Effect Of Gestagens Research Articles

High-performance thin-layer chromatography in combination with a yeast-based multi-effect bioassay to determine endocrine effects in environmental samples.

Effect-directed analysis (EDA) that combines effect-based methods (EBMs) with high-performance thin-layer chromatography (HPTLC) is a useful technique for spatial, temporal, and process-related effect evaluation and may provide a link between effect testing and responsible substance identification. In this study, a yeast multi endocrine-effect screen (YMEES) for the detection of endocrine effects is combined with HPTLC. Simultaneous detection of estrogenic, androgenic, and gestagenic effects on the HPTLC plate is achieved by mixing different genetically modified Arxula adeninivorans yeast strains, which contain either the human estrogen, androgen, or progesterone receptor. Depending on the yeast strain, different fluorescent proteins are formed when an appropriate substance binds to the specific hormone receptor. This allows to measure hormonal effects at different wavelengths. Two yeast cell application approaches, immersion and spraying, are compared. The sensitivity and reproducibility of the method are shown by dose-response investigations for reference compounds. The spraying approach indicated similar sensitivities and higher precisions for the tested hormones compared to immersion. The EC10s for estrone (E1), 17β-estradiol (E2), 17α-ethinylestradiol (EE2), 5α-dihydrotestosterone (DHT), and progesterone (P4) were 95, 1.4, 10, 7.4, and 15pg/spot, respectively. Recovery rates of E1, E2, EE2, DHT, and P4 between 88 and 120% show the usability of the general method in combination with sample enrichment by solid phase extraction (SPE). The simultaneous detection of estrogenic, androgenic, and gestagenic effects in wastewater and surface water samples demonstrates the successful application of the YMEES in such matrices. This promising method allows us to identify more than one endocrine effect on the same HPTLC plate, which saves time and material. The method could be used for comparison, evaluation, and monitoring of different river sites and wastewater treatment steps and should be tested in further studies.

Regulatory aspects of myogenic factors GDF-8 and Follistatin on the intake of combined oral contraceptives

Combined oral contraceptives are one of the most prescribed drugs in the western world. While there is little evidence regarding effects of estrogen or gestagens on muscle metabolism, androgens are well-known for their anabolic characteristics. In this study, we seeked to investigate potential correlations of the myokines GDF-8, IGF-1 and Follistatin with female sexual hormones and likewise possible interactions with combined oral contraceptives (Dienogest and Ethyl Estradiol) intake. We obtained serum samples of young healthy women to measure hormone correlations. Furthermore, we simulated combined oral contraceptive blood circulating hormone concentrations to identify myogenic effects on HSkM in vitro. GDF-8, IGF-1 and Follistatin showed concentration correlations (p = .005) in overall patients’ serum, while Follistatin as a promyogenic protein additionally showed a positive correlation with testosterone and estradiol (p < .05). Lower GDF-8 levels were also linked to a higher BMI (p = .009). Upon combined oral contraceptives (COC) intake, patients showed decreased GDF-8 (p = .006) but increased Follistatin (p = .0001) concentrations compared to patients without COC intake. In vitro, addition of Ethyl Estradiol and Dienogest to HSkM cells revealed a pro-myogenic, proliferative, chemosensitized pattern. Our data support a pro-myogenic effect of combined oral contraceptives.

EFFECT OF THE NEW STEROID DRUG GESTOBUTANOIL FOR HORMONE REPLACEMENT THERAPY OF GESTAGENOUS INSUFFICIENCY ON THE DEVELOPMENT OF FETUS AND OFFSPRING IN EXPERIMENTS ON RATS

A study of the embryo- and fetotoxic effects registered in the antenatal and postnatal periods of development after the intragastric administration of Gestobutanoil in two doses (equitherapeutic and 10 times higher) has been performed. It has been found that the introduction of gestagen Gestobutanoil in both doses did not affect the body weight of pregnant animals, which continued to increase progressively during pregnancy. The embryotropic effect of gestagen was revealed to be dose-dependent. The introduction of gestagen in a dose of 0.25 mg / kg (calculated equitherapeutic dose) had no adverse effect on the development of the embryo and fetus of rats. The gestagen in a dose of 2.5 mg / kg (10 equitherapeutic dose) had a negative effect on the course of pregnancy, which has been manifested by a statistically significant increase in preimplantation death and total fetal mortality, as well as a statistically significant decrease in the number of fetuses. It has been shown that intragastric administration of Gestobutanoil in doses of 0.25 and 2.5 mg / kg to female rats from the 6th to the 19th days of pregnancy did not adversely affect the early development of the first generation offspring (postnatal mortality, body mass dynamics, and development of rats’ sensory-motor reflexes).

