly half the embryos were retained in the oviducts at midday on day 5 of pregnancy, in contrast with control animals in which all embryos were transferred to the uterus. Shorter treatments were less effective. The rate of secretion of oestradiol from the ovary on days 2-5 of pregnancy in control rats was low in the morning and high in the afternoon. Treatment with 4-OH-A from days 2 to 5 reduced the secretory surges of oestradiol in the afternoon by 77% without significantly changing the progesterone output. Systemic testosterone levels were significantly increased by this treatment. To assess whether changes in the transport of ova were due to an increase in testosterone concentrations the influence of exogenous testosterone on embryo transport and oestradiol production was tested. Testosterone administered by subdermal implants from days 2 or 3 to day 5 disturbed embryo transport in a manner similar to that of 4-OH-A. The longest period of testosterone administration decreased ovarian oestradiol production by 82% without changing the secretion of progesterone.(ABSTRACT TRUNCATED AT 250 WORDS)

Action of anti-progesterone monoclonal antibody in blocking pregnancy after postcoital administration in mice.

Anti-progesterone monoclonal antibody injected intraperitoneally as a single dose 32 h post coitum completely blocked pregnancy in BALB/c and CBA nulliparous mice. The dose required was greater in CBA than in BALB/c females, but in both strains no implantation sites were detectable at autopsy on day 10 after mating. The action of the antibody in BALB/c females was associated with a failure to initiate an implantation response as indicated by the Pontamine Blue reaction. The most pronounced effect, however, was an arrest of embryonic development at a stage prior to cavitation. Plasma progesterone concentration in blood taken by cardiac puncture was greatly increased after treatment, by virtue of high-affinity binding by antibody in circulation. The results show that passive immunization against progesterone shortly after mating interferes with early hormone-dependent steps which are essential for normal embryonic development.

Immunoglobulins and monoclonal anti-blastocyst antibodies in the mouse uterine secretion during the early pre-implantation period.

A Sepharose bead blot technique was used to study immunoglobulins in the uterine secretion of mice during the pre-implantation stage. Secretion collected by Sepharose beads contained IgA, IgG, and IgM. The method could be made ten-fold more sensitive by using anti-mouse IgG or IgM conjugated to Sepharose beads. It has also been demonstrated that when injected intravenously, biotinylated purified immunoglobulins, both non-specific mouse myeloma IgG and IgM and specific anti-blastocyst IgG and IgM, is able to pass into the uterine cavity of the mouse. It was further shown that when injected systemically, anti-blastocyst antibodies can reach the blastocyst. Functionally active specific antibodies against morulae and/or blastocysts may, therefore, be able to influence the pre- and peri-implantation development of the embryo and could serve as a useful model for experiments directed towards the identification of immunological contraceptive procedures.

Temporal relationships critical to progesterone-induced acceleration of ovum transport.

Previous investigators have demonstrated that 2.5 mg fo progesterone, administered intramuscularly to rabbits on the day of ovulation and the 2 preceding days (Days -2, -1, and 0) significantly and consistently accelerates ovum transport. In contrast, when given on the day of ovulation and the 2 following days (Days 0, +1, and +2), progesterone does not accelerate ovum transport. The experiments reported were designed to define more precisely the temporal relationships critical to progesterone-induced acceleration of tubal ovum transport. Our observations suggest 3 important conclusions: 1) Progesterone, when given at least 1 day, and not more than 2 days, prior to ovulation does induce accelerated ovum transport. 2) The progesterone responsive mechanism is dose dependent. 3) The acceleration is partially antagonized if progesterone treatment is begun 3 days prior to ovulation.

PIP: These experiments were designed to determine the time relationship of progesterone administration to ovulation and to ovum transport rates in the rabbit. Ovulation was induced in New Zealand white does by iv injection of 100 units of human chorionic gonadotropin (hCG). The day this was given was designated Day 0. All animals were killed 24 hours after injection, oviducts removed, and ova counted. Ova in the uterine horns were also counted and ovulation points on the ovaries recorded. In control animals all ova were recovered in the oviducts. When 2.5 mg progesterone had been given 1 or 2 days before Day 0 only 34% of the ova were in the oviducts and 41% were in the uteri. When 2.5 mg progesterone was given 3 days before or on Day 0, the accelerated ovum transport did not occur. Treatment by progesterone for 4 days, starting 3 days before ovulation, diminished ovum acceleration. When 7.5 mg progesterone were given in a single dose either 3 days before or on Day 0, ovum transport was not altered but when given 1 or 2 days before Day 0 maximum ovum acceleration was induced. Results show that the effects of progesterone on ovum transport are related to both time and dose administered. To induce acceleration of ovum transport progesterone injection must be given earlier than 12 hours but not more than 60 hours before ovulation (p less than .001). This result is dose-dependent and is partially reduced if progesterone is given 3 days prior to ovulation. If the same holds true for humans, the results indicate that progester one should be given as a single dose within 36-60 hours prior to ovulation or about the time of the blood estrogen peak.

