Receptor Replenishment Research Articles

Replenishment and nuclear retention of oestradiol-17β receptors in rat uteri during postnatal development

1. 1. Uteri of 6–10-day-old rats do not show a late growth response to oestrogen (increase in rate of DNA synthesis and cell division) exhibited by fully competent (20 days or older) uteri. We posed the question whether the lack of the late growth response is due to an inability to replenish the cytoplasmic pool of oestrogen receptors or to curtailed retention of oestrogen binding in the nucleus. Uterine nuclear and cytoplasmic receptors were measured by a [ 3H]oestradiol-17β exchange assay, at 1, 3, 6, 14 and 24 h after oestrogen injection. 2. 2. The replenishment of cytoplasmic oestrogen receptors showed a similar pattern in the uteri of 6 and 10-day-old (partially responsive) and in 20-day-old (fully responsive) rats. 3. 3. Oestrogen was retained longer in uterine nuclei obtained from 5 and 10-day-old rats than in uterine nuclei of 20 and 25-day-old rats. 4. 4. Oestrogen receptors resistant to 0.4 M KCl extraction (residual receptors) were found in uterine nuclei of 6 and 25-day-old rats after oestrogen injection at all the times tested. The concentration of these residual receptors during the late period (6–24 h after injection) was not significantly different in uterine nuclei of 6-day-old and 25-day-old rats. 5. 5. We conclude that neither lack of oestrogen receptor replenishment nor curtailed retention of oestrogen binding in the nucleus is the factor which limits the complete responsiveness to oestrogen in uteri of rats during postnatal development.

The Uterine Endometrium and Isthmic Oviduct of the Ewe: Does Progesterone Act as an Antiestrogen?

The action of progesterone (P) on the estrogen stimulated uterine endometrium and isthmic oviduct of the ewe was studied. Changes in protein:DNA and RNA:DNA ratios, rate of in vitro protein synthesis, alkaline phosphatase and peroxidase activity and the level of cytoplasmic receptors for estradiol (E2) and P were measured in the endometrium. Only receptor effects were measured in the oviduct. Progesterone decreased the level of both receptors in each tissue but did not decrease protein:DNA and RNA:DNA ratios or protein synthesis; P had little effect on alkaline phosphatase activity and there was no detectable peroxidase activity. These findings argue against the concept, based on studies in the rat, that the initial antiestrogenic effects of P are mediated by a specific effect on E2 receptor replenishment.

The Relevance of Interaction Kinetics in Determining Biological Response to Estrogens

The study of the kinetics of the interaction between two ethynyl estradiol derivatives (llβ-methoxy-17-ethynyl-l,3,5(10)-estratriene-3,17β-diol and llα-methoxy-17-ethynyl-l,3,5(10)-estratriene-3,17β-diol), differing in the configuration of the 11-methoxy substituent, and the cytoplasmic estrogen receptor in mouse uterus and the study of their ability to induce short and long term estrogen responses (uterine weight increase, estrogen receptor replenishment, and progestin receptor induction) in the mouse have led to the following conclusions. Potent agonists are compounds which form slowly dissociating cytoplasmic receptor complexes. Low doses of weak agonists are able to induce short term responses to the same extent as potent agonists but are unable to maintain these responses; higher doses or frequent administration are required to maintain an effect. The degree of agonist activity is related to the nuclear estrogen receptor concentration, which is in turn related to the ability of the ligand to form a stable cytoplasmic receptor complex. Low doses of weak agonists can effectively compete with endogenous hormone for cytoplasmic receptor sites and thus prevent the endogenous hormone from inducing its full biological response. Since they are themselves unable to induce a full response, this competition results in antihormonal activity.

