A possible role for progesterone in the preovulatory gonadotropin surge through modulation of LHRH degrading activity

The preovulatory surge of gonadotropins is triggered by estradiol and enhanced to its full magnitude by progesterone. Progesterone may exert this effect through several mechanisms. One of the mechanisms is through the ability of progesterone to induce an increase in the hypothalamic content and release of LHRH. The purpose of this study was to determine if progesterone might not act through yet another mechanism and facilitate LHRH release of the proestrous gonadotropin surge through modulation of luteinizing hormone releasing hormone (LHRH) degrading activity. Sixty-day-old Sprague-Dawley rats were ovariectomized; 14 days later, the estradiol-progesterone milieu of proestrous was mimicked in these animals through the use of estradiol containing silastic implants and subcutaneous progesterone injections. The LHRH degrading activity of the hypothalamus, pituitary and serum were monitored subsequently at preselected time points. In the hypothalamus, estradiol alone was capable of inducing significant increase in degrading activity; progesterone alone had no effect; however, progesterone subsequent to estradiol priming suppressed the increase induced by estradiol alone. In the pituitary, neither estradiol alone nor progesterone alone nor progesterone subsequent to estradiol priming had any significant effect on degrading activity. In the serum, estradiol induced a rapid and significant increase in activity; progesterone alone suppressed activity; progesterone subsequent to estradiol priming induced a similar but more rapid suppression. Therefore, the overall tendency was for estradiol to stimulate and progesterone to suppress LHRH degrading activity in the tissues studied. The results of this study indicate that progesterone has the capacity to suppress LHRH degrading activity and may be one of the mechanisms capable of increasing the availability of LHRH to the anterior pituitary gland thereby facilitating the preovulatory gonadotropin surges.     

The effect of smoking on serum progesterone, estradiol, and luteinizing hormone levels over a menstrual cycle in normal women

Since smoking has been shown to affect serum progesterone and estradiol levels in postmenopausal women, we evaluated the levels of these hormones and luteinizing hormone (LH) over an entire menstrual cycle (17 points) in eight healthy nonsmokers and eight healthy smokers. The total length of the cycle and the lengths of the follicular and luteal phases did not differ between the groups. There was no difference in estradiol, progesterone, or LH levels during the periovulatory and luteal phases. Follicular-phase serum progesterone, which had a level 37% higher in smokers, showed a plateau in both groups (28.3 +/- 5.7 ng/dl versus 20.7 +/- 5.7; P less than 0.0001). Follicular-phase serum estradiol showed a rising curve in both groups. The mean value in smokers was slightly higher than that in nonsmokers (107 pg/ml versus 95; P approximately 0.05); during the early part of the follicular phase, prior to the rapid preovulatory increase, the difference was greater (23%) and of higher statistical significance (80 pg/ml versus 65; P less than 0.001). The follicular-phase LH levels of smokers were skewed downward from the levels in nonsmokers, presumably by negative feedback from the elevated estradiol and progesterone levels; the difference was significant (P less than 0.001). The elevations of serum progesterone and estradiol in smokers probably represent activation of adrenocortical secretion by smoking. The greater and more clear-cut rise of progesterone than of estradiol is probably due to the fact that essentially all of the follicular-phase serum progesterone is secreted by the adrenal, while only part of the follicular-phase serum estradiol comes from the adrenal (via androstenedione and estrone).     

In the absence of protein, estradiol suppresses meiosis of porcine oocytes in vitro

