A double antibody radioimmunoassay for free and conjugated estradiol-17 beta in cow's milk

A radioimmunoassay for free estradiol-17 beta, conjugated estradiol-17 beta or total (free + conjugated) estradiol-17 beta in defatted milk of cows is described. Conjugated estradiol-17 beta was hydrolyzed by enzymes of Helix pomatia juice. Estrogens were extracted with dichloromethane; no other purification step was required before radioimmunoassay because of the high specificity of the antiserum. Immunoprecipitation was used to separate bound and free estradiol-17 beta. Concentrations measured were corrected for procedural losses on a per sample basis. The assays were shown to be accurate and specific. The sensitivity was 1.3pg/ml for the assay of free estradiol-17 beta (5ml of milk extracted) and 2.9pg/ml for conjugated or total estradiol-17 beta (2 ml of milk hydrolyzed and extracted). Estrogens were measured in the milk of cyclic cows and in cows stimulated with pregnant mare serum gonadotropin (PMSG). A preovulatory increase was clearly observed. Wether or not the ovary was stimulated by PMSG, concentrations of estrogens were higher and the relative increase during the preovulatory peak was greater for conjugated estradiol-17 beta than for the free form. The assay of conjugated or total estradiol-17 beta in defatted milk should be a practical method for assessing preovulatory growth of follicles in cows.     

The influence of estradiol-17beta and progesterone on peripheral serum concentrations of luteinizing hormone and follicle stimulating hormone in the ovariectomized ewe

Serum gonadotropin concentrations were high and variable and fluctuated episodically in short and long term ovariectomized ewes. Treatment with solid silastic implants releasing progesterone (serum levels 1.81 +/- 0.16 ng/ml) had no consistent effect. Treatment with implants releasing estradiol-17beta significantly depressed mean serum gonadotropin concentrations and peak height to values usually seen in intact ewes. This occurred regardless of implant size and serum estradiol-17beta concentrations (range 11 +/- 0.3 pg/ml to 98 +/- 12.8 pg/ml). Progesterone and estradiol-17beta together significantly depressed the frequency of peaks in LH concentration. Following progesterone removal, 95% of the ewes treated with progesterone and estradiol-17beta implants experienced a transient increase in serum LH concentrations similar to the preovulatory surge in intact ewes. Eighty-four percent of the LH surges were accompanied by a surge in serum FSH concentrations. However, following progesterone removal, 5.1 +/- 2.1 FSH surges were observed over six days. Gonadotropin surges occurred regardless of estradiol-17beta implant size and with or without the influence of supplemental estradiol-17beta.     

Serum estradiol-17beta concentrations during spontaneous silent estrus and after prostaglandin treatment in the mare

Serum estradiol-17beta concentrations were determined during silent estrus in the mare. Relationships between serum estradiol-17beta concentration, corpus luteum regression, follicular development, ovulation, prostaglandin treatment and behavioral estrus were investigated. The expression of behavioral estrus was found to be related to the patterns of progesterone and estradiol-17beta secretion during the periovulatory period. When compared to normal estrous cycles, silent estrus was accompanied by a significantly lower maximum serum estradiol-17beta concentration (47.8 vs 34.6 pg/ml), a significantly longer interval from maximum estradiol-17beta concentration to ovulation (1.7 vs 4.0 days), and a significantly shorter interval from corpus luteum regression to ovulation (5.3 vs 2.8 days). Silent estrus following prostaglandin treatment was related to a significantly shorter interval from prostaglandin treatment to ovulation (3.6 +/- 0.4 days) than from normal corpus luteum regression to ovulation (5.3 +/- 0.3 days). Silent estrus appeared to be related to changes in follicular estradiol-17beta secretion and to the pattern of its secretion as related to regression of the corpus luteum. There appeared to be not only less estradiol-17beta present, but also less time available after luteal regression for it to interact with the central nervous system to elicit the changes necessary to cause behavioral estrus. There fore, unusual relationships between luteal function and folliculogenesis can result in one type of silent estrus. Significant correlations (P<0.05) were found between follicle size and serum estradiol-17beta concentration whenever behavioral estrus occurred [follicle diameter in mm = 0.96 (serum estradiol-17beta in pg/ml) + 6.08 and 0.73 (serum estradiol-17beta + 13.32 for control and normal estrus following prostaglandin treatment groups, respectively]. During silent estrus, however, no significant correlations between follicle size and serum estradiol-17beta concentration were observed.     

