Steroid hormone formation in human ovarian follicles in vitro.

Ovarian follicles of 5 to 15 mm in diameter were isolated from 45 ovaries of 34 patients in the follicular and luteal phases of the cycle. Three experiments were done. In the first, follicles were minced and incubated in Krebs-Ringer bicarbonate buffer containing 1 to 2muCi of testosterone-4-14C in the presence or absence of 100 IU human chorionic gonadotropan (hCG). In the second, minced follicles were incubated with 100 muCi of sodium acetate-I-14C under identical conditions. In the third, ten follicles from a single patient in the late proliferative stage of endometrial dating were cut in halves and incubated with 100 muCi of acetate-I-14C under identical conditions. The minced follicle preparation was capable of aromatizing testosterone-4-14C into radioactive estrone and estradiol in significant amounts. Incorporation of radioactive acetate into pregenolone, progesterone, 17-hydroxypregnenolone, 17-hydroxyprogesterone, dehydroepiandrosterone, androstenedione, testosterone, estradiol and estrone was assessed by reverse dilution analysis with recrystallization to constant specific activity. The major radioactive products formed were androstenedione and 17-hydroxyprogesterone in the latter two experiments. Dehydroepiandrosterone was one of the major steroids in the second experiment. The minor products were testosterone, progesterone and pregnenolone. Smaller, but definite incorporations of radioactive acetate into estradiol and estrone occurred in the second experiment. On histological examination, the follicles were characterized by atretic changes. This distribution pattern of radioactive acetate among the steroids was considered to represent the steroidogenic profile of unstimulated or atretic follicles.     

Steroid levels after intramuscular injection of radioactive estradiol-17beta, estrone, progesterone and testosterone in the bovine

Eight 2 year old Hereford cows from days 8 to 12 of the estrous cycle were injected intramuscularly with 5 ml of corn oil containing 5 mg of estradiol-17beta (two cows), estrone (two cows), progesterone (two cows) or testosterone (two cows). Each cow treated with estradiol received 494 microc of estradiol-17beta-6, 7 H3 and each cow treated with estrone received 492 microc of estrone-6, 7 H3. Each cow treated with progesterone or testosterone received 400 muc of H3 compound labeled in the 7 position. Total urine was collected by urethral catheterization of the cows treated with estrogens. Blood samples for plasma and serum were collected via jugular cannulae. Blood and urine samples from estrogen-treated cows were collected hourly for the first 24 hr, at 2 hr intervals for the next 26 hr, at 4 hr intervals for the next 12 hr and at 12 hr intervals until background was reached. Blood samples were collected hourly from 1 to 8 hr after injection from progesterone or testosterone-treated cows. Plasma and serum levels of radioactive estradiol-17beta, estrone, progesterone and testosterone were similar. Blood levels of radioactivity peaked at 2 hr post-injection in cows receiving estradiol-17beta and at 3 hr in cows receiving estrone. Blood levels of labeled estradiol-17beta and estrone were nondetectable by 54 hr and 83 hr, respectively. Peak urinary excretion of radioactivity was reached at 7 hr for estradiol-17beta and at 14 hr for estrone and nondetectable levels were reached by 95 hr for estradiol-17beta and 14 hr for estrone. At these times, 15.5% of the total dose of radioactive estradiol-17beta and 17.5% of the injected estrone had been excreted in the urine. Peak blood and urinary excretion levels were reached earlier for radioactive estradiol-17beta than for estrone, and excretion of estradiol-17beta was completed more rapidly. No difference was found in plasma and serum levels for any steroid studies; thus, endogenous steroid titers in blood plasma and serum are not different in the cow.     