The effect of estradiol on the spontaneous maturation of porcine oocytes was investigated. Cumulus-enclosed (intact) and cumulus-free (denuded) oocytes were cultured in the presence of estradiol-17 beta (0 to 10 microgram/ml) in a chemically defined bicarbonate-buffered medium that contained either dextran or BSA, or in a complex Hepes-buffered medium that was supplemented with serum. After 24 hr, chromatin spreads were prepared and meiotic maturation was scored. The biochemical integrities of the cumulus cells were assessed by determination of the estradiol and progesterone content of spent media after culture of intact oocytes in the presence of 0.5 X 10(-6) M testosterone and 10 microgram/ml follicle-stimulating hormone. Estradiol did not significantly affect the onset of maturation of either intact or denuded oocytes that were cultured in medium containing either BSA or serum. In serum-supplemented medium, however, the progression of maturation beyond metaphase I was significantly affected by the steroid in a dose-dependent manner. The steroid significantly inhibited the release from meiotic arrest of both types of oocyte cultured in medium supplemented with dextran. Supplementation of all media with testosterone and FSH significantly stimulated the synthesis of estradiol by the cumulus cells, compared with that of control groups. The synthesis of progesterone, however, was significantly stimulated by testosterone and FSH only in the BSA and serum-supplemented media. It is concluded that exogenous estradiol has the capacity to arrest meiosis in vitro but that this capacity can only be expressed if no exogenous protein(s) is present. In the absence of exogenous protein, progesterone synthesis by the adherent cumulus cells is minimal.     

Antagonism of estrogen-induced prolactin release by progesterone

Previous work from our laboratory has shown that during the process of nuclear occupancy of the progesterone receptor complex (1-2 h), nuclear estradiol receptors of the anterior pituitary are depleted. The purpose of this study was to determine whether the depletion of nuclear estradiol receptors by progesterone had functional biological significance. The ovariectomized (26 days of age) immature rat was used as the model for analysis of this question. The ability of estradiol to release prolactin from the anterior pituitary was the function chosen to determine the biological significance of the progesterone and estradiol interactions. In response to estradiol exposure (2 micrograms/rat), prolactin release reached peak values from 8 h to 12 h and returned to control levels by 24 h. A second injection of estradiol 13 h after the initial injection stimulated a second increase in serum prolactin at 25 h. This model of two injections of estradiol 13 h apart served to provide adequate levels of anterior pituitary progesterone receptors and elevated serum prolactin levels upon which superimposed progestin modulation could be examined. A single injection of progesterone (0.8 mg/kg BW) 1 h before the second estradiol injection blocked the increase in serum prolactin. This action was a receptor-mediated event because progesterone had no effect without estrogen priming or when the progesterone antagonist RU486 was used. Finally, when the interval between the progesterone and second estradiol injection was extended to 4 h, a time period when progesterone does not deplete pituitary nuclear estrogen receptors, the estrogen-induced increase in serum prolactin was not blocked.(ABSTRACT TRUNCATED AT 250 WORDS)     

Effect of steroid hormones on vaginal epithelial cells: Anin vitro model for steroid hormone action

Conditions have been standardized to maintain rat vaginal epithelial cellsin vitro with more than 95% viability. Cultured epithelial cells were used to study the effects of normal fetal calf scrum, estradiol and progesterone on the incorporation of [³H]-uridine in RNA and incorporation of [¹⁴C]-aminoacids in proteins. While fetal calf serum and estradiol stimulate the incorporation of both uridine and afno acids, progesterone did not show any effect. Estradiol treated vaginal cells show typical fcroridges (indicative of keratinization of cells) in contrast to estradiol deprived cells, which show microvilli on cell surface when examined in scanning electron microscope.     

Prolactin, LH, and estradiol-17 beta in utilization of lipoprotein substrate by porcine granulosa cells in vitro

Porcine granulosa cells cultured under serum free conditions responded by increased progesterone secretion to the addition of the leuteotropic hormones, LH, prolactin, and estradiol. Provision of extracellular substrate for steroidogenesis in the form of porcine high density lipoprotein or low density lipoprotein enhanced progesterone accumulation by granulosa cell cultures. Estradiol, LH, and prolactin all greatly increased progesterone accumulation in the presence of either high or low density lipoproteins. Increases in progesterone accumulation following addition of prolactin or LH in combination with estradiol suggested the presence of a synergistic interaction among leuteotropins. Pre-exposure of granulosa cell cultures to estradiol increased the subsequent stimulatory effect of prolactin on lipoprotein utilization. It is concluded that all three leuteotropins function to enhance and may interact in the utilization of extracellular lipoprotein substrate for progesterone synthesis.     