Oxytocin and bovine parturition: a steep rise in endometrial oxytocin receptors precedes onset of labor.

Oxytocin receptors were measured in myometrium and intercaruncular endometrium of cows during pregnancy and parturition. Concentrations of estradiol-17 beta, estrone, and progesterone in peripheral blood were also measured. Receptor concentrations in the endometrium rose almost 200-fold from Day 20 to term (p < 0.0001, ANOVA), from 40 +/- 11 to 7300 +/- 1430 fmol/mg protein. Myometrial receptor concentrations increased 10-fold from 180 +/- 36 fmol/mg on Day 20 to 1850 +/- 360 fmol/mg protein at term (p < 0.0001, ANOVA). During labor, endometrial receptors (6600 +/- 1300 fmol/mg) remained at prelabor values, whereas myometrial receptor concentrations had decreased to 1190 +/- 316 fmol/mg (not significant) and declined further postpartum. Plasma concentrations of progesterone declined from 4-5 ng/ml to about 2 ng/ml between Days 250 and 282 and dropped to < 0.2 ng/ml shortly before delivery. Plasma concentrations of estrone and estradiol-17 beta were below 10-20 pg/ml until Day 230. Estrone concentrations were significantly (p < 0.05) increased by Day 250 and estradiol-17 beta by Day 270, and then both rose rapidly. During labor, plasma estrone was 1135 +/- 245 pg/ml and plasma estradiol-17 beta was 226 +/- 131 pg/ml. The molar ratio of estrone and estradiol-17 beta to progesterone rose from less than 0.01 to 4.4 during labor, and was correlated with oxytocin receptor concentrations in endometrium (r = 0.5160, p < 0.001), but not those in myometrium (r = 0.0122). The regulation of oxytocin receptors by ovarian hormones in the two tissues may therefore differ.(ABSTRACT TRUNCATED AT 250 WORDS)     

Catalytic competence, a new criterion for affinity labeling. Demonstration of the reversible enzymatic interconversion of estrone and estradiol-17 beta covalently bound to human placental estradiol-17 beta dehydrogenase.

Human placental estradiol-17beta dehydrogenase is rapidly inactivated upon treatment with 3-bromoacetoxyestrone. Pseudo-first order kinetic data are obtained and inactivation is accompanied by incorporation of 1 mol of 3-acetoxyestrone/mol of subunit (Mr =34,000). Treatment of the inactivated enzyme with (4S)-[4-2H]DPNH results in the formation of covalently bound [17alpha-2H]estradiol-17beta, which can be released by hydrolysis and identified by gas chromatography-mass sepctrometry. When (4R)-[4-2H]DPNH was used, deuterium was not transferred. Thus, the normal stereochemistry of hydridetransfer is preserved for both partners. After treatment with p-mercuribenzoate, affinity-labeled estradiol-17beta dehyrogenase is no longer able to caralyze reduction its covalently bound estrone; in the presence of DPNH and native enzyme, however, reduction occurs, demonstrating that affinity-labeled enzyme can itself serve as subtrate for native estradiol-17beta dehydrogenase. The reversible enzymatic interconversion of covalently bound estrone was demonstrated using a transhydrogenase assay. The ability of an enzyme to catalyze its normal reaction with a covalently bound substrate is termed catalytic competence, and is considered to be a new criterion for affinity labeling.     

A sensitive and specific enzymeimmunoassay for serum testosterone.