Steroid metabolism in human teratocarcinoma cell line PA 1

Human ovarian teratocarcinoma cells of line PA 1, (Zeuthen et al., 1979[1]) used as model for early embryonic cells, were analyzed for their in vitro capacity to convert steroids. The cells were incubated for 20 h with radioactive pregnenolone, progesterone, dehydroepiandrosterone, androstenedione, testosterone or estradiol-17 beta, or with non-radioactive progesterone, 6 alpha- or 6 beta-hydroxyprogesterone, 3 beta-hydroxy-5 alpha-pregnan-20-one, dehydroepiandrosterone or estradiol-17 beta. The metabolites were analyzed by thin layer chromatography or studied by gas chromatography-mass spectrometry. The results indicate that PA 1 cells are able to metabolize, although to a restricted amount, a variety of steroids, most markedly progesterone. The metabolites were almost exclusively found in the medium. The main metabolite of progesterone was 3 beta, 6 alpha-dihydroxy-5 alpha-pregnan-20-one. Minor formation of progesterone from pregnenolone could be detected. Human chorionic gonadotropin did not have any effect on pregnenolone metabolism. No formation of estradiol-17 beta or estrone from dehydroepiandrosterone, androstenedione or testosterone could be detected. However, estradiol-17 beta was shown to be converted mainly to estrone. These findings indicate that undifferentiated PA 1 teratocarcinoma cells like certain mouse teratocarcinoma cells, seem not to be steroidogenic but are capable of metabolizing naturally occurring steroid hormones and their precursors.     

Enzyme induction in Streptomyces hydrogenans. V. Characterization of testosterone-17 beta-dehydrogenase and its induction by steroids]

Testosterone 17beta-dehydrogenase can be enriched from Streptomyces hydrogenans. The enzyme dehydrogenizes testosterone with Km=13muM and estradiol-17beta with Km=21muM to the corresponding 17-ketoderivatives. NAD forms NADH with Km=125muM. The enzyme is strongly inhibited by androstandione and 17alpha-methyltestosterone. The Ki for 17alpha-methyltestosterone is 18muM. The enzyme activity increases with increasing pH up to alkali-mediated denaturation at about pH 10. The optimum temperature is at 45 degrees C. If Streptomyces hydrogenans is cultivated in the absence of steroids, the specific activity of testosterone 17beta-dehydrogenase in the cytosol of the microorganisms amounts to 10 mU/mg protein, and increases up to 10-fold if the cells are cultivated in the presence of certain steroids. Testosterone, alpha-dihydrotestosterone, beta-dihydrotestosterone, estradiol-17beta, and 17alpha-methyltestosterone are very effective inducers. Thus, for the first time, the ability of estradiol-17beta to induce an enzyme synthesis in a microorganism is shown. The steroid-dependent induction is inhibited by testosterone acetate and rifamycin SV. Cyproterone, however, does not decrease the testosterone-dependent enzyme induction of testosterone 17beta-dehydrogenase.     

Androgen metabolites in bovine follicular fluid

The values of C21-steroids, Delta4-androgens, estrogens as well as 5alpha-reduced steroids have been determined in follicular fluid obtained from superovulated and untreated cows. In the three cows treated with a hormone regimen to induce superovulation, the levels of progesterone and estradiol determined in 3 to 6 follicles per cow ranged from 65 to 448 ng/ml and 1.9 to 8.6 ng/ml, respectively while the concentrations of androstenedione and testosterone varied between 1.5 to 2.5 ng/ml. Low levels of dihydrotestosterone and androstane-3alpha, 17beta-diol (approximately 30 to 50% of Delta4-androgens) were found in the bovine follicular fluid. In untreated cows, the follicular steroid concentrations were divided into two groups on the basis of the ratio between estrogen and Delta4-androgen concentrations. In estrogen-rich follicles, the ratio of estrogens Delta4- androgens was higher than 1 and in estrogen-poor follicle, the ratio of estrogens Delta4- androgens was lower than 1. Pregnenolone, dehydroepiandrosterone, androst-5-ene-3beta, 17beta-diol, progesterone, androstenedione and testosterone levels were not significantly different in the two groups while the levels of estradiol and estrone were approximately 100-fold higher in the estrogen-rich group. The concentrations of 5alpha-reduced steroids particularly, dihydrotestosterone, androstane-3alpha, 17beta-diol and androsterone as well as their glucuronides which were found at values extremely low (under 1 ng/ml) were not significantly different in both groups. The results indicate that low levels of 5alpha-reduced steroids and their glucuronides are present in bovine follicular fluid and their concentrations remained fairly stable either in estrogen-rich or estrogen-poor groups.     