Effect of particle size on time course of transport of surrogate ova through the rabbit oviduct.

The time-course of transport of plastic microspheres of 100, 200, 400, 800, and 1000 mu diameter through the rabbit oviduct was studied under various hormonal conditions. During estrus, the rabbit oviduct rapidly transports particles of 200 mu diameter to the uterus. After ovulation, the passage of these particles is delayed, first at the ampullary-isthmic junction, and then along the isthmus, perhaps influenced by a second site of resistance at the uterotubal junction. Particles of 100 mu diameter are not subject to these delays but pass rapidly to the uterus. Our data support the idea that the rapid passage of 100-mu spheres reflects a size-related inability of the resisting "gates" to limit their passage. Pharmacologic doses of estrogen act to prolong or increase the resistance to particle passage at the ampullary-isthmic junction and apparently also accelerate transport through the isthmus. Progesterone presumably increases the efficiency of the "gate" at the ampullary-isthmic junction, hastens transport through the isthmus, and seems to "open the gate" at the utero-tubal junction. Further experiments are required to prove or negate these inferences.

Probable influence of ova and embryo prostaglandins in the differential ovum transport in pregnant and cycling rats.

In this study we explored the possible underlying mechanism(s) of the differential transport of unfertilized and fertilized ova in cycling and pregnant rats. The number of ova recovered from rat oviducts and uterus was not significantly different in estrus, metestrus and diestrus but dropped sharply at proestrus. When estrus rats were injected with indomethacin (10(-6)), a well known inhibitor of cyclooxygenase, delivered into both ovarian bursae, and sacrificed next day at metestrus, the number of ova in the oviduct was significantly smaller (p less than 0.025) than in controls at metestrus. On the other hand, when diestrus rats were injected with PGE1 (10(-6)) delivered into both ovarian bursae, and sacrificed next day at proestrus, no ova were found in the oviducts, and only a few of them were in the uterus. When fertilized ova were recovered from oviducts and uteri at day 4 of pregnancy (corresponding to proestrus of cycling rats) an average of 4 embryos were still found in the oviducts, proving a differential ovum transport between cycling and pregnant rats. In order to establish if there exists any ova or embryo releasing factor responsible for this difference, the prostaglandins released to the incubation medium by ovum or 3-day embryo were measured. Unfertilized ova produced significantly more PGE1 (p less than 0.05) than PGE2 or PGF2 alpha. The same pattern of PG production was observed with incubated embryos, but in this case the amount of PGE1 released was significantly higher (p less than 0.01) that the PGE1 released by unfertilized ova.(ABSTRACT TRUNCATED AT 250 WORDS)

Influence of ova within rat oviducts on spontaneous motility and on prostaglandin production.

The present study was performed in order to explore the influence of ova present within rat oviducts on: a) tubal spontaneous motility and b) oviduct prostaglandin production. It was found that the isometric developed tension (IDT) of tubes isolated from proestrous rats (preovulatory oviducts) was significantly higher (P less than 0.01) than the IDT of tubes from rats at estrus and at metestrus (postovulatory oviducts). After flushing the oviducts with KRB solution (i.e., after removing existing ova) the IDT of the oviducts obtained from estrous rats increased significantly (P less than 0.01), whereas the IDT of tubes isolated from proestrous rats (i.e., preparations without ova) was not modified. On the other hand, isolated tubes containing their corresponding ova released into the suspending solution significantly more PGE1 than PGE2 or PGF2 alpha (P less than 0.005). It was particularly interesting to find that after flushing the oviducts, tissue production of PGE1, PGE2 and PGF2 alpha was similar. Finally, when dose response curves for PGE1 and for PGE2 on the spontaneous contractions of oviducts isolated from rats at proestrus, estrus and metestrus were constructed, both PGs evoked an inhibitory inotropic action. The ED50 for PGE1 in tubes from estrous rats was significantly smaller (P less than 0.01) than that for metestrous animals but significantly greater (P less than 0.01) than that observed in oviducts from proestrous rats. The ED50 for PGE2 did not change in the different tested periods of the sex cycle. Results reported herein suggest the possibility that the ova present within rat oviducts, may influence their own transport along the tubes by modifying the amount of prostaglandins produced by the oviducts or via their own prostaglandin synthesis.