Elevated yolk progesterone moderates prenatal heart rate and postnatal auditory learning in bobwhite quail (Colinus virginianus).

Previous studies have established that yolk hormones of maternal origin can influence physiology and behavior in birds. However, few studies have examined the effects of maternal gestagens, like progesterone, on chick behavior and physiology. We tested the effects of experimentally elevated egg yolk progesterone on embryonic heart rate and postnatal auditory learning in bobwhite quail hatchlings. Quail chicks were passively exposed to an individual maternal assembly call for 10 min/hr during the 24 hr following hatching. Preference for the familiarized call was tested at 48 hr following hatching in three experimental groups: chicks that received artificially elevated yolk progesterone (P) prior to incubation, vehicle-only controls (V), and non-manipulated controls (C). Resting heart rate of P, V, and C embryos were also measured on prenatal day 17. The resting heart rate of P embryos was significantly higher than both the V and C embryos. Chicks from the P group also showed an enhanced preference for the familiarized bobwhite maternal call when compared to chicks from the C and V groups. Our results indicate that elevated yolk progesterone in pre-incubated bobwhite quail eggs can influence arousal level in bobwhite embryos and postnatal perceptual learning in bobwhite neonates.

Synthetic gestagens exert differential effects on arterial thrombosis and aortic gene expression in ovariectomized apolipoprotein E-deficient mice.

Combined hormone replacement therapy with oestrogens plus the synthetic progestin medroxyprogesterone acetate (MPA) is associated with an increased risk of thrombosis. However, the mechanisms of this pro-thrombotic effect are largely unknown. The purpose of this study was to: (i) compare the pro-thrombotic effect of MPA with another synthetic progestin, norethisterone acetate (NET-A), (ii) determine if MPA's pro-thrombotic effect can be antagonized by the progesterone and glucocorticoid receptor antagonist mifepristone and (iii) elucidate underlying mechanisms by comparing aortic gene expression after chronic MPA with that after NET-A treatment. Female apolipoprotein E-deficient mice were ovariectomized and treated with placebo, MPA, a combination of MPA + mifepristone or NET-A for 90 days on a Western-type diet. Arterial thrombosis was measured in vivo in a photothrombosis model. Aortic gene expression was analysed using microarrays; GeneOntology and KEGG pathway analyses were conducted. MPA's pro-thrombotic effects were prevented by mifepristone, while NET-A did not affect arterial thrombosis. Aortic gene expression analysis showed, for the first time, that gestagens induce similar effects on a set of genes potentially promoting thrombosis. However, in NET-A-treated mice other genes with potentially anti-thrombotic effects were also affected, which might counterbalance the effects of the pro-thrombotic genes. The pro-thrombotic effects of synthetic progestins appear to be compound-specific, rather than representing a class effect of gestagens. Furthermore, the different thrombotic responses elicited by MPA and NET-A might be attributed to a more balanced, 'homeostatic' gene expression induced in NET-A- as compared with MPA-treated mice.

Does Adjuvant Long-Acting Gestagen Therapy Improve the Outcome of Hysteroscopic Endometrial Resection in Women of Low-Resource Settings with Heavy Menstrual Bleeding?

To test for the hypothesis of the beneficial effect of long-acting gestagens as an adjuvant postoperative therapy on the outcome of hysteroscopic transcervical endometrial resection (TCRE) in women of low-resource settings and suffering from ovulatory heavy menstrual bleeding (menorrhagia).

Does Adjuvant Long-Acting Gestagen Therapy Improve the Outcome of Hysteroscopic Endometrial Resection in Women of Low-Resource Settings With Heavy Menstrual Bleeding?