The determination of five steroids in avian plasma by radioimmunoassay and competitive protein-binding.

A method has been developed for the simultaneous determination of testosterone, 5alpha-dihydrotestosterone and corticosterone, or of estrone, estradiol-17beta and corticosterone, after separation on a Celite:propylene glycol:ethylene glycol column (6:1.5:1.5 w/v/v). The lower quarter of the column was packed with a Celite: water mixture (3:1 w/v) as a stationary phase (glycol) 'trap'. This effectively prevented leaching of the glycols into the eluate as the concentration of ethyl acetate in the mobile phase was increased to elute the more polar steroids. In addition, a second system utilizing a Celite: ethylene glycol column (2:1 w/v) for the separation of estrone and estradiol-17beta is described. Testosterone, 5alpha-dihydrotestosterone, estrone and estradiol-17beta were measured by radioimmunoassay and corticosterone by a competitive protein-binding technique. Reliability criteria are presented showing that the assay systems used are accurate and reproducible. Plasma-steroid levels of eight avian species are also presented and compared with those found by other investigators.     

Identifications of radioactive steroid estrogen conjugates in blood plasma of laying hens after intramuscular injection of (4--14C)-estrone.

[4--14C] Estrone was injected intramuscularly into six laying hens. Fifty minutes later the hens were exsanguinated. The plasmas were examined for conjugates of radioactive phenolic steroids by recovery on columns of Amberlite XAD-2 or by extraction with tetrahydrofuran followed by chromatography on a column of DEAE-Sephadex A-25 in a gradient of NaCl. The biggest Sephadex chromatographic fraction (50,4% of total) contained about 42% of its radioactivity as estradiol-17alpha-3-sulfate and 18% as estradiol-17beta-3-sulfate and the remaining 40% was identified tentatively as estradiol-17alpha-17-sulfate plus a small proportion of estradiol-17beta-17-sulfate. The second biggest Sephadex chromatographic fraction (12.7% of total) was a mixture of conjugates not further identified. Minor fractions identified comprised estrone-beta-glucuronide (2.8%), estradiol-17alpha-3-beta-glucuronide (2.8%), estradiol-17beta-3-beta-glucuronide (2.3%) and estrone sulfate (6.0%). Evidence was obtained for the presence of small proportions of estradiol-17alpha disulfate and estradiol-17beta disulfate.     

Conversion of steroids in bovine blood in vitro

Blood samples collected from eight Braunvieh cows between the sixth and eighth month of gestation were allowed to stand with and without anticoagulant at 20 degrees C and 0 degrees C for different time periods. In these samples the degree of in vitro conversion of gestagens, androgens and estrogens was investigated. The concentrations were measured by radioimmunoassay. After 24 h at 20 degrees C, the levels of pregnenolone, progesterone, 17alpha-hydroxyprogesterone, androstenedione, dehydroepiandrosterone and estrone decreased to 62, 29, 25, 10, 34 and 44%, respectively, of the initial value and those of 17alpha, 20beta-dihydroxyprogesterone, epitestosterone and estradiol-17alpha increased to 385, 800 and 852%, respectively. The conversion was slower in clotted blood. The concentrations of testosterone and estradiol-17beta were consistent over the 24 h period. There was no marked decrease of the steroid concentration after 24 h of incubation of whole blood at 0 degrees C and of plasma at 20 degrees C. After the addition of (3)H-steroids, conversion could be demonstrated by thin-layer chromatography and autoradiography. These results demonstrate that all investigated hormones except testosterone and estradiol-17beta were metabolized by bovine blood cells.     