Temporal relationships critical to estrogen-induced delay of ovum transport.

Previous studies have shown that a pharmacologic dose of estradiol given at the time of injection of human chorionic gonadotropin (HCG) significantly delays the transport of ova through the rabbit oviduct and that the site of delay is located at the ampullary-isthmic junction. The current experiments investigated the effects of varying the timing of the estrogen injection on ovum transport rates. Fifty-eight rabbits received injections of either Depot-estradiol or crystalline estradiol at various times before or after the injection of HCG. "Tube-locking" resulted when estrogen was administered before or at the time of HCG administration, whereas acceleration occurred when estrogen was given 12 to 24 hours after HCG. The results demonstrate that pharmacologic doses of estrogen exert two effects on ovum transport in the rabbit: (1) delay at the ampullary-isthmic junction and (2) acceleration once that junction has been traversed. The possible application of these results to contraception is discussed.

Egg transport in the rabbit oviduct following ampullary resection and microsurgical end-to-end anastomosis.

Surgical alteration of rabbit oviducts was employed to investigate the mechanisms of ovum transport. Resection of a 5- to 10-mm segment of midampulla was followed by microsurgical end-to-end anastomosis. One to two months later, direct observations were made, in situ, of ampullary egg transport. Stained cumulus masses were transported normally to the site of anastomosis: two-thirds of those studied were delayed at that point for an average of 2 1/2 minutes. Normal transport then resumed and continued to the ampullo-isthmic junction. In all but one instance, blockage of muscular activity with isoproterenol prevented transport beyond the anastomosis site, demonstrating the presence of an obstacle to ciliary transport. These studies reveal an important facilitative role for muscle in improving the reliability of ampullary egg transport. Considering species differences in egg transport mechanisms, it is suggested that ampullary-ampullary anastomosis in women might have a poor prognosis for re-establishing fertility.

Pharmacologic modification of the time course of ovum transport in guinea pigs.

Various agents were examined for their effects on ovum transport in the guinea pig. Estrogen significantly accelerated ovum transport in this species. The experiments further demonstrated that estrogen did not act by inducing prostaglandin synthesis, nor by altering plasma progesterone levels. The estrogen-induced acceleration was significantly antagonized by tamoxifen, an antiestrogen that acts by interfering with estrogen receptor synthesis. Cycloheximide also antagonized the effects of estrogen on ovum transport. These data suggest that the modification of ovum transport by estrogen is due to the entrance of estrogen into the nuclei of target cells, and subsequent protein synthesis. Although we assume that this action occurs at the level of the oviduct, our experiments do not prove this assumption.

PIP: The effects of various agents on the transport of ovum was tested in 39 guinea pigs. The administration of estrogens significantly accelerated this process. Estrogens did not act through the induction of prostaglandin synthesis or by altering plasma progesterone levels. Estrogen-induced ovum acceleration was significantly antagonized by tamoxifen, an antiestrogen that acts by interfering with estrogen receptor synthesis. Cycloheximide also antagonized the effects of estrogen on ovum transport. A modification of ovum transport by estrogens is thought to be due to the entrance of estrogen into the nuclei of the target cells, and through subsequent protein synthesis. It is assumed that this process takes place at the oviduct level, although the experiment did not prove this.

Failure of castration to modify estradiol-induced tube-locking of rabbit ova.

Exogenous estradiol, administered in the appropriate dose at the proper time in relation to ovulation, "locks" ova within the rabbit oviduct. The mechanism by which estrogen acts on the oviduct has not been elucidated. In these experiments, ovaries were removed from rabbits at various times prior to the transfer of donor ova and the administration of estrogen. The significant number of ova remaining in the oviducts 72 hours later demonstrates that ovaries are not essential to estrogen-induced "tube-locking."