Mechanisms involved in the regulation of steroid receptor levels

The turnover of steroid receptors comprises: synthesis in the cytoplasm, translocation into the cell nucleus and degradation at a still unknown site. From studies on the oestradiol/receptor system, the following conclusions can be drawn. 1. The in-vivo uptake of oestradiol by target cell nuclei is receptordependent. Steroid and receptor are translocated from the cytoplasm in a 1:1 ratio. A recycling of receptor is undetectable after pulse administration of oestradiol. Receptor replenishment in the cytoplasm is accomplished by synthesis. 2. The nuclear uptake of receptor-in contrast-proceeds also in the absence of oestradiol. Both forms of receptor, monomer and “activated” dimer are present in oestrogen-free nuclei. 3. Oestradiol enhances the “nucleotropy” and the turnover rate of receptor. 4. Oestrogenicity and antioestrogenicity are apparently linked to effects exerted on the “nucleotropy” of receptor and its ability to interact with the relevant nuclear structures.

The effects of antioestrogens on the oestrogen receptor

In order to probe the source of the antagonistic properties of triphenylethylene antioestrogens in oestrogen induced uterine growth, we have studied both low- and high-affinity antioestrogens. In in vitro competition studies H-1285 was shown to have about 1100% the affinity for the oestrogen receptor when compared with 17-β-oestradiol (E 2) (100%) which is in sharp contrast to H-1067 and U-11, 100A (UA) which have 25 and 4% binding affinities, respectively. H-1285 and H-1067 both caused an increase in uterine DNA and wet weight, but were impeded oestrogens. H-1285 and H-1067 were also antioestrogenic because they inhibited the E 2-induced DNA and wet weight responses. The in vivo nuclear and cytoplasmic oestrogen receptor responses to UA, H-1067 and H-1285 were also investigated and compared to the E 2 response. All antioestrogens studied caused a prolonged nuclear retention of the oestrogen receptor, in contrast to the rapid uptake followed by a rapid deletion seen with E 2. H-1285 receptor complexes, however, displayed a delayed uptake, which did not reach maximal nuclear levels until 24 h. Cytoplasmic oestrogen receptor replenishment with E 2 was immediate and rapid. All the antioestrogens studied demonstrated delayed and slow receptor replenishment, about 25% the E 2 induced rate. E 2 injected at 48 h subsequent to an injection of H-1285 caused a rapid 1-h nuclear translocation of the oestrogen receptor followed by a complete nuclear “fallout” of receptor, a phenomenon previously seen only with oestrogens. No salt-resistant oestrogen receptor complex was measurable with a high-affinity antioestrogen H-1285 or with low affinity antioestrogens, H-1067, UA or enclomiphene during 1–12 h, but the salt-resistant form was seen with E 2 and diethylstilboestrol at 1–6 h. All of the oestrogen responses (wet weight, protein content, DNA and protein synthesis) exhibited the same increase at 24 h whether single 5 μg E 2 or H-1285 injections were administered. However, by 48 and 72 h, H-1285 consistently gave higher responses than did E 2. E 2 administered at 48 h subsequent to a previous E 2 injection caused a dramatic increase in oestrogenic responses, in contrast to E 2 given subsequent to H-1285 which caused only a slight increase in these responses. These data demonstrate the ability of H-1285 to inhibit oestrogen induced uterine responses.

Receptor Depletion and Replenishment Processes: In vivo Regulation of Gonadotropin Receptors by Luteinizing Hormone, Follicle Stimulating Hormone and Ethanol in Rat Testis

The effect of in vivo injections of human lutemnizing hormone (hLH), human follicle stimulating hormone (hFSH) and ethanol on gonadotropin receptor sites in rat testes was studied. We found that hLH (32 IU) and hFSH (30 IU) reduced the number of their respective binding sites shortly after administration. The hLH was more rapid and effective in its action than hFSI-I. Hormone specificity was also evident in the depletion process. The hLH could not effectively deplete testicular hFSH receptors during the first 3 days and vice versa. Prolonged administration of ethanol (5 ml of 20% v/v ethanol in saline/injection; 2 injections! day for 7 days) also led to depletion of gonadotropmn receptor sites in rat testes but without the specificity displayed by hLH and hFSH. The receptor replenishment process commenced when ethanol was withdrawn and was essentially complete by Day 20 (or 13 days after ethanol withdrawal). The ability of ethanol to reduce gonadotropmn binding through receptor depletion may be a possible cause of testicular atrophy, reduced steroidogenesis and impotency, problems frequently encountered in chronic alcoholics.