Steroids and follicular rupture at ovulation

The preovulatory surge of gonadotropins stimulates follicular steroidogenesis and changes from estrogen as the major product to progesterone. We shall overview the studies dealing with the role of ovarian steroidogenesis in follicular rupture at ovulation. Several inhibitors of steroidogenesis blocked follicular rupture in vivo. Likewise, RU 38486 partially blocked ovulation triggered by hCG. Collectively, these data support the knowledge that follicular steroidogenesis is required for ovulation. Recent studies confirmed the essential role of plasminogen activator (PA) in follicular rupture. The LH stimulation of PA activity was partially blocked by several inhibitors of steroidogenesis and it could be restored by the addition of progesterone, testosterone and estradiol-17 beta, but not the non-aromatizable 5 alpha-dihydrotestosterone. Gonadotropic stimulation enhanced only the synthesis of tissue type PA (t-PA) and not that of urokinase. Likewise, inhibition of steroidogenesis, reduced only the synthesis of t-PA and was reversed by addition of estradiol-17 beta. It seems, therefore, that follicular steroids, most probably estrogen, are involved in the preovulatory rise in follicular t-PA activity.     

In vitro metabolism of testosterone by cultured Sertoli cells and the effect of FSH.

Cultures of Sertoli cells isolated from testes of 18-and 36-day-old Long Evans rats were used to investigate their capacity to metabolize testosterone and the effect of FSH on such metabolism. Three different approaches were used: 1) investigation of the metabolism of radiolabeled testosterone under saturating substrate conditions; 2) study of the metabolism of radiolabeled testosterone utilizing trace amounts of high specific activity substrates; 3) the utilization of radioimmunoassay for measurement of estradiol-17 beta. The following steroids were isolated and identified by recrystallization to constant specific acitvity from the control and FSH-treated cultures; testosterone (unconverted substrate), androstenedione, dihydrotestosterone, 3 alpha-hydroxy-5 alpha-androstan-17-one and 5 alpha-androstane-3 alpha, 17 beta-diol. Radioimmunoassay data suggests that the Sertoli cells produce an estradiol-17 beta-like compound from unlabeled testosterone and that this production is stimulated by FSH. However, the radioactive metabolite from all our studies that behaved chromatographically like estradiol--17 beta failed to crystallize to constant specific activity, while in each experiment, authentic radiolabeled estradiol-17 beta added as recovery tracer did. The data demonstrate that : 1) cultures of Sertoli cells from immature rats have 5 alpha-reductase, 3 alpha- and 17 beta-hydroxysteroid oxidoreductase activities; 2) these enzymes may be affected by FSH; 3) based on radiolabeled metabolic techniques, Sertoli cells were unable to biotransform testosterone to estradiol-17 beta even in the presence of FSH.     

Intrafollicular steroids and anti-mullerian hormone during normal and cystic ovarian follicular development in the cow

Development of follicular cysts is a frequent ovarian dysfunction in cattle. Functional changes that precede cyst formation are unknown, but a role for anti-Müllerian hormone (AMH) in the development of follicular cysts has been suggested in humans. This study aimed to characterize intrafollicular steroids and AMH during follicular growth in a strain of beef cows exhibiting a high incidence of occurrence of follicular cysts. Normal follicular growth and cyst development were assessed by ovarian ultrasonography scanning during the 8 days before slaughtering. Experimental regression of cysts was followed by rapid growth of follicles that reached the size of cysts within 3-5 days. These young cysts exhibited higher intrafollicular concentrations of testosterone, estradiol-17beta, and progesterone than large early dominant follicles did in normal ovaries, but they exhibited similar concentrations of AMH. Later-stage cysts were characterized by hypertrophy of theca interna cells, high intrafollicular progesterone concentration, and high steroidogenic acute regulatory protein mRNA expression in granulosa cells. Progesterone and AMH concentrations in the largest follicles (> or =10 mm) and cysts were negatively correlated (r = -0.45, P < 0.01). Smaller follicles (<10 mm) exhibited higher intrafollicular testosterone and estradiol-17beta concentrations in ovaries with cysts compared to normal ovaries. During follicular growth, AMH concentration dropped in follicles larger than 5 mm in diameter and in a similar way in ovaries with and without cysts. In conclusion, enhanced growth and steroidogenesis in antral follicles <10 mm preceded cyst formation in cow ovaries. Intrafollicular AMH was not a marker of cystic development in the cow, but low AMH concentrations in cysts were associated with luteinization.     