Serum levels of prolactin during the ovarian cycle,pregnancy, and lactation in the common marmoset (Callithrix jacchus)

A homologous double-antibody radioimmunoassay developed for humans was used to measure serum prolactin, progesterone, and estradiol in common marmosets. In the ovarian cycle of common marmosets, serum progesterone began to increase after an estradiol surge, attained a peak level, and then declined before the ensuing pre-ovulatory rise in estradiol. During the luteal phase, the change in serum concentrations of estradiol was synchronized with that of progesterone. During the ovarian cycle there was no consistent change in serum prolactin concentrations. During the last 75 days of pregnancy the prolactin level was higher than during the ovarian cycle and the first 70 days of pregnancy. Moreover, during lactation, mothers with suckling twin infants had a higher prolactin level than during the final stage of pregnancy.     

Seasonal variations in the responses of luteal and follicular luteinizing hormone-stimulated adenylyl cyclases to estradiol implants and implant removal in pseudopregnant rabbits

Summer rabbits appeared to be less sensitive to the presence of exogenous estradiol or to the withdrawal of exogenous estradiol than were Winter rabbits. Suppression of the luteinizing hormone(LH)-stimulated adenylyl cyclase of corpora lutea by estradiol-filled silastic capsules, which resulted in serum estradiol concentrations approximately 3.8 X control (high level), appeared to be greater in Winter rabbits than in Summer rabbits. Both high level and low level (2.1 X control) estradiol implants suppressed follicular LH-stimulated adenylyl cyclase in Winter rabbits but neither size capsule had an effect in Summer rabbits. Serum LH concentrations were equally low in Winter rabbits using either high or low level estradiol implants, while only the high level implants caused a decrease in serum LH in Summer rabbits. Withdrawal of the exogenous estradiol caused a precipitous fall in serum progesterone concentrations in Winter rabbits; estradiol withdrawal in Summer rabbits was without effect.     

Effect of indomethacin, cycloheximide, aminoglutethimide on ovarian steroid and prostanoid levels during ovulation in the gonadotropin-primed immature rat

It has become popular to use the gonadotropin-primed immature rat to study ovulation. The ovarian content of progesterone, estradiol, PGE₂, PGF_2α, and 6-keto-PGF_1α during the ovulatory process was determined in this model. Also, the effect of three anti-ovulatory agents on the ovarian levels of the above substances was determined. At 23 days of age, Wistar rats were primed with pregnant mares serum gonadotropin (PMSG) sc, and two days later the ovulatory process was initiated with human chorionic gonadotropin (hCG) sc. The ovarian follicles began rupturing 12 h later. Ovaries were assayed for the two steroids and prostanoids at 2-h intervals befored and several 4-h intervals after ovulation. The ovarian estradiol level increased slightly between 0 and 2 h after hCG, while the progesterone level increased sharply between 2 and 4 h after hCg--at a time when the estradiol declined markedly. All three prostanoids increased concomitantly with progesterone. When the PG synthesis was blocked by indomethacin treatment at 1 h before hCG, ovarian progesterone levels still incrased. In contrast, when steroidogenic activity was inhibited by aminoglutethimide, the ovarian prostanoid levels also decreased. Cycloheximide had little effect on the steroids and prostanoids. It is concluded that ovarian prostanoid synthesis might be influenced by ovarian steroid output.     

Endocrine profiles during the estrous cycle and pregnancy in the Baird's tapir (Tapirus bairdii)