Glucocorticoid modulation of prolactin receptors on mammary cells of lactating mice

Prolactin receptors were monitored by measuring 125I-labeled prolactin binding to collagenase-dissociated mammary epithelial cells of lactating BALB/c mice. Specific receptors for iodine-labeled prolactin with an apparent dissociation constant ( K d) of 0.99 · 10 −9 M were present on the dissociated mammary cells. The binding was inhibited by ovine prolactin, human growth hormone and human placental lactogen but not by follicle stimulating hormone, luteinizing hormone, thyroid stimulating hormone, bovine growth hormone or insulin. Adrenal ablation of nursing mothers caused a reduction of the number of prolactin receptors and this effect was preventable by hydrocortisone therapy. Hydocortisone injections to mothers 3 days after adrenalectomy also induced a replenishment of the prolactin receptors on the mammary cells. Injections of progesterone failed to sustain the high level of mammary cell prolactin receptors in adrenalectomized animals. Stimultaneous injections of hydrocortisone and progesterone to animals 3 days after adrenalectomy caused a partial suppression of the stimulatory action of hydrocortisone alone. The results suggest that hydrocortisone can exert a modulatory influence on mammary cell prolactin receptors in non-hypophysectomized post-partum mice without altering the dissociation constant ( K d) of the receptors.

Cytoplasmic oestrogen receptor replenishment: oestrogens versus anti-oestrogens.

The effect of the in vivo administration of various triphenylethylene antioestrogens and physiological (0.05 microgram) versus pharmacological (0.5--5 microgram) doses of oestradiol-17 beta (Oe2) on the uterotrophic process in general and the nuclear accumulation and cytoplasmic depletion and replenishment of uterine oestrogen receptor was determined. Regardless of the dose of Oe2 the changes in uterine wet weight, total protein and incorporation of [3H]thymidine into DNA and [14C]leucine into protein were the same at 24 h. The anti-oestrogen receptor was also studied. The net increase, above control, of cytoplasmic oestrogen receptor at 24 h and 48 h after Oe2 injection was approximately 0.35 and 0.77 pmoles/uterus, respectively. The effect of anti-oestrogens (U-11,100A, CI628, en- and zuclomiphene) on the increase in cytoplasmic oestrogen receptor at 24 h and 48 h measured from maximally depleted levels was nearly identical to the Oe2 induced net increase. This suggests that in both cases these particular increases represent newly synthesized receptor and that Oe2 causes some receptor replenishment through a recycling process.

Changes in the concentration of uterine cytoplasmic oestrogen receptors induced by doxorubicin and methotrexate.

The effects of doxorubicin and methotrexate on the oestradiol-induced depletion, replenishment and subsequent increase beyond the normal value (overshoot) in the number of uterine cytoplasmic oestrogen receptors were investigated in intact rats. Injection of doxorubicin (0.5 mg/kg, i.v.) or methotrexate (1mg/kg, i.m.) 5 min after a single i.p. injection of 10 ng oestradiol-17beta (which is able to induce a 50% depletion in the number of oestrogen receptors) caused a significant increase in the oestradiol-induced depletion. Both drugs inhibited the replenishment and the overshoot phases until 48 h after treatment, although the effect was more marked with doxorubicin. Experiments in vitro showed that both methotrexate and doxorubicin affected the capacity of oestradiol-17beta to bind to specific cytoplasmic receptors, inducing an increase in binding when used at low concentrations and a decrease at higher concentrations. The effects of doxorubicin and methotrexate on the depletion, replenishment and overshoot of oestrogen receptors seemed to be partly dependent on the inhibition of protein synthesis and partly due to direct action on the binding of oestradiol-17beta to its receptors.