Estradiol-17 beta inhibition of androgen uptake, metabolism and binding in epididymis of adult male rats in vivo: a comparison with cyproterone acetate

The effects of estradiol-17 beta on androgen uptake, metabolism and binding were studied in rat epididymis in vivo in comparison with cyproterone acetate. Steroids (250 ug/100 g body weight) were injected 5 min prior to 3H-testosterone in castrate rats. Estradiol-17 beta inhibited 3H-testosterone uptake into epididymal cytosol by 58% as compared to 38% by cyproterone acetate. 3H-Testosterone uptake into epididymal nuclei was inhibited 95% by estradiol-17 beta and 83% by cyproterone acetate. Total bound radioactivity in cytosol fractions was reduced to a greater extent by estradiol-17 beta than cyproterone acetate when either 3H-testosterone or 3H-dihydrotestosterone was injected. Binding of 3H-dihydrotestosterone to nuclear receptors was completely abolished by estradiol-17 beta; whereas approximately 20% binding remained in the nuclear extract after cyproterone acetate treatment. Metabolism of 3H-testosterone in vivo was also altered by estradiol-17 beta, resulting in diminished conversion to 3H-dihydrotestosterone. Cyproterone acetate, on the other hand, did not affect 3H-testosterone metabolism. Estradiol-17 beta and cyproterone acetate inhibited in vitro binding of 3H-dihydrotestosterone to the intracellular cytoplasmic receptor, but not the intraluminal androgen binding protein (ABP). These data suggest that estradiol-17 beta may have a more potent antiandrogenic effect on the epididymis than cyproterone acetate due to inhibition of 5 alpha reduction of testosterone as well as binding to the androgen receptor.     

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)     

In vitro synthesis of estrogen glucuronides and sulfates by human renal tissue.

17beta-[6,7-3H]Estradiol (E2) was incubated with slices and homogenates of adult human renal tissue. The metabolites formed were identified by chromatography on DEAE-Sephadex, thin layer chromatography and crystallization with carrier steroids or steroid derivatives. The major metabolites formed by slices were estradiol-17-glucuronide (E217G), estrone sulfate and estradiol-3-sulfate. This is the first report of in vitro synthesis of estrogen sulfates by adult renal tissue. Minor quantities of the 3-glucuronides of estrone and estradiol were also found. An oxygen atmosphere appeared to stimulate the production of E217G. A time study with tissue slices showed similarities between the in vitro pattern of glucuronide synthesis and the excretion pattern of these compounds seen in earlier in vivo studies. Homogenates fortified with uridine diphosphoglucuronic acid formed the same pattern of glucuronide products but in lesser amounts. No sulfates were formed under these conditions. Testosterone did not act as a substrate in the experimental conditions used.     

Identification of some plasma metabolites of 6, 7-3-H-estrone glucosiduronate in male dogs.

Following the constant infusion of 6, 7-3-H-estrone glucosiduronate in male dogs for a period of 120 minutes, the radioactive metabolites present in the plasma were separated by solvent partition, DEAE-Sephapadex, Celite partition and thin layer chromatography. The identities of the individual estrogens and estrogen conjugates were confirmed by specific activity determinations after chromatography in several different solvent systems, enzyme hydrolysis and steroids and their derivatives. Most of the radioactivity in the plasma was identified as estrone glucosiduronate. The major metabolite present was estradiol-17-beta-3-glucosiduronate. Small amounts of estradiol-17-alpha-3-glucosiduronate and free estrone were also identified. Three other minor conjugates were separated, but positive identification could not be made.     

