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.     

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.     

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.     

Oxytocin-stimulated production of prostaglandin F2alpha by isolated endometrium of rabbit: modulation by ovarian steroids

To test the hypothesis that ovarian steroid hormones modulate oxytocin-induced release of prostaglandin F2alpha (PGF2alpha) from uterine endometrium, 2 ovariectomized rabbits were pretreated with progesterone (5 mg/day for 10 days), 2 with estradiol-17 beta (25 microgram/day for 10 days), 2 with both steroids, and one with sesame oil only. On the last day of treatment, endometrial fragments were excised and incubated in vitro with or without oxytocin (100 muU/ml). Although endometrium from rabbits pretreated with combined steroids released more PGF2alpha immediately after excision than did tissue from animals pretreated with either steroid by itself, endometrium from animals pretreated with estradiol-17 beta alone released the most PGF2alpha during sustained incubation in vitro. Moreover, only this tissue exhibited significant oxytocin-dependent release of PGF2alpha. At the dosages used, progesterone completely antagonized both of these effects of estradiol-17 beta. The results support the hypothesis that ovarian steroid hormones regulate oxytocin-dependent release of PGF2alpha from endometrial cells. A posible mechanism of action is suggested.     

The effect of some sex hormones on the isovolumetric pressure curve of the rabbit's left ventricle

The effect of some sex hormones (estradiol-17α), estradiol- 17ß, estrone, progesterone and testosterone) on the mechanical activity of rabbit left ventricle was studied in concentrations within the physiological range. Investigations were carried out during the first ten minutes after the introduction of the steroid. Only progesterone and estradiol-17ß affect the isovolumetric pressure curve; the other steroids do not exhibit any effect. The effects of estradiol-17ß and progesterone counterbalance each other. Estradiol-17ß enhances the rate of pressure development and decreases the time to reach peak pressure. Progesterone decreases both the rate of pressure development and the time to reach peak pressure.     

Intratesticular unconjugated steroids in elderly men

As an extension of our studies on the influence of age on testicular function and with the aim of detecting whether the decline in testosterone production by aged testes is accompanied by a block in the biosynthetic chain leading from cholesterol to testosterone, we determined in the testis of young and elderly men, who died suddenly either from a cardiac incident or from accident, intratesticular steroids: pregnenolone, 17 hydroxypregnenolone (3 beta, 17 alpha-dihydroxy-5-pregnen-20-one), dehydroepiandrosterone, androstenediol, (5-androsten-3 beta, 17 beta-diol), progesterone, 17 hydroxyprogesterone, androstenedione, 17 beta-estradiol as well as testosterone, dihydrotestosterone (5 alpha-androstan-17 beta-ol-3-one) and androstanediol (5 alpha androstane-3 alpha, 17 beta-diol). The intratesticular steroid pattern in elderly men was essentially characterized by a decrease of the 5-ene steroid concentration, whereas we did not observe a decrease in the 4-ene steroids, progesterone concentration being even significantly higher in the aged testes. There was no evidence for a decrease in either lyase or 17-hydroxylase activity. It is suggested that the steroid pattern as observed in the aged testes is the consequence of a decreased oxygen supply, due to a decreased testicular perfusion.     

Effects of sex steroid hormones on differentiation of adipose precursor cells in primary culture

The effects of estradiol-17 beta and progesterone on multiplication, differentiation and lipid filling of adipose precursor cells were examined in primary cell cultures of cells prepared from adipose tissue of both male and ovariectomized female rats. Progesterone down to a concentration of 10(-7) mol/liter, alone or in the presence of estradiol-17 beta stimulated the development of glycerophosphate dehydrogenase and lipoprotein lipase activity. Estradiol-17 beta alone had no effects. These effects were essentially parallel to increases in the rate of lipid filling of the cells. Furthermore, the formation of cells with a lipid vacuole greater than 20 micron was markedly stimulated, suggesting that new fat cells were formed by the stimulation of differentiation of the adipose precursor cells. No effects of the sex steroid hormones were seen on the rate of multiplication. These results suggest a role of sex steroid hormones in the regulation of triglyceride storage capacity in adipose tissue by facilitating the differentiation of precursor cells to form new adipocytes.     