Estrogen action in the mouse uterus: An additional nuclear event

An in vivo injection of 3H-estradiol to an ovariectomized mouse resulted in the appearance of two increases of nuclear receptor: the first at 1 hr and the second occurring at approximately 7–8 hrs post-injection. These findings were substantiated by quantifying cytoplasmic and nuclear receptor levels with the exchange assay after injections of unlabeled estradiol. Cytosol receptor levels were depleted at 0.5–1 hr and replenished from 2–10 hr with values at 15–24 hr significantly greater than those at zero time. Nuclear receptors decreased precipitously to near control values after the initial translocation event. At 7–8 hr a second smaller nuclear increase occurred which then declined to near zero levels. Both of these nuclear events appear to be dose related. Similar experiments using cycloheximide did not abolish either of these events, but did significantly diminish the cytosol receptor replenishment.

Relationship between the estrogen induced protein IP and other parameters of estrogenic stimulation. A hypothesis

Data concerning the estrogen-induced protein “IP” discovered by Notides and Gorski are reviewed. Estrogen specificity of the IP response, physiological occurrence of IP and inducibility of an IP-like protein in target tissues other than the rat uterus are considered. Available information concerning the relative ability of different estrogenic and anti-estrogenic compounds to promote IP synthesis, the uterotrophic responses and the receptor depletion-replenishment cycle is examined. From this the hypothesis is proposed that IP induction may be related with receptor replenishment. Recent data concerning the “receptor cycle” are briefly reviewed in order to envisage further extension of our hypothesis.

Progesterone inhibition of estrogen receptor replenishment in ovine endometrium.

The influence of exogenous progesterone and estradiol on the concentration of total cytoplasmic and nuclear estrogen receptor in endometrium of ovariectomized ewes was studied and the effect of the respective steroid treatments on the receptor-mediated translocation of estrogen to the nucleus determined. 16 ovariectomized ewes were assigned to treatment groups as follows: Group 1 controls; Group 2 estradiol-17beta (EZ) implants (silastic capsules packed with 54 + or -3 mg); Group 3 progesterone (P) (15 mg twice daily); and Group 4 EZ + P. Control and P-treated animals were implanted with empty silastic capsules. Blood samples were collected on Days 0 and 2-7 for analysis for EZ and P. Ewes were sacrificed on Day 7 and the uteri were removed. Total specific nuclear and cytoplasmic estrogen binding sites were determined by use of tritiated estradiol exchange assay. Tritiated E2 bound to total cytoplasmic and nuclear receptors was greater (p less than .05) in endometirum of E2 treated ewes (10.4 + or -7.27 and .23 + or -.06 fmoles/mcg DNA respectively) than in endometrium of control P- and E2 + P-treated animals (1.62 + or -.26 and .13 + or -.02 fmoles/mcg DNA respectively). An average of 81 + or -1% of the tritiated E2 bound to available cytoplasmic receptors was translocated to the nucleus during incubation of endometrium of ewes from all treatment groups. The concentration of tritiated E2 specifically bound in the cytoplasm and translocated to the nucleus during incubation was greater in the endometrium of controls P-treated and E2 + P-treated groups. Concentrations of labeled E2 in the cytoplasm after in vitro translocation was greater in the endometrium of E2-treated ewes than in the endometrium of the other treatment groups. It is suggested that P antagonizes estrogen binding in the uterus by decreasing the concentration of cytoplasmic estrogen receptor.

Progesterone modulation of estrogen-stimulated uterine biosynthetic events and estrogen receptor levels