Involvement of steroid hormones on in vitro maturation of pig oocytes

The purpose of this study was to determine if the addition of steroid hormones into the culture medium could influence the in vitro maturation of pig oocytes. The cumulus-oocyte complexes (COCs). collected from follicles of 2-5 mm diameter, were matured in steroid-free medium supplemented with various concentrations of estradiol-17beta (0-3000 ng/ml), progesterone (0-5000 ng/ml) and testosterone (0-300 ng/ml). The COCs were cultured for 42 h, then fertilized in vitro. We analyzed nuclear and cytoplasmic maturation with lacmoid stain 20 h after in vitro insemination. We observed no significant effect (P > 0.05) on the percentage of oocytes completing nuclear or cytoplasmic maturation or the number of sperm penetrating each oocyte for any concentration of progesterone, estradiol-17beta or testosterone. Similarly, adding a combination of those hormones to the medium did not significantly (P > 0.05) affect any of the criteria. In order to determine if there was a possible secretion of steroids during maturation, we added COCs, denuded oocytes and stripped cumulus cells to drops of a steroid-free medium and cultured them for 42 h, after which we analyzed the medium, before and after culture, for the presence of progesterone, estradiol-17beta and testosterone by radioimmunoassay (RIA) analysis. COCs, as well as cumulus cells alone, secreted similar amounts of estradiol (43.3 and 37.5 pg/ml, respectively) and progesterone (4.24 and 4.79 ng/ml, respectively) into the maturation medium. A small amount of estradiol (28.8 pg/ml) was also detected when oocytes were cultured alone. These results indicate that no steroids need to be added to the maturation medium of pig oocytes and that the COCs secrete steroids during maturation. It is possible that the amounts produced by the COCs fulfill any requirement for steroids if these steroids are required for either nuclear or cytoplasmic oocyte maturation.     

Synthesis and aromatization of 19-norandrogens in the stallion testis

The results of the measurement of 19-nortestosterone in the testiscular artery and vein of the stallion, the very low levels of this steroid in the peripheral blood of geldings and the similar patterns of increase in the peripheral levels of 19-nortestosterone and testosterone after hCG stimulation, show that 19-nortestosterone, like testosterone, is essentially synthesized in the testis. This testicular origin was confirmed by the ability of testicular tissue to synthesize 19-norandrogens from [4-14C]androgens in vitro. 19-Nortestosterone was 50% conjugated in the peripheral blood and almost entirely conjugated after biosynthesis in vitro. The sequence of appearance of steroids in the peripheral blood after a single injection of 10,000 IU hCG suggests that, in the equine testis, 19-norandrogens are produced by a specific C10-19 desmolase (estrene synthetase), stimulable by hCG. 19-Nortestosterone was aromatized into estradiol-17 beta by stallion testicular microsomes. The affinity of the aromatase for 19-nortestosterone was very low compared to that for testosterone. At low and presumably physiological levels, and at a high testosterone/19-nortestosterone ratio, testosterone did not inhibit 19-nortestosterone aromatization by more than 53%. Thus, 19-nortestosterone may be aromatized in vivo in the testis in spite of the endogenous concentrations of androgens. However, the low velocity of 19-nortestosterone aromatization by testicular microsomes at roughly physiological concentrations suggests that 19-norandrogen aromatization may only participate slightly in the testicular estrogen production. These results suggest that in the equine testis, two aromatizing enzyme systems may exist: one which aromatizes both androgens and 19-norandrogens, and a minority system more specific for 19-norandrogens.     

In vitro metabolism of estrone 2,4,6,7-3H and 4-androstene-3,17-dione-1,2-3H in submandibular gland and submandibular gland cancer tumor

Tritiated estrone and 4-androstene-3, 17-dione were incubated with slices of human submandibular gland and submandibular gland cancer tumor. Metabolites were separated by paper and thin layer chromatography. Estradiol-17β and testosterone were identified as metabolites. The metabolites were identified by reverse isotope dilution and the preparation of derivatives (formation of acetates). The amounts of estradiol-17β and testosterone formed in the tumor were significantly higher than that formed in the submandibular gland.     

Validation of radioimmunoassay systems for the measurement of 11-keto- and 11 beta-hydroxytestosterone in teleost blood

Highly specific antisera for 11-keto- and 11 beta-hydroxytestosterone have been raised in sheep. Assay systems for the simultaneous measurement of 11-ketotestosterone, 11 beta-hydroxytestosterone, testosterone, progesterone and estradiol-17 beta were validated for Ictalurus nebulosus plasma and Carassius auratus serum. In males of both species 11-ketotestosterone and testosterone were the major steroids detected. In females, testosterone and estradiol-17 beta were the predominant steroids measured. Data from samples taken at different stages of the annual cycle suggest that seasonal fluctuations in gonadal steroid secretion occur in I. nebulosus and C. auratus.