A very sensitive enzymeimmunoassay for testosterone was developed using testosterone-penicillinase conjugate and an antibody to testosterone-3-(O-carboxymethyl)oxime-bovine serum albumin. The specificity of the assay was demonstrated by the fact that estradiol-17 beta, estrone, estriol, progesterone, 17 alpha-hydroxy-progesterone, dehydroepiandrosterone, androstenedione, cortisol and cortisone were ineffective in crossreacting with testosterone while dihydrotestosterone was 8 times less crossreactive as compared to testosterone. The minimum detectable amount of testosterone was 10-15 pg per assay tube. Intra-assay and inter-assay coefficients of variation for samples containing 0.3-6ng/ml of testosterone were 6-8% and 8-10%, respectively. A high degree of correlation (r = 0.97) was observed between serum testosterone values obtained by enzymeimmunoassay and radioimmunoassay. The levels of testosterone in the sera of normal men and women and those in hypogonadal males following stimulation with human chorionic gonadotropin determined by this enzymeimmunoassay appear similar to those reported by other investigators.     

Gonadal steroid hormone level in blood plasma and milk of primiparous and multiparous nonpregnant and pregnant buffaloes

Changes in the concentration of progesterone and estradiol-17beta were measured by radioimmunoassay in 11 primiparous and 17 multiparous buffaloes at estrus and daily post insemination and in 6 nonbred buffaloes at 6 hour intervals from 4 days before expected estrus to one day after estrus. Plasma progesterone concentration at estrus was 0.1 ng/ml which rose to a peak level of 3.47 ng/ml on day 17. It fluctuated around this level in those animals which conceived, but followed a declining trend in those which failed to do so and attained lowest values on the day of next estrus. Temporal changes of the hormone revealed that the occurrence of major decline varjed between 16 and 62 h before estrus. The average concentration in milk was about three to four times higher than in plasma. The concentration of estradiol-17beta about 23.50 pg/ml at estrus and fluctuated around 10 pg/ml in animals that returned to estrus with a peak around estrus. Temporal changes of hormone revealed that peak level occurred 8-17 h before estrus. The concentration of estradiol in pregnant animals fluctuated around 10 pg/ml. The concentration in milk was about 2-3 times higher than in plasma. There was no significant (P > 0.05) difference in the concentrations of progesterone and estradiol-17beta between primiparous and multiparous animals.     

Immunoaffinity chromatography and a biotin-streptavidin amplified enzymeimmunoassay for sensitive and specific estimation of estradiol-17β

A sensitive test system has been developed for estimation ofestradiol-17β (E₂) in bovine plasma. Plasma extracts are first purified by a selective immunoaffinity chromatography (IAC) using an antibody raised against estradiol-6-carboxymethyloxime-bovine serum albumin and immobilized to Sepharose. The eluate was analysed by a competitive enzyme immunoassay (EIA) on microtitration plates. For the assay the wells of microtitration plates were coated with affinity purified sheep IgG (antirabbit IgG) that binds the hormone specific antibody raised in rabbits against estradiol-17-hemisuccinate-bovine serum albumin. E₂ is estimated by displacement of biocytinyl-E₂, that was produced by ligation of estradiol-17β, d-glucuronic acid and biocytin. Bound biocytinyl-E₂ is detected after binding of streptavidin-peroxidase and colour production by the enzyme. A very high amplification was possible with this technique and the absolute detection limit amounted to ≈120fg/well at 94% relative binding. By combination of IAC and EIA the following levels of E₂ were found in bovine plasma: male or female calves <2.7pg/ml, cycling cow 0.5–7 pg/ml, cow during last month of pregnancy 9–310 pg/ml, mature bull 5–30 pg/ml. However, up to 1110 pg E₂/ml were found in plasma of a calf after treatment with an illicit hormone preparation used for growth promotion; after 21 days levels declined to 6 pg/ml which is hardly different from controls. In conclusion, the IAC/EIA can be used for sentitive estimation ofestradiol-17β in plasma from all type of cattle and for control of improper use of E₂ after commitment of a threshhold level.     