Serum samples were collected 1–3 times weekly from two Baird's tapirs (Tapirus bairdii) for 6 months in 1987–1988, and for more than 3 consecutive years beginning in 1989 to characterize hormone patterns during the estrous cycle and pregnancy. Based on serum progesterone concentrations, mean (±SEM) duration of the estrous cycle (n = 20) was 30.8 ± 2.6 days (range, 25–38 days) with a luteal phase length of 18.1 ± 0.4 days (range, 15–20 days). Mean peak serum progesterone concentrations during the luteal phase were 1.35 ± 0.16 ng/ml, and nadir concentrations were 0.19 ± 0.03 ng/ml during the interluteal period. Distinct surges of estradiol preceded luteal phase progesterone increases in most (14/20) cycles. Gestation length was 392 ± 4 days for three complete pregnancies. Mean serum progesterone concentrations increased throughout gestation and were 1.83 ± 0.13, 2.73 ± 0.13, and 4.30 ± 0.16 ng/ml during early, mid- and late gestation, respectively. Serum estradiol concentrations began to rise during mid-gestation, increasing dramatically during the last week of pregnancy. Patterns of serum estriol and estrone secretion during pregnancy were similar to that observed for estradiol. In contrast to progesterone and estrogens, serum cortisol concentrations were unchanged during pregnancy or parturition. Females resumed cycling 16.2 ± 2.0 days after parturition (n = 4) and, on two occasions, females became pregnant during the first postpartum estrus. These data suggest that the tapir cycles at approximately monthly intervals and that increases in serum progesterone are indicative of luteal activity. The interluteal period is relatively long, comprising approximately 40% of the estrous cycle. During gestation, progesterone concentrations are increased above luteal phase levels, and there is evidence of increased estrogen production during late gestation. The absence of increased cortisol secretion at the end of gestation suggests that this steroid does not play a major role in initiating parturition in this species. © 1994 Wiley-Liss, Inc.     

Serum and fecal steroid analysis of ovulation,pregnancy, and parturition in the black rhinoceros (Diceros bicornis)

Studies were conducted to determine: (1) if fecal hormone metabolite concentrations correlated with serum estrogen and progesterone concentrations, follicular activity and reproductive behavior in the black rhinoceros (Diceros bicornis) and (2) if threshold values of respective fecal metabolite concentrations correlated with pregnancy. Blood and fecal samples were collected, in conjunction with transrectal ultrasound and behavior observations, for an 18-month period from one black rhinoceros female. Subsequently, serial fecal samples were collected from 13 females in 10 zoos. Quantitative analysis of serum progesterone (P₄) and estradiol (E₂) was performed by radioimmunoassay (RIA): analysis of fecal estrogen metabolites (E) and fecal progesterone metabolites (P) were performed by enzyme immunoassay (EIA). Serum P₂ concentrations identified two luteal phase patterns and two nadirs which corresponded with behavioral estrus. Fecal E patterns indicated a sharp peak which corresponded with breeding. concentrations of fecal P illustrated identifiable nadirs and several peaks which corresponded to serum P₄ nadirs and luteal phases. Serum P₄ concentrations were not different between the luteal phase and pregnancy. Fecal P concentrations started to rise above luteal phase concentrations approximately 150 days postbreeding and remained elevated until immediately before parturition. Serum E₂ and fecal E concentrations rose and subsequently declined after parturition. In the fecal samples from seven pregnant females, fecal P concentrations were similarly elevated compared to six nonpregnant females. Results indicated that fecal steroid metabolites accurately reflected serum steroid hormone concentrations and that the measurement of P and E concentrations permitted the characterization of the estrous cycle, the diagnosis of pregnancy, and the onset of parturition. Zoo Biol 16:121–132, 1997. © 1997 Wiley-Liss, Inc.     

小熊猫繁殖周期血清雌二醇和孕酮含量变化

为研究小熊猫繁殖周期血清雌二醇、孕酮含量变化规律,采用化学发光免疫分析法连续16 次测定了2只成体雌性小熊猫血清雌二醇和孕酮含量变化,历经发情间期、发情期和两次妊娠期;连续9次测定了7只小熊猫妊娠期的孕酮含量变化。结果:(1)发情间期,小熊猫血清雌二醇的水平一直维持在低水平(基础水平),进入发情前期,血清雌二醇水平明显升高,在发情期一直维持高水平,配种后迅速降至基础水平; (2)小熊猫血清孕酮含量在发情间期和发情期均维持在较低水平,直至发情期过后才出现升高,在妊娠期一直维持高水平,峰值出现在5 月;(3)发情的小熊猫不论妊娠与否,在妊娠期内血清孕酮含量均维持在高水平。研究表明:小熊猫血清雌二醇、孕酮含量变化能直接反映其繁殖规律,雌二醇对启动雌性小熊猫季节性繁殖起重要作用;在妊娠期内小熊猫血清孕酮含量升高不能作为判断小熊猫妊娠的标准;雌性小熊猫在妊娠期有假孕现象。     