ObjectiveTo test for the hypothesis of the beneficial effect of long-acting gestagens as an adjuvant postoperative therapy on the outcome of hysteroscopic transcervical endometrial resection (TCRE) in women of low-resource settings and suffering from ovulatory heavy menstrual bleeding (menorrhagia). DesignRandomized controlled trial (Canadian Task Force classification I). SettingLow-resource tertiary care university hospital. PatientsSeventy-one premenopausal women with established ovulatory menorrhagia. InterventionAfter randomization, 37 patients were treated with TCRE plus gestagen and 34 patients with TCRE alone. Measurements and Main ResultsVariations in menstrual patterns and bleeding scores, as well as amenorrhea and repeat surgery rates with treatment, were determined 1 year after resection. In those who continued to menstruate at 6 months, treatment with TCRE plus gestagen was associated with a significant reduction in the number of days bleeding from 7.2 to 3.4 (p ≤ .0001), increased cycle length from 24 to 30 (p = .02), a 60% reduction in dysmenorrhea score from 62 to 25 (p ≤ .0001), and a 60% reduction in premenstrual syndrome score from 55 to 22 (p = .04). Amenorrhea rates at 12 months in the TCRE plus gestagen and TCRE alone groups were 40% versus 26% (p = .02), with combined amenorrhea and hypomenorrhea rates of 75% versus 64% (p = .02), respectively. At 12 months, repeat surgery rates were higher in the TCRE alone group (21% vs 3%, p < .05). ConclusionIn a low-resource setting, the adjuvant postoperative long-acting gestagen therapy has proven to be superior in inducing amenorrhea after hysteroscopic TCRE.

Progestagens for human use, exposure and hazard assessment for the aquatic environment

Little information is available on the environmental occurrence and ecotoxicological effects of pharmaceutical gestagens released in the aquatic environment. Since eighteen different gestagens were found to be used in France, preliminary exposure and hazard assessment were done. Predicted environmental concentrations (PECs) suggest that if parent gestagens are expected to be found in the ng l −1 range, some active metabolites could be present at higher concentrations, although limited data on metabolism and environmental fate limit the relevance of PECs. The biological effects are not expected to be restricted to progestagenic activity. Both anti-androgenic activity (mainly for cyproterone acetate, chlormadinone acetate and their metabolites) and estrogenic activity (mainly for reduced metabolites of levonorgestrel and norethisterone) should also occur. All these molecules are likely to have a cumulative effect among themselves or with other xenoestrogens. Studies on occurrence, toxicity and degradation time are therefore needed for several of these compounds.

Serum lipoprotein changes in female rats treated with progesterone or synthetic gestagens alone or in combination with estradiol. I. Total and fractionated cholesterol and lipoprotein pattern.

In adult female rats, the effects of progesterone (P), norethisterone acetate (NEA), levonorgestrel (LNG), dienogest (DEG) and chlormadinone acetate (CMA) given alone at doses of 2.5 or 10 mg/kg p.o. or in combination with s.c. implanted estradiol (E2) on total cholesterol (TC), high-density-lipoprotein cholesterol (HDL-C), low-density-lipoprotein cholesterol (LDL-C) and very-low-density-lipoprotein cholesterol (VLDL-C) were investigated. Additionally, the lipoprotein pattern was determined using agarose gel electrophoresis. Synthetic gestagens derived from nortestosterone (NEA, LNG, DEG) lowered TC by reduction of HDL-C and LDL-C, whereas E2 induced an increase of these lipids. The decrease of HDL-C and LDL-C caused by the gestagens was also found in E2 pretreated rats. In contrast, the pregnane related CMA and P given alone did not diminish TC, HDL-C or LDL-C. But they partly reversed the enhancing effect of E2 on the HDL-C fraction following their simultaneous administration. The results suggest that the cholesterol lowering effects of gestagens are mediated rather via androgen than via progesterone receptors.

Effects of Progesterone and Its Antagonist Mifepristone on Progesterone Receptor A Expression in Human Umbilical Vein Endothelial Cells

Effects of female steroid hormones on endothelial cells are gaining increased importance due to several studies on the effects of hormonal treatment on cardiovascular risk. Recent data argue for an improvement of endothelium-derived relaxation and impaired vascular contraction by estradiol, whereas progesterone and testosterone might entail contrary effects. So far, gestagenic influence on endothelial cell physiology is poorly understood. Human umbilical vein endothelial cells (HUVECs) exposed to the female sex hormones estradiol and progesterone show expression of estrogen receptor-β (ERβ) and progesterone receptor A (PR-A), and are negative for ERα and PR-B. The aim of this study was to analyze the expression and stimulation of PR-A and -B in HUVECs after stimulation with progesterone and PR antagonists that are commercially available. PR-B expression or upregulation was abrogated after application of progesterone or antagonists to HUVECs. Expression of PR-A could be significantly upregulated with progesterone and mifepristone. Unexpectedly, stimulation with the progesterone antagonist RU486 (mifepristone) was accomplished by an upregulation of PR-A expression in our study. We conclude that gestagenic effects on HUVECs independent of modulators are mediated via the PR-A.