The effect of progesterone on estrogen-stimulated biosynthetic events — namely, glucose metabolism, fluid imbibition, DNA synthesis, and replenishment of estrogen receptor — is studied in uteri of immature (22–24-day-old) rats and immature estrogen-primed rats. Pretreatment with progesterone alone (2 mg s.c./rat) for up to 48 h does not markedly influence the subsequent uterine response to estrogen in terms of 2-h deoxyglucose metabolism, 3-h uterine wet weight or 24-h DNA synthesis rate. Moreover, progesterone alone increases uterine glucose metabolism and the rate of DNA synthesis, as does estradiol (5 Mg), although the magnitude of the responses obtained is lower with progesterone. However, simultaneous exposure to progesterone plus estradiol (E + P) for over 12 h results in a decreased uterine responsiveness to subsequent estrogen as monitored by glucose metabolism and uterine wet weight stimulation. This decreased responsiveness to estrogen after exposure to E + P correlates with a decreased content of uterine cytosol estrogen receptor. After treatment with E (5 Mg) plus P (0.5 or 2 mg), or E alone, the initial movement of the estrogen receptor into the nucleus is similar, but by 12 h after E + P the rate of reappearance of the cytosol receptor is slower and the levels of cytosol receptor are depressed, reaching only 50–60% of that seen after E alone. This depression of cytosol receptor replenishment is most marked with the steroid progesterone, although exposure to estradiol (5 μg) plus high doses of dihydrotestosterone or testosterone (2 mg), but not low doses (0.5 mg), results in slightly depressed levels of cytosol receptor at 12–48 h. Cortisol (0.5 or 2 mg) is without any effect. Administration of either progesterone or the inhibitor cycloheximide or actinomycin D at zero time (along with the estradiol) or up to 4–6 h following estradiol injection is markedly effective in decreasing the levels of estradiol cytoplasmic receptor (to approx. 40–50% of the control, E alone, level by 12 h) in 3-day estrogen-primed rat uteri. The striking similarity in the time course and extent of the cycloheximide-, actinomycin D-, and progesterone-sensitive inhibition of estrogen receptor replenishment suggests that progesterone may influence uterine sensitivity to estrogen by interfering with the de novo synthesis of new estrogen receptors.

Different nuclear binding sites for antiestrogen and estrogen receptor complexes.

Experiments were performed to determine the in vivo effect of various estrogens and anti-estrogens on the nuclear accumulation and retention of estrogen receptors, the cytoplasmic levels of estrogen receptors, and the formation of salt-resistant and salt-extractable forms of the nuclear estrogen receptor in immature rat uteri. A 5 mug injection of estradiol-17beta (E2) or diethylstilbestrol (DES) resulted in a maximal nuclear translocation of the receptor complexes by 1 h with a subsequent rapid decrease of both the estradiol receptor complex (ERC) and diethylstilbestrol receptor complex (DRC) to levels found in uteri of saline-injected rats by 12 h. However, the antiestrogens U-11,100A, zuclomiphene and enclomphene (100 mug/injection) resulted in a slower nuclear accumulation of receptor complex which continued to increase through 24 h. The cytosol receptor levels with E2 and DES were depleted to 10--20% of control levels within 1 h, but then were replenished so that they were above control levels by 24 h. The clomiphene-type compounds also showed an initial depletion of cytosol estrogen receptor, but the antiestrogens were almost ineffective in receptor replenishment. The estrogen receptor translocated to the nuclear fraction by estrogens demonstrated both salt-extractable (0.3M KCl) and salt-resistant forms at 1--6 h, whereas the clomiphene-type compounds resulted in the formation of only a salt-extractable form of the estrogen receptor at all times. By 12--24 h after injection, the salt-resistant forms of the ERC and DRC were no longer present. The effect of varying the dosage of injected E2 (0.05 mug-5 mug) resulted in the formation of an identical amount of salt-resistant ERC at 1--2 h, whereas the total amount of nuclear ERC (salt-resistant and salt-extractable) varied with the injected dose of E2. However, at 6 h, the amount of salt resistant ERC varied with the injected dose of E2 (0.005-5 mug). These results suggest that the nuclear salt-resistant form (formed by estrogens only) of the estrogen receptor is required for true uterine growth, whereas the nuclear salt-extractable form may be only sufficient for short term estrogenic responses.