Intrafollicular concentrations of steroids and steroidogenic enzymes in relation to follicular development in the mare

The objective of the present study was to determine the changes in follicular fluid steroid concentrations and in granulosa cell steroidogenic enzyme expression during the follicular phase, in relation to follicular size and physiological status in the mare. Follicular fluid and follicular cells were recovered by ultrasound-guided follicular punctures either around the time of emergence of the dominant follicle, at the end of the dominant follicle growth, or at the preovulatory stage, after injection of gonadotropin to induce ovulation. Cellular relative amounts of steroidogenic acute regulatory protein (StAR), P450-side chain cleavage (P450(scc)), 3beta-hydroxysteroid dehydrogenase (3betaHSD), 17alpha-hydroxylase, and aromatase were assessed by semiquantitative Western blot and densitometry. Follicular fluid was assayed for cholesterol concentrations by colorimetric assay and for progesterone, testosterone, and estradiol-17beta concentrations by RIA. Intrafollicular concentrations of progesterone and estradiol-17beta significantly increased in the dominant follicle during growth. After injection of gonadotropin, follicular maturation was characterized by a decrease in estradiol-17beta concentrations and a further increase in progesterone concentrations. Granulosa cells from dominant follicles had increased levels of StAR, P450(scc), 3betaHSD, and aromatase during growth, but decreased levels during maturation. Levels of StAR, P450(scc), 3betaHSD, and aromatase, as well as progesterone and estradiol-17beta, were lower in granulosa cells from subordinate than from dominant follicles. We did not observe a relationship between the steroidogenic activity of follicles and the capacity of their enclosed oocytes to complete meiosis in vitro.     

Steroid hormone formation in bovine ovarian follicles.

In an attempt to assess histophysiological implication of the follicular compartment of the bovine ovary in steroid hormone formation and the effect of human chorionic gonadotropin (hCG) in vitro on follicular steroidogenesis, minces of follicular tissues from non-gravid bovine ovaries were incubated with radioactive testosterone or acetate in the presence and absence of hCG. Significant amounts of estrone and estradiol-17beta were formed on incubation with testosterone-4-14C; hCG decreased the conversion approximately by 30%. The major radioactive products formed from acetate-l-14C were androstenedione and testosterone with lesser amounts of dehydroepiandrosterone and 17-hydroxyprogesterone. In addition, small amounts of progesterone, 17-hydroxypregnenolone, estrone and estradiol-17beta were formed. Histology of the dissected follicle specimens was characterized by dominant theca cells undergoing luteinization with small amounts of granulosa cells, which showed neither proliferation nor luteinization. The pattern of distribution of radioactivity among the steroids formed from acetate-14C was considered to represent steroidogenic profile of bovine atretic follicles. The addition of hCG in vitro increased the overall incorporation of radioactive acetate into the steroids approximately by 50%, although the range of increase was not uniform in the individual steroids under the exprimental conditions.     

In situ endocrine disrupting effects of cadmium on the reproduction of Ruditapes decussatus

The gulf of Gabès is well known for its multiple contaminations which are closely associated with the past and present industrial and anthropogenic activity. In order to evaluate in situ the cadmium bioaccumulation in the clam Ruditapes decussatus sampled in this area, we measured cadmium concentrations in the whole soft tissues and in the digestive gland during one year. Energy reserves (proteins, glycogen, lipids) were monitored during gametogenesis in the remainder (mantle, adductor muscle, foot, siphons) of clams. Sex steroids (progesterone, testosterone, estradiol-17beta) were also measured by radio-immunoassay (RIA) in the gonads. The results showed that cadmium varied according to the reproductive cycle. High values were recorded at the resting stage in both sexes, during the gamete development and maturation in males and at the ripening and partially spawn stages in females. No significant changes in protein levels were observed in both females and males. However, high levels of glycogen and lipid reserves were recorded during the gametogenesis activity (from the development to the partially spawn stages). For steroid analyses, the results showed no important changes in testosterone levels all along the reproductive cycle. Fluctuations of sex steroids (especially progesterone and estradiol-17beta) could suggest their possible role as endogenous modulators of gametogenesis in R. decussatus. Correlation matrix and multiple regression models were also proposed to describe negative and significant relationship between cadmium and (glycogen, progesterone, testosterone and estradiol-17beta). These correlations may suggest the involvement of cadmium in the dysfunction of energy status and the endocrine disruption which could impede reproduction.     

Specificity of inhibition by steroids of porcine oocyte maturation in vitro.