Comparative analysis of the effect of gestagens, antiestrogencytostatics, and androstenes on the viability of tumor and normal cells

The hormonal compound with the highest cytostatic activity against MCF-7 tumor cells (human breast cancer, BC) and the lowest activity against normal cells (rat skin fibroblasts) was sought among gestagens, androstenes, and antiestrogencytostatics. It was found that antiestrogencytostatics and androstenes had the highest cytostatic activity against tumor cells whereas gestagens and antiestrogencytostatics were least active against fibroblasts. Studies of the activity of the hormonal compounds in combination with doxorubicin on the viability of MCF-7 and rat skin fibroblasts found that all investigated compounds with the exception of dehydroepiandrosterone (DHEA) intensify the cytostatic activity of doxorubicin against tumor cells, the greatest effect seen for antiestrogencytostatics. A chemoprotective effect of androstenes on normal cells was noted.

Tissue expression of the nuclear progesterone receptor in male non-human primates and men

In females, progesterone is associated with reproductive functions. In males, its role and the expression of its genomic receptor are not very well understood. In attempts to achieve a hormonal male contraceptive method, gestagens are used to downregulate gonadotropin and sperm production. It is therefore essential to understand the mechanism of action of progesterone at the molecular level in males, especially in primates. This investigation was undertaken: (a) to determine whether the genomic progesterone receptor is expressed in males; and (b) to locate it in various organs that are potential targets of gestagens. Human tissues were obtained at surgery for benign prostatic hyperplasia or prostate cancer and at autopsy. Non-human primate tissues were obtained at autopsy. This study was performed by analyzing the genomic progesterone receptor by immunohistochemistry, Western blot and RT-PCR. The nuclear progesterone receptor was expressed in pituitary and hypothalamus of both monkeys and men. In the testis progesterone receptor expression was found in a few peritubular and interstitial cells, but not in germ cells. In addition, expression was detected in the epididymis, prostate and male mammary gland. Reverse transcriptase (RT)-PCR experiments indicated that progesterone receptor A and B are expressed in all tissues analyzed. These data exclude direct genomic effects of gestagens at the spermatogenic level but indicate that a male contraceptive based on gestagens might have some effects on other tissues, such as the epididymis, prostate and mammary gland.

Regulation of expression of the retinoic acid-synthesising enzymes retinaldehyde dehydrogenases in the uteri of ovariectomised mice after treatment with oestrogen, gestagen and their combination

The active metabolite of vitamin A, retinoic acid (RA), plays an important role in the female reproductive system. The synthesis of RA is tightly regulated by the activity of retinaldehyde dehydrogenases (Raldh). Among these, Raldh1 and Raldh2 exhibit specific temporal and spatial expression patterns in the mouse uterus, both during the oestrous cycle and early pregnancy. In the present study, we have assessed whether oestradiol and progesterone directly influence the uterine expression of Raldh1 and Raldh2 in ovariectomised mice. We investigated the effect of gestagen (promegestone 0.3 mg kg(-1) bodyweight), oestrogen (oestradiol 3 microg kg(-1) bodyweight) and their combination on the uterine expression of Raldh2. Expression was analysed using in situ hybridisation and quantified using real-time detection reverse transcription-polymerase chain reaction. The results show that the expression of Raldh2 is rapidly (within 1-4 h) induced in stromal cells by oestrogen, but not by gestagen, treatment, whereas combined oestrogen + gestagen treatment leads to a more prolonged (48 h) response. In contrast, oestrogen, but not progesterone, treatment downregulates (within 4-24 h) Raldh1 expression in the uterine glandular epithelium. We conclude that the uterine RA concentrations are regulated by oestrogens via an effect on the expression of the Raldh synthesising enzymes. Such a regulation is consistent with the natural fluctuations of Raldh expression during the oestrous cycle, early pregnancy and blastocyst implantation.