ENDOMETRIAL HYPERPLASIA AND CARCINOMA: HISTOPATHOLOGY AND HORMONAL FACTORS

The actions of estrogen (E) and progestogen (P) on normal endometrium are reviewed in order to give insight into endometrial hyperplasia and carcinoma. The entry of E from plasma may be protein-mediated. Human endometrium slices metabolize estradiol-17beta to an extent which varies with the stage of the cycle. Endometrium with cystic or anovulatory hyperplasia or adenocarcinoma behaves as proliferative endometrium. P in rats blocked E-induced replenishment of cytoplasmic receptors and caused a decrease in the quantity of cytoplasmic E receptors. Estrone and estriol had less affinity in binding to receptors than estradiol-17beta and this is important as a factor in the biological effect of these steroids. It has been noticed that the length of time the estrogen-receptor complex remains in the nucleus is a factor of critical importance for the biological potency as well. Weak estrogens may be as effective as estradiol-17beta if administered often enough. P enters the target cells through simple diffusion. Estradiol-17beta induces synthesis of new P uterine receptors while P itself seems to inactivate its own receptor. The role of E in the development of endometrial cancer is discussed. Results from a Swedish study suggest that prolonged estrogen stimulation unopposed by P can produce endometrial hyperplasia with a progression of changes leading to endometrial carcinoma. It is suggested that until the role of E in endometrial carcinoma is elucidated E should be administered only for established indications and only in a cyclical manner.

The excitation and inhibition of sexual receptivity in female hamsters by progesterone: time and dose relationships, neural localization and mechanisms of action.

Ovariectomized hamsters were administered estradiol benzoate (EB) and 44 h later, progesterone (P.) Lordosis behavior was induced. When an additional dose of P was given up to 24 h prior to or 24 h after the EB, EB-P facilitation of lordosis was inhibited. Additional hamsters were given varying doses of P (25-200 mu) following EB using both excitatory and inhibitory paradigms. Inhibition of EB-induced lordosis was effected with a lower dose of P than was the facilitation of EB induced lordosis by P. Hamsters were also given intracerebral implants of P using excitatory and inhibitory paradigms. No excitatory loci were found. Inhibition of EB-induced lordosis was effected by implants in the posterior hypothalamus and anterior mesencephalon, but not by diencephalic implants. Other hamsters were administered tritiated estradiol (E2) plus P prior to, concurrent with, or shortly after the E2. P had no effect upon the accumulation of E2 by any brain sites, although E2 was found to concentrate to a greater degree in the diencephalon than in the mesencephalon or cortex. The estrogen-induced depletion and replenishment of hypothalamic cytosol estrogen receptors was also studied. Concurrent P treatment had no effect upon the receptor depletion-replenishment process. It was concluded that P can both facilitate and inhibit estrogen-induced lordosis and that the inhibitory effects of P are not upon estrogen-sensitive cells in the brain.

Kinetics of glucocorticoid-receptor complexes in rat thymus cells

Using cortisol and dexamethasone, we have studied the kinetic behavior of glucocorticoid receptors in intact rat thymus cells at 37°C. As reported previously, translocation of the cortisol-receptor complexes from “cytoplasmic” to nuclear-bound form takes about 1 min. At 26°C this time is doubled. Consistent with these observations and with the current view that formation of nuclear-bound complex must be preceded by formation of cytoplasmic complex, on addition of hormone to cells at 37°C the initial time-course of formation of cytoplasmic complex precedes that of nuclear complex by about 30 s. After cells have achieved a steady state with [ 3H]-dexamethasone, however, a “chase” of unlabelled dexamethasone lowers nuclear levels of [ 3H]-dexamethasone several minutes before cytoplasmic levels, suggesting formation of nuclear complex by direct reaction of hormone with nuclear-bound receptors. Following depletion of cytoplasmic receptor by incubation with a high concentration of dexamethasone and then sudden lowering of steroid concentration by dilution, the time-course of replenishment of cytoplasmic receptors appears to lag significantly behind the time-course of removal of nuclear-bound steroid. The lag is roughly the same magnitude as the time-constant of dissociation of the glucocorticoid used, and is much shorter with cortisol. The pathway of replenishment thus may be different from simple reversal of the pathway of depletion of cytoplasmic receptor, and may depend on the dissociation rate or affinity constant of the steroid.