Comparable testosterone responses are produced by constant infusion of LH or GnRH in normal men

Luteinizing hormone (LH) was infused continuously at a rate of 1.3 IU/min to 4 normal adult men. A 4 to 5-fold increase in serum LH was noted by 8 hours. Serum FSH declined steadily throughout the infusion period in the face of rising concentrations of gonadal steroids. Basal plasma testosterone of 4.7 +/- 0.4 ng/ml rose progressively to a peak of 11.1 +/- 0.9 ng/ml at hour 56 (p less than 0.005). A similar pattern was demonstrated by plasma androstenedione. Plasma 17 alpha-hydroxyprogesterone rose from a basal concentration of 0.81 +/- 0.14 ng/ml to a peak concentration of 2.6 +/- 0.3 ng/ml at hour 36 of the infusion and subsequently declined. A similar course was followed by serum estradiol-17 beta, which achieved a maximal concentration of 70.0 +/- 10.4 pg/ml at hour 36. Results are compared to those obtained with continuous infusion of GnRH in normal adult men. Testosterone responses were similar, whereas elevations in 17 alpha-hydroxyprogesterone and estradiol were higher following GnRH infusion. This difference may be consequent upon a direct gonadal effect of GnRH, or may be secondary to local regulation of testicular steroidogenesis by estradiol-17 beta.     

Sex hormones correlated with sex skin swelling and rectal temperature during the menstrual cycle of the pigtail macaque (Macaca nemestrina).

Daily measurement of serum luteinizing hormone, estradiol-17beta, and progesterone were made during the menstrual cycle in nine pigtail macaques (Macaca nemestrina). All data were normalized to the day of the luteinizing hormone peak. Serum estradiol-17beta increased from approximately 100 pg/ml during the early follicular phase to 442 +/- 156 pg/ml during the maximum midcycle concomitant with the luteinizing hormone peak, and a small increase in serum estradiol-17beta was observed during the luteal phase coincident with the progesterone peak. Serum progesterone values increased slightly at the time of the luteinizing hormone peak and increased from 0.2-0.3 ng/ml during the midfollicular phase to peak levels of 8.3 +/- 1.75 ng/ml 9 days after the luteinizing hormone surge. Serum luteinizing hormone remained low and relatively constant throughout the early and midcycle, then sharply increased approximately four-fold to peak values of 6.25 +/- 0.9 ng/ml. Sex skin swelling increased slowly during the follicular phase and declined slowly throughout the early luteal phase. Rectal temperature did not change significantly throughout the menstrual cycle. The similarity of plasma sex hormone changes during the menstrual cycle between women and the pigtail macaque suggested that this nonhuman primate should be a useful animal model for studying human reproduction.     

Evidence for estrogen-dependent blastocyst formation in the pig

The physiological significance of estradiol-17beta for the early embryonic development in the pig was investigated in vitro by four different experimental designs. A total of 1635 morphologically intact morulae were cultured in vitro in Krebs-Ringer bicarbonate solution supplemented with 10% heat-inactivated lamb serum, and the blastocyst formation rate (BFR) was recorded after 24 or 48 h. The addition of estradiol-17 beta (0.1 nM, 1 nM, 100 nM), progesterone (100 nM, 500 nM) or cortisol (100 nM) to the culture medium did not affect BFR (95 to 100%, Experiment 1). Similarly, adding charcoal-stripped lamb serum to the medium instead of normal lamb serum in the absence or presence of 1 nM estradiol-17 beta had no effect (93 to 95% BFR, Experiment 2). The antiestrogen Nafoxidine, however, at a concentration of 15 micrograms/ml, significantly (p less than 0.01) reduced BFR to 13.3 +/- 5.8% compared to controls (93.3 +/- 4.2%, Experiment 3). Supplementation with estradiol-17 beta (1 nM) in the presence of 15 micrograms/ml Nafoxidine significantly (p less than 0.01) improved BFR to 57.2 +/- 8.9%. Higher concentrations of estradiol-17 beta (100 nM, 100 microM) did not further enhance BFR. The stimulatory effects of estradiol-17 beta were specific since the BFR remained low in the presence of 100 nM progesterone (10.0 +/- 4.5%) or 100 nM cortisol (3.3 +/- 3.3%). Addition of 5% estradiol-17 beta-antiserum to the culture medium (Experiment 4) significantly (p less than 0.01) reduced BRF to 51.9 +/- 6.7% compared to controls (93.1 +/- 2.2%).(ABSTRACT TRUNCATED AT 250 WORDS)     

Radioimmunoassay of estradiol-17beta in bovine peripheral plasma with and without chromatography.