Evaluation of the effects of various gestagens on insulin sensitivity, using homeostatic model assessment, in postmenopausal women on hormone replacement therapy

The objective of the present study was to compare the effects of various gestagens on insulin sensitivity in postmenopausal women on hormone replacement therapy (HRT). This prospective study enrolled 156 postmenopausal women who had menopausal status for at least 6 months. Group 1 was treated with 17β-estradiol (E2; 2 mg) plus norethisterone acetate (NETA; 1 mg); Group 2 was given E2 (2 mg) plus medroxyprogesterone acetate (MPA; 2.5 mg); Group 3 was given E2 (2 mg) plus dydrogesterone (DG; 10 mg); and Group 4 was given E2 (2 mg) plus micronized progesterone (MP; 100 mg). Group 5 was the surgical menopausal group and was given only E2 (2 mg) continuously. All 156 subjects completed the 3-month follow-up on the trial. The patients were analyzed by using homeostatic model assessment (HOMA) for insulin sensitivity before treatment and 3 months after treatment, comparing the effects of various HRT regimens on insulin sensitivity. No significant differences were found in the baseline characteristics of the patients (p > 0.05). There were no significant differences in mean values of HOMA before HRT among the five groups (p > 0.05). There were statistically significant differences in mean values of HOMA only in Group 1 (E2 + NETA) and Group 3 (E2 + DG) after HRT (p > 0.05). E2 + NETA and E2 + DG were found to improve insulin sensitivity in postmenopausal women after 3 months of treatment, whereas E2 + MPA, E2 + MP and E2 only did not show such an effect in postmenopausal women.

The effect of gestagens on bone turnover in sheep

SummaryThe effects of two gestagens on bone turnover were investigated in three groups of female sheep comparable in age and weight. Group A (n = 10) was given a single intramuscular injection of 50 mg chlormadinon acetate (CMA), group B (n = 9) received 140 mg medroxyprogesterone acetate (MPA), and control group C (n = 9) received 5 ml physiological NaCI-solution. Plasma progesterone was measured weekly over 3 months. Bone mineral density (BMD) in the right distal tibia and calcaneus were determined every four weeks during the four months of the study duration, using peripheral quantitative computed tomography (pQCT). At the same times total alkaline phosphatase (AP) was determined. No significant changes of BMD were found in the three groups during the four months. Total alkaline phosphatase showed a significant decrease of 36-45% in all groups during the first six weeks. Sheep seem to have a comparable bone metabolism to humans with respect to the short term effect of gestagens, i. e. MPA and CMA.

Effect of glucocorticoids and gestagens on glutathione redox-system in rat skin involved in dermatitis

In the model of experimental dermatitis the effect of hydrocortisone and two gestagens (endogenous progesterone and synthetic mecygestone) on the level of glutathione redox-system components (reduced glutathione, glutathione reductase, and glutathione peroxidase) in rat dermis and epidermis is characterized by different degree of activation of this system.

Current drug therapy recommendations for the treatment of endometriosis.

The principal symptoms and signs of endometriosis are tissue lesions and pelvic pain. These occur to varying degrees, with a chronic pattern and a tendency for deterioration with time. Patients with endometriosis often also have fertility problems, but the relationship between this and the signs and symptoms of the disease is inconsequent; the basic pathophysiology is not exactly known. Although an immunological defect resulting in an inflammatory reaction around discharged menstrual debris in the pelvic cavity has been shown, no treatments based on this process are available. Estrogen often plays an important role in the progression of lesions and pain. Therefore, the aim of treatment usually has been to downregulate the ovaries and/or given antiestrogenic drugs as an alternative to surgical removal. As complete downregulation of the ovaries and hypoestrogenaemia does not seem to be crucial, achievement of amenorrhoea seems to be sufficient. This means that women may continue to have circulating estrogen levels so that severe hypoestrogenic adverse effects such as bone demineralisation, dry vagina, psychiatric symptoms or anabolic/androgenic effects of gestagens can be avoided. However, as both symptoms and the dependence of hormones may vary between and within women, the treatment needs to be individualised. There are a number of available treatments for endometriosis on the market and it is important for the doctor to know how to reach the therapeutic window of these treatments for each woman. It is also important to inform the patient about the different possibilities so that the treatment with the least impact on her quality of life can be chosen. When the therapeutic window has been identified, the treatment may then either be continued for a long period of time or be repeated when needed.