In order to examine the influence of several steroids on the process of oocyte maturation, denuded (adherent cumulus granulosa cells mechanically removed) and intact (cumulus granulosa cells left attached) porcine oocytes were cultured in the presence or absence of estradiol-17 beta, estradiol-17 alpha, testosterone, cortisol, progesterone, or the nonsteroidal estrogen diethyl stilbestrol (all at 10 microgram/ml) in defined medium that contained either BSA or dextran. Estradiol-17 beta was the only steroid to exert a significant inhibitory effect on the maturation of denuded oocytes, and did so only in BSA supplemented medium. The inhibition was reversible in that oocytes, cultured in steroid-free medium after initial culture in estradiol-17 beta medium, resumed meiotic maturation. Oocytes took up 3H-estradiol-17 beta in both media, although less radiolabel entered oocytes in BSA supplemented medium. The majority of label in the oocytes, when cultured with either medium, was not displaced by excess radioinert estradiol-17 beta or progesterone, nor were the oocytes saturated even when cultured in 10(-6) M estradiol-17 beta. Autoradiography of sectioned oocytes after culture in 3H-estradiol-17 beta has shown that there was no selective accumulation of silver grains over the germinal vesicle as was the case with granulosa cell nuclei. This observation suggests that estradiol-17 beta may not act at the level of the oocyte nucleus.     

17 beta-hydroxysteroid and 20 alpha-hydroxysteroid dehydrogenase activities of human placental microsomes: kinetic evidence for two enzymes differing in substrate specificity

During storage at 4 degrees C, the 17 beta-hydroxysteroid dehydrogenase activity of human placental microsomes with estradiol-17 beta was more stable than that with testosterone. In order to evaluate the basis for this difference, kinetics with C18-, C19-, and C21- steroids as substrates and/or inhibitors was studied in conjunction with an analysis of the effects of detergents. Both 17 beta-hydroxysteroid dehydrogenase (17 beta-HSD) and 20 alpha-hydroxysteroid dehydrogenase (20 alpha-HSD) activities were detected. At pH 9.0, apparent Michaelis constants were 0.8, 1.3, and 2.3 microM for estradiol-17 beta, testosterone, and 20 alpha-dihydroprogesterone, respectively, 17 beta-HSD activity with testosterone was inhibited by estradiol-17 beta, 5 alpha-dihydrotestosterone, 5 beta-dihydrotestosterone, 20 alpha-dihydroprogesterone, and progesterone. In each case 90 to 100% inhibition was observed at 50 to 200 microM steroid. Activity with 20 alpha-dihydroprogesterone was similarly sensitive to inhibition by C19-steroids. By contrast, 25 to 45% of the activity with estradiol-17 beta was not inhibited by high concentrations of C19- or C21-steroids and differed from the 17 beta-HSD activity with testosterone and the major fraction of that with estradiol-17 beta by being insensitive to solubilization by detergent. These results are consistent with an association of two dehydrogenase activities with human placental microsomes. One recognizes C18-, C19-, and C21-steroids as substrates with comparable affinities. The second appears to be highly specific for estradiol-17 beta. The former activity may account for most if not all of the oxidation-reduction at C-17 of C19-steroids and at C-20 of C21-compounds at physiological concentrations by term placental tissue.     

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.     

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.     

A transgenic mouse model for investigating the response of the upstream region of whey acidic protein (WAP) gene to various steroid hormones.

The limitations of studies of clarification of response elements of whey acidic protein (WAP) gene to hormones using mammary cell lines has been shown. We studied the response of the upstream region (2.6 kb) of WAP to various steroid hormones using gonadectomized mWAP/hGH transgenic mice. Ovariectomy or castration for transgenic mice was performed at 10 days or 30 days post partum. Various steroid hormones were administered daily for 10 days to the gonadectomized transgenic mice after they reached 2 months of age. Prior to the hormonal administration and 24 hr after the final administration, blood was collected and the hGH levels in the plasma was measured by RIA. Daily doses of estradiol-17 beta were significantly more effective at increasing hGH levels in transgenic females ovariectomized at 10 days post partum than progesterone of an equal dose. A combined dose of progesterone and of estradiol-17 beta significantly amplified the increase of hGH levels accompanied by the great development of mammary glands, compared to a dose of progesterone alone. Corticosterone induced only a slight increase of hGH, while testosterone had no effect. The doses of gonadal steroid hormones did not induce an increase in hGH levels and development of mammary glands in the castrated transgenic males. The results showed that the response of 5' region of WAP requires at least some extended development of the mammary gland and that the 2.6 kb upstream region of the exogenous WAP gene contained the element responsive to ovarian hormones.     