An assay of estradiol-17beta (E217beta) in bovine peripheral plasma is described. The plasma is incubated with an antiserum to E217beta-BSA and the gamma-globulin fraction precipitated with ammonium sulphate. After extraction with diethyl ether E217beta in the precipitate is estimated by radioimmunoassay using a specific antiserum against E217beta-6-BSA. Plasma concentrations of E217beta during the normal estrous cycle determined by this method and by a method involving Sephadex LH-20 chromatography range from 4 to 23 pg/ml.     

Seasonal variation in serum levels of steroid hormones and their relation with seminal quality and libido in buffalo bulls

Blood samples from 15 breeding male Murrah buffaloes were collected during the winter, summer and monsoon seasons. Seminal characteristics and sexual behaviour were also studied. Serum samples were analysed for testosterone, progesterone and estradiol-17beta levels by radioimmunoassay. The studies showed significantly lower values for testosterone during winter (0.53 +/- 0.06 ng/ml) than during summer (1.22 +/- 0.19 ng/ml) and monsoon (1.06 +/- 0.12 ng/ml). The progesterone level was lowest during monsoon (84 +/- 9 pg/ml), intermediate during winter (115 +/- 14 pg/ml) and highest during summer (224 +/- 24 pg/ml). The mean level of estradiol-17beta was almost double (9 +/- 0.7 pg/ml) during monsoon as compared to winter (5 +/- 0.1 pg/ml). The correlations between hormone levels, seminal characteristics and sexual behaviour were of low magnitude.     

Highly sensitive second-antibody enzyme immunoassay for determination of estradiol-17beta concentration in blood plasma of the mithun (Bos frontalis)

The objective of the present study was to develop and validate a simple, sensitive, quick and economic enzyme immunoassay (EIA) for estradiol-17beta (E2) in mithun (Bos frontalis) plasma on microtiter plates using a second-antibody coating technique and hormone-horseradish peroxidase as a label. For the assay, the wells of microtiter plates were coated with affinity-purified goat anti-rabbit IgG that binds the hormone-specific antibody. One milliliter of mithun plasma was extracted using benzene and 50 microl of 300 microl volume reconstituted with assay buffer was run in the assay along with standards ranging from 0.10-100 pg/well prepared in assay buffer. The sensitivity of the assay was 0.72 pg/ml. The intra- and inter-assay coefficients of variation were below 10%, and the extraction efficiency was >93%. Linearity of recovery of the added hormone concentrations was recorded. The assay developed was further validated biologically by estimating the hormone concentrations in six female and five male mithun calves, 12 cyclic mithuns for the entire reproductive cycle, and four pregnant mithun cows. The EIA developed can estimate low concentrations of E2 (2.2-5.2 pg/ml) in growing calves as well as very high concentrations of the hormone during pregnancy (E2=85.6-143.5 pg/ml). Apart from being non-radioactive, the assay developed has several advantages over conventional radioimmunoassays: it is more sensitive, less labor intensive, simpler to perform, and less time consuming. In conclusion, the EIA procedure described herein is sufficiently reliable, economic, safe, quick and sensitive to estimate the hormone at all physiological levels in bovine plasma.     