Chin marking behavior, sexual receptivity, and pheromone emission in steroid-treated, ovariectomized rabbits

The effect of daily injections of estradiol benzoate (1 or 10 micrograms) and of progesterone (10 mg) on chin marking activity, sexual receptivity, and emission of nipple-search pheromone in ovariectomized rabbits was investigated. Both estradiol treatments resulted in a significant increase in all three measures over baseline and control group levels within 1-3 days, and withdrawal in a return to pretreatment levels within 2 weeks (Experiment I). In contrast, the administration of progesterone to such estradiol-primed does resulted in an almost immediate suppression of chin marking and lordosis, but in marked enhancement of pheromone emission and aggressive behavior (Experiment II). However, progesterone given alone to nonprimed does had no effect on any of these measure (Experiment III). The response profiles resulting from these treatments correspond well to patterns reported for intact does during estrus (= estradiol alone), pregnancy (= estradiol plus progesterone), and at parturition (= progesterone withdrawal).     

Dynamics of Blood Serum Levels of Follicle-Stimulating Hormone, Luteinizing Hormone, Prolactin, Estradiol, and Progesterone in Right- and Left-Handed Women

The dynamics of follicle-stimulating hormone (FSH) luteinizing hormone (LH), prolactin (PRL), estradiol (E₂), and progesterone were studied in left- and right-handed women having a stable 28-day menstrual cycle. The hormones were determined by enzyme immunoassays on days 3, 8, 10, 13, 16, 22, 26, and 28 of the menstrual cycle. The data showed that bllod serum levels of FSH, LH, PRL, and E₂are higher in left-handed in comparison to right-handed women (p< 0.001). On days 10 through 28 of the cycle, the level of progesterone is also higher in left-handed women (p< 0.001). The dynamic of these hormones in left-handed and right-handed women appeared to remain within the normal limits. These findings indicate that the handedness correlates with the dynamucs of serum levels of these hormones. Higher serum levels of hormones in left-handed women suggests that they have higher levels of the functional activity of the hypophysis–ovarian axis and prolactin axis.     

Evidence for the preferential utilization of esterified cholesterol in progesterone production by rabbit corpora lutea

Our previous studies show that lipoproteins stimulate progesterone secretion by rabbit luteal cells in vitro and that estradiol modifies this effect. This study examines the relationship between estradiol and serum lipoproteins for progesterone production by rabbit corpora lutea in vivo. Using morphometric analysis, we determined that estrogen treatment of hysterectomized pseudopregnant (E-hyst) rabbits increased luteal lipid volume by mid-pseudopregnancy without altering serum progesterone levels. Treatment of E-hyst rabbits with 4-amino-3,4,pyrazolo pyrimidine (APP) during early to mid-pseudopregnancy reduced serum cholesterol levels without decreasing serum progesterone concentrations. However, 3-hydroxy-3 methyl glutaryl-CoA reductase activity was increased. Thus, in the presence of exogenous estrogen, serum cholesterol is esterified and stored rather than converted directly into progesterone. APP-treatment of E-hyst rabbits during late-pseudopregnancy, when estrogen receptor levels are low, increased serum progesterone levels and reduced intracellular lipid content. Thus, stored lipid is the primary source of cholesterol for progesterone synthesis. In addition, estrogen, via estrogen receptor, is important in maintaining steady progesterone output despite fluctuations in serum lipoprotein levels. A working model for cholesterol utilization by rabbit luteal cells is presented, which suggests that stored cholesterol esters, derived from both endogenous and exogenous sources, is the key source or cholesterol for progesterone production. Furthermore, we propose that estradiol regulates the uptake and storage of cholesterol and its rate of metabolism into progesterone.     