Structural requirements for progesterone binding to the hepatic endoplasmic reticulum in the female rat

Microsomes isolated from the liver of the female rat specifically bind progesterone. The progesterone-microsomal complex shows highly specific characteristics. The binding is probably associated with the carbonyl groups at positions C-20 and C-3. Other steroids compete for microsomal binding sites less effectively. Competition for progesterone binding sites by other steroids in percentages: testosterone 33; testosterone propionate, 9; 17-methyltestosterone, 23.2; cortisol, 6.4; estradiol-17 beta, 1.8; 17 alpha-ethynyl estradiol, 4.7; mestranol, 1.0; norethynodrel, 4.5; ethisterone, 7.1; lynestrenol, 4.3; medroxyprogesterone, 23.3; medroxyprogesterone acetate, 15.2; 5 alpha-pregnane-3,20-dione, 47.6; 5 beta-pregnane-3,20-dione, 20.7; pregnenolone, 14.8; 6-methylpregnenolone, 1.2; 16 alpha-methylpregnenolone, 3.8%; 20 beta-hydroxy-4-pregnen-3-one, 2.8; 3 beta-hydroxy-5 alpha-pregnan-20-one, 5.2; 4-pregnene-3 beta, 20 beta-diol, 2.1; 11 alpha-hydroxyprogesterone 21.0; 16 alpha-hydroxyprogesterone, 7.9; 17-hydroxyprogesterone, 26.7; 16 alpha, 17-epoxyprogesterone, 2.7; 16 alpha-methylprogesterone, 3.8; 6-methylpregnenolone, 1.2; 16 alpha-methylpregnenolone, 3.8; promegestone, 27.0. 3 beta-Hydroxy-5 beta-pregnan-20-one, 3 alpha-hydroxy-5 beta-pregnan-20-one, 5-pregnene-3 beta,20 beta-diol, 5-pregnene-3 beta, 20 alpha-diol; 5 alpha-pregnane-3 beta, 20 beta-diol, 5 alpha-pregnane-3 beta, 20 alpha-diol, 5 beta-pregnane-3 alpha, 20 alpha-diol, 5 beta-pregnane-3 alpha, 20 alpha-diol diacetate, 5 beta-pregnane-3 alpha, 20 beta-diol, 3 alpha, 17-dihydroxy-5 beta-pregnan-20-one, 17-hydroxypregnenolone, 6-methyl-17-hydroxypregnenolone, pregnenolone-16 alpha-carbonitrile, dihydrotestosterone and cholesterol show no competition at all. The varying degree of competition by different steroids is unrelated to their lipid solubility.     

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.     

Uptake, distribution and binding of vertebrate and invertebrate steroid hormones and time-dependence of ponasterone A binding in Calliphora vicina. Comparisons among cholesterol, corticosterone, cortisol, dexamethasone, 5 alpha-dihydrotestosterone, 1,25-dihydroxyvitamin D3, ecdysone, estradiol-17 beta, ponasterone A, progesterone, and testosterone.

The presence of specific binding sites for radiolabelled vertebrate-type and arthropod-type steroid hormones was investigated in several organs including salivary gland, and central nervous system of third instar Calliphora vicina larvae by thaw-mount autoradiography. Ponasterone A, a 20-hydroxyecdysone agonist and 20-hydroxyecdysone are the only steroids which bind to nuclear high affinity binding sites. These binding sites are DNA associated while nucleoli show no tracer binding. Ecdysone, an endogenous 20-hydroxyecdysone precursor, is taken up by target cells but no significant nuclear binding occurs. 1,25-Dihydroxyvitamin D3 concentrates in cytoplasm only and its uptake is highest compared to all other steroids. Progesterone and testosterone show weak accumulation in the cytoplasm, while for cholesterol, corticosterone, cortisol, dexamethasone, dihydrotestosterone and estradiol-17 beta, no noticeable uptake occurs. For ponasterone A, a clear time dependence of uptake and intracellular distribution is visible, suggesting the existence and involvement of specific ecdysteroid uptake and transport mechanisms. These results suggest the presence of binding sites for various mammalian steroids in insects. Whether vertebrate steroid hormones or metabolites of them play a role in insects or whether the uptake and binding is based on chemical similarities alone without biological significance remains to be further investigated.