Changes in circulating inhibin levels during pregnancy and early lactation in the Japanese monkey

Changes in immunoreactive (ir-) inhibin concentrations in serum throughout pregnancy and early lactation up to one month after parturition were characterized in 6 Japanese monkeys (Macaca fuscata fuscata) by a heterologous radioimmunoassay (RIA) based on a bovine RIA. Serum levels of FSH, LH/monkey chorionic gonadotropin (mCG), estradiol-17 beta, and progesterone were also monitored for the entire period. Ir-inhibin levels in the serum were low (under 0.5 ng/ml) before conception. Three marked increases in serum ir-inhibin levels were found during pregnancy. The first increase was noted during early pregnancy, with a peak (2.2 +/- 0.2 ng/ml) at Day 22 of pregnancy (Day 0 = day of LH surge). The second increase was noted after Day 38 until Day 72 of pregnancy, when a peak value was noted (19.0 +/- 1.4 pg/ml). Plateau levels were maintained until late pregnancy, and a final rise was evident near the term with a peak (36.7 +/- 3.8 ng/ml) at Day 158 of pregnancy, 5 days before parturition. After parturition, ir-inhibin levels in the serum plummeted to nonpregnant levels within one day, and were maintained during early lactation. The first rise in serum inhibin during pregnancy was parallel to the rise of mCG and estradiol-17 beta, and the second and third rise were well correlated with serum estradiol-17 beta. Serum FSH was maintained at low levels throughout pregnancy, followed by a slight increase after parturition when serum inhibin decreased abruptly. Both bioactivity and immunoreactivity of inhibin were detected in the placental homogenates obtained at 120 days of pregnancy.(ABSTRACT TRUNCATED AT 250 WORDS)     

Variation of steroid concentrations during the reproductive cycle of the clam Ruditapes decussatus: a one year study in the gulf of Gabès area

Progesterone, testosterone and estradiol-17beta were measured by radio-immunoassay (RIA) in the gonads of the clam Ruditapes decussatus. The reproductive cycle was also investigated. Our study covered a period of one year, from September 2003 to August 2004. The chosen site "Kerkennah", located out of industrial effluents, belongs to the gulf of Gabès area (Tunisia). Steroids varied from 178 to 2459 pg g(-1) wet mass for progesterone, from 40 to 326 pg g(-1) wet mass for testosterone and from 10 to 235 pg g(-1) wet mass for estradiol-17beta in females. However in males, these steroids ranged from 304 to 2303 pg g(-1) wet mass for progesterone, from 81 to 381 pg g(-1) wet mass for testosterone and from 48 to 168 pg g(-1) wet mass for estradiol-17beta. The reproductive cycle of R. decussatus, investigated by histological examination of gonadic sections, showed that gametogenesis occurred from April to February in males and from April to November in females. Progesterone and testosterone increased at the end of gametogenesis in both sexes. The highest estradiol-17beta was recorded at the beginning of vitellogenesis in females. Fluctuations in the levels of sex steroids during the reproductive cycle suggest their possible role as endogenous modulators of gametogenesis in R. decussatus. Although this species is considered as gonochoristic, 0.83% of hermaphrodites were observed.     

Development and validation of chemiluminescent immunoassay for vitellogenin in five salmonid species.

A highly sensitive and specific chemiluminescent immunoassay (CLIA) was developed for quantification of vitellogenin (Vg) in five salmonids. The CLIA for salmon Vg was performed using the two-site method, with anti-masu salmon beta'-component as primary antibody and chemiluminescent acridinium-labeled anti-rainbow trout lipovitellin F(ab)'(2) as the second antibody. Using cutthroat trout Vg as the standard, the working range of the CLIA was from 60 pg to 500 ng Vg/ml. Intra- and inter-assay coefficients of variation ranged from 3.04 to 6.67% and 3.23 to 5.86%, respectively. For the various salmonid species, serially diluted samples of serum from vitellogenic fish ran parallel to their purified Vg standard curve in the CLIA. In male cutthroat trout maturing during the 4 months before spawning, serum Vg levels ranged from 1.56 to 8000 ng/ml. High levels of Vg in some individuals may have resulted from temporary elevation of estradiol-17beta levels in the same fish during December or January (1-2 months before spawning). This is the first report on changes in serum Vg levels in maturing male trout using CLIA, the most sensitive assay for Vg yet developed.     