Sex-Related Differences in the Regulation of Cytoplasmie Estrogen Receptor Levels in Responsive Tissues of the Rat

Cytoplasmic estrogen receptors were measured in the anterior pituitary and hypothalamus of intact adult Holtzman male and female rats at various time intervals following injection of 10 mug of 17beta-estradiol. Following equivalent depletion of receptors at 1 h in either sex, replenishment of cytosol receptor levels progressed at a greater rate and to a higher level in the male than in the female. Cycloheximide only partially inhibited replenishment, and its blocking effect was similar in males and females. The administration of 5 mug of 17beta-estradiol to adult males elicited less depletion than did 10 mug, and a completely cycloheximide-sensitive replenishment phase. Patterns of depletion and replenishment in immature rats following 0.1 mug of 17beta-estradiol were the same for males and females. In an analysis of the effects of neonatal androgenization, Charles River rats were used and it was initially demonstrated that no sex differences in receptor levels existed between normal males and females. After an injection of an androgenizing level of testosterone propionate on day 3 of age, a full normal complement of receptors was found in estrogen-responsive tissues at 90--100 days of age. The functional dynamics of receptor depletion and replenishment under the influence of exogenous estrogen, however, indicated that the anterior pituitary of the androgenized female responded differently from that of the untreated female, and similarly to that of the male. The hypothalamic receptor dynamics in the male were unaltered by early androgen administration, but estrogen-induced depletion of receptors in the female hypothalamus was considerably less extensive in the androgenized animal than in the untreated control. Whereas cytoplasmic estrogen receptor levels are essentially independent of sex in the rat, the present results indicate a sex-linked difference in the dynamics of receptor turnover in the anterior pituitary and hypothalamic response to 17beta-estradiol.

Estrogen‐receptor binding: Relationship to estrogen‐induced responses

The relationship between nuclear binding of the receptor-estrogen complex (R-E) and estrogenic responses was examined. Nuclear-bound R-E was measured by the [3H] estradiol exchange assay, which permits the quantification of R-E as a function of either endogenous or non labeled estrogen. The quantity of nuclear R-E fluctuated as a function of blood levels of estrogen during the estrous cycle in the rat. This indicates that the accumulation of the R-E complex by the nucleus of uterine cells may be of physiologic significance and under the control of endogenous estrogen.

Actions of Pregnant Mare Serum Gonadotropin in the Immature Female Rat: Correlative Changes in Blood Steroids, Gonadotropins, and Cytoplasmic Estradiol Receptors of the Anterior Pituitary and Hypothalamus11

Several blood steroids, serum gonadotropins and cytosol estradiol receptors of the anterior pituitary and hypothalamus were quantified in immature female rats which were induced to ovulate with pregnant mare's serum gonadotropin (PMSG). Studies revealed that serum levels of progesterone, 17-hydroxyprogesterone, testosterone, androstenedione and estradiol were initially elevated at 6 PM (day 30) after administration of 8 IU of PMSG at 10 AM day 30. Serum levels of estradiol and testosterone rose progressively from day 30 through the AM of day 32. A further increase in serum concentrations of progesterone, 17-hydroxyprogesterone, androstenedione, testosterone, and dehydroepiandrosterone occurred on the PM of day 32 whereas serum estradiol levels declined. Serum levels of all steroids declined on the day of estrus (33) and only progesterone levels were further elevated on day 34 (diestrus). Dihydrotestosterone concentrations were minimally altered by PMSG treatment. Saline administration resulted in no significant alterations in levels of any steroid quantified from day 29 to 34 in control animals. A progressive decline in cytosol estradiol receptor content of the anterior pituitary and hypothalamus was documented following PMSG treatment of intact female rats; there was no depletion of receptors following PMSG administration to ovariectomized immature rats. Maximal depletion of cytosol estradiol receptors occurred on day 32 with replenishment of cytosol estradiol receptor levels on estrus (day 33). The preovulatory gonadotropin surge was found to occur on the PM of day 32 after maximal receptor depletion. The cycle of depletion and replenishment of receptors was repeated during a second spontaneous estrous cycle four days later which coincided with a rise and fall in serum estradiol levels. It is suggested that the depletion of cytosol estradiol receptors of the anterior pituitary/hypothalamic unit may be causally related to the preovulatory gonadotropin surge resulting from PMSG administration to immature female rats. In addition, changes in blood steroids and gonadotropins after PMSG treatment are similar to those reported for proestrus-estrus-diestrus I of the normal adult estrous cycle. These findings further demonstrate the validity of the PMSG-primed immature female rat preparation as a model for the estrous cycle of the adult rat.