Peripheral blood concentrations of progesterone and oestradiol during human pregnancy and delivery

To evaluate the significance of progesterone and estradiol in human uterine activity during pregnancy and delivery the blood concentrations of these hormones were monitored weekly during the last trimester of pregnancy and at the onset of labour in 15 women, and before and 3 hours after the induction of term delivery in 83 parturients. Neither plasma concentrations of progesterone or estradiol nor the ratio of progesterone to estradiol changed significantly during the last trimester of pregnancy or at the onset of delivery. After the induction of delivery parturients with initial progesterone dominance (ratio of progesterone to estradiol higher than 5 before induction) demonstrated a significant fall in serum concentration of progesterone and in the ratio of progesterone to estradiol while estradiol concentration rose significantly. In estrogen dominant women (progesterone to estradiol ratio equal to or lower than 5) the serum concentration of progesterone and the ratio of progesterone to estradiol rose significantly during the 3 hours after the induction of delivery. Our results suggest that the peripheral blood levels of progesterone and estradiol do not correlate with the tissue biochemical changes which prepare the uterine cervix and myometrium for delivery. The observation that the ratio of progesterone to estradiol decreased in progesterone-dominant and increased in estrogen-dominant women stresses the importance of a well balanced equilibrium of these hormones for prostaglandin metabolism during human delivery.     

Correlation between serum and fecal concentrations of reproductive steroids throughout gestation in goats

Non-invasive techniques such as the measurement of fecal steroids are now widely used to monitor reproductive hormones in captive and free-ranging wild-life. These methods offer great advantages and deserve to be used in domestic animals. The aim of the present study was to determine the endocrine profile of dairy goats throughout pregnancy by the quantification of fecal progestins and estrogens and assess its correlation with serum concentrations. Blood and fecal samples were collected weekly from 11 adult, multiparous goats, from mating through pregnancy and 2 weeks post-partum. The extraction of estradiol and progesterone fecal metabolites was performed by dilution in ethanol. The radioimmunoassay (RIA) in solid phase was used to quantify serum 17beta-estradiol (estradiol) and progesterone, as well as their fecal metabolites. The mean concentrations of both fecal and serum estradiol started to increase between weeks 7 and 11, reached peak values near parturition and then decreased sharply (range: 19.8+/-5.8 ng/g of feces to 608.6+/-472.4 ng/g of feces and 0.007+/-0.005 ng/ml to 0.066+/-0.024 ng/ml). An increase in both fecal and blood progestagens occurred in the second week, mean concentrations remained greater until week 20, and then decreased in the last week of gestation and 2 weeks post-partum (range: 108.8+/-43.6 ng/g of feces to 3119.5+/-2076.9 ng/g of feces and 0.12+/-0.04 ng/ml to 13.10+/-4.29 ng/ml). The changes in blood and fecal hormone concentrations were analyzed and compared throughout gestation for each single goat, for each breed and for the whole group. Results indicated that matched values of serum and fecal hormone concentrations were correlated (r=0.79; p<0.001 for progesterone and r=0.84; p<0.001 for estradiol mean concentrations in the whole group). Regression analysis showed that logarithmic model allows significant prediction of serum from fecal concentrations with an R(2)=0.729 (y=0.013ln x-0.021) for estradiol and R(2)=0.788 (y=3.835ln x-18.543) for progesterone. Neither fecal nor serum concentrations were affected by the breed but a significant effect of the number of fetuses on progestin concentrations was found. Therefore, the profiles of progesterone and estradiol fecal metabolites reflect the serum concentrations of the same hormones in pregnant goats.     

Effect of indomethacin, cycloheximide, and aminoglutethimide on ovarian steroid and prostanoid levels during ovulation in the gonadotropin-primed immature rat

It has become popular to use the gonadotropin-primed immature rat to study ovulation. The ovarian content of progesterone, estradiol, PGE2, PGF2 alpha, and 6-keto-PGF1 alpha during the ovulatory process was determined in this model. Also, the effect of three anti-ovulatory agents on the ovarian levels of the above substances was determined. At 23 days of age, Wistar rats were primed with pregnant mares serum gonadotropin (PMSG) sc, and two days later the ovulatory process was initiated with human chorionic gonadotropin (hCG) sc. The ovarian follicles began rupturing 12 h later. Ovaries were assayed for the two steroids and prostanoids at 2-h intervals before and several 4-h intervals after ovulation. The ovarian estradiol level increased slightly between 0 and 2 h after hCG, while the progesterone level increased sharply between 2 and 4 h after hCG--at a time when the estradiol declined markedly. All three prostanoids increased concomitantly with progesterone. When the PG synthesis was blocked by indomethacin treatment at 1 h before hCG, ovarian progesterone levels still increased. In contrast, when steroidogenic activity was inhibited by aminoglutethimide, the ovarian prostanoid levels also decreased. Cycloheximide had little effect on the steroids and prostanoids. It is concluded that ovarian prostanoid synthesis might be influenced by ovarian steroid output.     