Calcium and depolarization-dependent effect of pregnenolone derivatives on uterine smooth muscle

1. 1. The effects of several gestagens (pregnenolone [1 to 30 μm], 20α-hydroxypregnenolone [1 to 30 μM]), and 20β-hydroxypregnenolone [1 to 30 μm]) on rat uterine contraction induced by KC1 (60 mM) and CaCl 2 (30 μM to 6 mM) have been assayed. 2. 2. The three drugs relaxed the tonic contraction induced by KCI in a concentration-dependent way. The respective EC 50 values were: 27.6±1.58 μM (pregnenolone), 4.1±0.12 μM (20α-hydroxypregnenolone), and 11.2±1.04 μM (20β-hydroxypregnenolone). CaCl 2 (1 to 10 mM) totally counteracted the relaxing effect of pregnenolone but only partially compared to that of 20α- or 20β-hydroxy-pregnenolone. 3. 3. CaC1 2 (30 μM to 6 mM) produced concentration-dependent contraction of rat uterus in medium lacking calcium plus 30, 60, or 90 mM of KC1. The EC 50 values of CaCl 2 were: 0.38±0.072, 0.183±0.015, and 0.183±0.015 μM in a medium with 30, 60, or 90 mM of KCI, respectively. 4. 4. Pregnenolone (10 μM) did not significantly modify the EC 50 of CaC1 2 in a medium with 30, 60, or 90 mM of KCI. However, 20β-hydroxypregnenolone (10 μM) antagonized, in a noncompetitive manner, the concentration-response curve to CaC1 2. 5. 5. 20α-Hydroxypregnenolone (4 μM) antagonized the concentration-response curve to CaCl 2 in a competitive manner. This antagonism was directly related to the concentration of KCI in the medium. 6. 6. Our results suggest a different calcium antagonist effect of the three gestagens assayed.

Gestagens and heart

The use of gestagen hormones (progestogens, progestogens, progestins) in modern medicine is mainly based on their effect on the reproductive sphere of the female body, including menstrual dysfunction (luteal phase of the cycle), contraception, pregnancy support, correction of hormonal homeostasis during menopause, antitumor activity in some hormone-dependent oncological diseases. The presence of antiandrogenic properties in a number of progestogens (C21-derivatives), which have found application as antihormonal therapy for certain endocrine diseases in women (hirsutism) and for malignant neoplasms of the male genital organs, should be highlighted. The variety of influences exerted by natural gestagen hormones and their synthetic analogs on the human body also encompasses a change in their action of cardiovascular activity. Moreover, according to some authors, the heart and large vessels are the targets of the action of gestagens. It is assumed that a change in the hormonal characteristics of the female body during the normal menstrual cycle does not cause existing changes in the reactivity of the cardiovascular system. However, fluctuations in cardioactive and vasoactive parameters can be observed even with physiological differences in the level of endogenous progestogens and estrogens (menstrual cycle, pregnancy, menopause). The greatest severity of maladaptation phenomena in cardiac activity (according to the mathematical analysis of cardiointervalograms) was revealed precisely at peak concentrations of female sex hormones within the estrous cycle - in the phases of proestrus and metaestrus. During menopause and postmenopause, when the cardioprotective effect of endogenous estrogens and gestagens is significantly reduced, a violation of cardiac and vasomotor activity can be detected, often requiring hormone replacement therapy with estrogen-gestagen drugs.

Progesterone and pregnanolone derivatives relaxing effect on smooth muscle

1. 1. The effect of gestagens, 5α-hydroxyprogesterone (10 −6−6 × 10 −5 M), 5β-hydroxyprogesterone (10 −6−3 × 10 −5 M), progesterone (6 × 10 −6−10 −4 M), pregnanolone (10 −6−10 −5 M), allopregnanolone (10 −6−10 −4 M) and epipregnanolone (10 −6−6 × 10 −5 M) on rat uterine contractions induced by KCl (60 mM), has been assayed. 2. 2. All drugs assayed relaxed the tonic-contraction induced by KCl in a concentration-dependent way. The respectives IC 50 were 31.3 ± 4.1 × 10 −6 M (progesterone), 8.9 ± 0.8 × 10 −6 M (5α-hydroxyprogesterone 3.8 ± 0.3 × 10 −6 M (5β-hydroxyprogesterone), 3.1 ± 0.1 × 10 −6 M (pregnanolone), 21.2 ± 3.1 × 10 −6 M (allopregnanolone) and 6.3 ± 1.3 × 10 −6 M (epipregnanolone). This relaxing effect was partially or totally counteracted by CaCl 2 (1–10 mM) 3. 3. Cycloheximide (10 μg/ml) significantly shifted to the right the effect of allopregnanolone but not the effect of the other drugs. Actinomycin D (5 μg/ml) did not modify the effect of allopregnanolone. 4. 4. Our results suggest that the relaxing effect of gestagens in the rat uterus could be related to inhibition on calcium influx and mainly occur through non-genomic mechanisms.