An ovum capture inhibitor (OCI) in endometriosis peritoneal fluid: an OCI-related membrane responsible for fimbrial failure of ovum capture.

The present study was conducted to assess the mechanism of in vitro interference with fimbrial ovum capture by the ovum capture inhibitor (OCI) which we have recently demonstrated in endometriosis peritoneal fluid (PF). A golden hamster oviduct exposed to either endometriosis or nonendometriosis PF for 20 minutes at 37 degrees C was examined by scanning electron microscopy. Exposure of the oviduct to endometriosis PF reduced fimbrial activity of ovum capture and developed an OCI-related membrane on the fimbria, by which fimbrial cilia were completely concealed. This was not the case for nonendometriosis PF. Subsequently, an oviduct having been exposed to endometriosis PF was retrogradely flushed, by which the OCI-related membrane was ballooned and removed. The flushed oviduct resumed its activity of ovum capture. The OCI-related membrane appeared a cause of OCI interference with fimbrial ovum capture by preventing the contact between the fimbrial cilia and the cumulus oophorus.

Effects of prostaglandin F2alpha on egg transport and in vivo egg recovery from the vaginas of rabbits.

The effects of prostaglandin F2alpha (PGF2alpha) on egg transport and vaginal egg recovery were examined in rabbits. Little disturbance of egg transport was induced by an injection of 2 or 5 mg/kg of PGF2alpha 12 hours after mating. A remarkable disturbance of egg transport was seen when 5 mg/kg were injected 22 hours after mating. The administration of PGF2alpha24 hours after mating led to maximal vaginal egg recovery: 48.4% and 54.8% of eggs were recovered by 51 hours after treatment, and implantation rates were 3.2% and 6.5% after treatment with 2 and 5 mg/kg, respectively. The earliest eggs were recovered vaginally 3 hours after administration of PGF2alpha at 24, 36, 48, or 96 hours after mating. The recovery of slightly bloody washing fluids at the first and second vaginal washings in animals treated 24 or 36 hours after mating appears to indicate that severe uterine contractions were induced by PGF2alpha.

Effect of 15(S)-15-methyl prostaglandin F2alpha on human oviductal motility and ovum transport.

The effects of an intravenous infusion of 15(S)-15-methyl prostaglandin F2alpha (PGF2alpha) on oviductal motility and ovum transport were studied in women who were scheduled for elective tubal sterilization. Infusion rates of 0.38 microgram/kg/hour or higher caused an increase in oviductal motility in all patients. Lower infusion rates did not always cause a stimulation of motility. Low infusion rates generally caused an increase in the amplitude of contractions without any effect on basal oviductal tone. The higher infusion rates usually caused a large increase in basal tone as well as an increase in the amplitude of contractions. Ova were recovered from the oviducts of five patients who had received an intravenous infusion of 15(S)-15methyl PGF2alpha. The ova were recovered from the ampulla in three patients, from the ampullary-isthmic junction in one patient, and from the isthmus in one patient. Since one would expect to recover ova from the oviducts at similar times under normal circumstances, there was no evidence that this prostaglandin treatment caused an acceleration of ovum transport. These data support the conclusion that a PGF analog which stimulates oviductal motility does not necessarily also accelerate ovum transport in women.

Administration of prostaglandin F2alpha for the recovery of fertilized eggs from the vaginas of rabbits.

Nonsurgical recovery of fertilized eggs from the vaginas of prostaglandin F2alpha (PGF2alpha)-treated rabbits was attempted. Eighty-two eggs were collected by the vaginal washing method, and the earliest egg was recovered 20 hours after mating. Twenty eggs recovered vaginally 24 to 107 hours after mating were transferred to six synchronized, pseudopregnant rabbits to test the viability of the eggs; four newborns, two fetuses, and one implantation site were obtained. These results indicate that some viable eggs are discharged into the vaginas of rabbits treated with PGF2alpha.

Hormonal control of oviductal ciliary activity: effect of prostaglandins.

Prostaglandins have been shown to exist in the walls of the oviduct and to effect strongly oviductal muscle contractions and egg transport. Our observations, using laser light-scattering spectroscopy, indicate that the "natural" prostaglandins F2 alpha, E1, and E2 can stimulate ciliary activity in cultures of ciliated epithelium of the rabbit oviduct. These findings suggest a new alternative to explain the effect of prostaglandins on oviductal egg transport.