Validation of the mechanisms proposed for the stimulatory and inhibitory effects of progesterone on gonadotropin secretion in the estrogen-primed rat: a possible role for adrenal steroids.

The stimulatory and inhibitory effects of progesterone on luteinizing hormone (LH) and follicle-stimulating hormone (FSH) secretion were found to be dependent on the length of estrogen exposure in ovariectomized estrogen-primed rats. Progesterone suppressed LH and FSH secretion when administered 16 hours after a single injection of estradiol to ovariectomized rats. If the estradiol treatment was extended over 40 hours by two injections of estradiol 24 hours apart, progesterone administration led to a highly significant elevation of both serum LH and FSH levels 6 hours later. In addition to the direct stimulatory effect on LH and FSH release, progesterone, when injected 1 hour before, was able to antagonize the suppressive effect of a third injection of estradiol on LH and FSH release. In the immature ovariectomized estrogen-primed rat, 10 IU of ACTH brought about a release of progesterone and corticosterone 15 minutes later and LH and FSH 6 hours later. Progesterone, but not corticosterone, appeared to be responsible for the effect of ACTH on gonadotropin release. The synthetic corticosteroid triamcinolone acetonide brought about LH and FSH release in the afternoon, while cortisol, similar to corticosterone, was unable to do so. Nevertheless, triamcinolone acetonide and cortisol brought about increased secretion of FSH the following morning.     

Estrophilic 3 alpha,3 beta,17 beta,20 alpha-hydroxysteroid dehydrogenase from rabbit liver--II. Mechanisms of enzyme-steroid interaction

Binding of [3H]estradiol, [3H]testosterone and [3H]progesterone to purified NADP-dependent estrophilic 3 alpha,3 beta,17 beta,20 alpha-hydroxysteroid dehydrogenase (EHSD) from rabbit liver cytosol has been examined. The three steroids bind to the enzyme with moderate [corrected] affinity (Ka congruent to 10(7) [corrected] M-1 at 4 degrees C) and equal binding capacity. High-rates were shown for both association and dissociation processes. The steroids competitively inhibited the binding of each other to EHSD. At the same time, their relative binding affinities (RBA) were dependent on the nature of [3H]ligand. The results of RBA determinations for 72 steroids and their analogues by inhibition of [3H]progesterone binding to EHSD suggest that androgens and gestagens bind preferentially to the same site on EHSD molecule, while estrogens (at least by their D-ring) bind to another site. The assumption that EHSD molecule has more than one binding site for steroids is corroborated by (i) substrate inhibition revealed for a number of steroids; (ii) the estrogen ability to potentiate 20 alpha-reduction of progesterone; (iii) stimulatory effect of 5 alpha (beta)-androstane-3 alpha (beta), 17 beta-diols on [3H]testosterone and progesterone binding; and (iv) reciprocal effect of NADP on [3H]estradiol and [3H]testosterone binding to EHSD. Significant differences in sensitivity to pH and changes in NaCl concentration upon metabolism and binding of various steroids have been found. At concentrations of 16 mM dithiothreitol potentiated catalytic conversion of some steroids and had no effect on metabolism of others. Both the affinity for steroids and binding capacity of EHSD are found to be cofactor-dependent. It is speculated that EHSD has a complex active center including at least two mutually influencing steroid-binding sites tightly related with cofactor-binding site. The polyfunctionality of EHSD may be due to both the excess of functional protein groups that form individual constellations upon binding of any steroid and also to conformational lability of EHSD molecule implying alternative orientations of steroids at the binding site.