Binding of 2-hydroxyestradiol and 4-hydroxyestradiol to estrogen receptors from human breast cancers

The binding of catechol estrogens, epoxyenones and methoxyestrogens was evaluated using estrogen receptors in cytosol prepared from human breast cancers. The relative affinity of 2-hydroxyestradiol, a metabolite formed in vitro from estradiol-17 beta by breast cancer cells, was indistinguishable from that of estradiol-17 beta. 4-Hydroxyestradiol, which is also a metabolite of estradiol-17 beta, associated with the estrogen receptor with a relative affinity approximately 1.5-fold greater than that of estradiol-17 beta. Epoxyenones and methoxyestrogens were weak competitors compared to the binding of estradiol-17 beta, exhibiting relative affinities 3% or less than the affinity of estradiol-17 beta. Sucrose density gradient centrifugation revealed that both 2- and 4-hydroxyestradiol inhibited the binding of estradiol-17 beta to both the 4S and 8S isoforms of the estrogen receptor in a competitive manner, with a Ki = 0.94 nM for 2-hydroxyestradiol and a Ki = 0.48 nM for 4-hydroxyestradiol. It can be concluded that these data demonstrate a specific receptor-mediated estrogenic action for both of these catechol estrogens.     

Relationship between seasonal plasma estradiol-17 beta and testosterone levels and in vitro production by ovarian follicles of amago salmon (Oncorhynchus rhodurus)

Plasma estradiol-17 beta and testosterone levels were assessed by radioimmunoassay during the sexual maturation of female amago salmon (Oncorhynchus rhodurus). Estradiol-17 beta levels gradually increased during vitellogenesis (June to September), reached a peak in September (about 16 ng/ml) and rapidly decreased in mature and ovulated fish (about 3-4 ng/ml) in October. The seasonal pattern of plasma testosterone levels lagged behind and followed that of estradiol-17 beta during vitellogenesis, but levels remained high in mature and ovulated fish (90-110 ng/ml). Estradiol-17 beta levels and the gonadosomatic index (GSI) values correlated well during vitellogenesis: GSI values showed a linear increase, and reached a peak (29.9 +/- 1.4) in October. Values were extremely low in ovulated fish (1.2 +/- 0.2). In vitro production of estradiol-17 beta and testosterone by ovarian follicles in response to partially purified chinook salmon gonadotropin (SG-G100) was examined monthly using 18-h incubations. Throughout the vitellogenic period SG-G100 stimulated both estradiol-17 beta and testosterone production: the steroidogenic response of follicles increased from June (about 2 ng/ml estradiol-17 beta; 0.1 ng/ml testosterone) to September (about 10 and 14 ng/ml, respectively). In October full-grown immature follicles which could be induced to mature in vitro by hormone treatment produced large amounts of testosterone (about 130 ng/ml) but not estradiol-17 beta. Postovulatory follicles also produced testosterone but the values were low (10 ng/ml) compared with full-grown immature follicles. Very low levels of estradiol-17 beta were produced by postovulatory follicles.(ABSTRACT TRUNCATED AT 250 WORDS)     

Serum and urinary estrone sulfate during the menstrual cycle, measured by a direct radioimmunoassay, and fate of exogenously injected estrone sulfate

Serum and early-morning urinary levels of estrone sulfate during the menstrual cycle were measured by a direct radioimmunoassay without hydrolysis. These levels were high and showed prominent peaks [serum, 2.67 +/- 0.37 ng/ml (mean +/- SE); urine, 5.82 +/- 2.3 micrograms/l] around the day of the preovulatory estradiol-17 beta peak, and increased again during the luteal phase. Following intravenous injection of estrone sulfate, serum estrone sulfate, estrone and estradiol-17 beta were measured. The conversion of estrone sulfate to estrone and/or estradiol-17 beta was very small during their transit in the general circulation.