Experimental therapy of fetuses affected with congenital adrenal hyperplasia (CAH) has been reported by administering dexamethasone (Dex) to pregnant women at risk for carrying a CAH fetus. Such prenatal therapy can almost wholly eliminate virilization of the external genitalia of affected female fetuses, but only when treatment is started before 9 weeks of gestation. As it is not known whether the hypothalamic-pituitary-adrenal axis is functional at this time, and as the minimal effective doses of Dex are substantially supraphysiologic for the fetus, the mechanism of action of prenatal Dex treatment has been unclear. To assess the possibility that Dex might act directly on the adrenal, we used human adrenocortical NCI-H295A cells, an established model of the human fetal adrenal. Short term (6 h) incubation of these cells with Dex decreased synthesis of 11-deoxycortisol and 17alpha-hydroxyprogesterone and increased synthesis of deoxycorticosterone (DOC), but only at very high concentrations of Dex (> or =10 microM) that affect P450c17 (17alpha-hydroxylase/17,20 lyase) as a competitive inhibitor; no effects were seen at lower concentrations corresponding to those used in prenatal treatment. When NCI-H295A cells were treated with up to 100 microM Dex for 72 h, dot-blot analysis showed no changes in the abundance of the mRNAs for P450scc (cholesterol side-chain cleavage enzyme), P450c17, or 3beta-hydroxysteroid dehydrogenase II (3betaHSDII). When NCI-H295A cells were transfected with promoter/reporter constructs for the human genes for P450scc and P450c17, 24-h treatment with Dex at doses up to 100 microM had no effect on the transcription of these constructs. Because the regulation of steroidogenic processes in NCI-H295A cells closely resembles such regulation in the adrenal, we suggest that Dex may not act by an intra-adrenal mechanism in the experimental prenatal treatment of CAH.
Damian G Romero, Ming Yi Zhou, Licy L Yanes, Maria W Plonczynski, Tanganika R Washington, Celso E Gomez-Sanchez and Elise P Gomez-Sanchez
steroidogenesis, regulate adrenal cells through G-protein-mediated intracellular signaling pathways. G-proteins are heterotrimeric proteins composed of Gα , Gβ , and Gγ polypeptides. When the ligand binds its seven-trans-membrane G-protein-coupled receptor (GPCR
Guillaume Arguin, Yannik Regimbald-Dumas, Marc-Olivier Fregeau, Annabelle Z Caron and Gaetan Guillemette
coupling, tyrosine kinase receptors, Ca 2+ entry channels). Therefore, by acting at different levels of the Ca 2+ signalling cascade, PKC is providing diverse negative feedback loops that may produce additive inhibitory effects on intracellular Ca 2
F. A. Antoni, J. Hoyland, M. D. Woods and W. T. Mason
Stress provokes a cohort of homeostatic reflexes by the central nervous, the immune as well as the metabolic control systems of the body. These powerful adaptive responses, which can cause a collapse of body homeostasis in the absence of feedback inhibition, are suppressed by adrenal glucocorticoid hormones. A prominent and physiologically significant early action of glucocorticoids that requires the induction of newly synthesized messenger RNA and protein is the suppression of ACTH release by anterior pituitary corticotroph cells. It is demonstrated here that glucocorticoids inhibit stimulated ACTH secretion in pituitary corticotroph tumour (AtT-20) cells by reducing stimulus-evoked intracellular free calcium transients. Thus, the data show for the first time that intracellular calcium signals may be modified by rapidly induced proteins. It is proposed that this is a general mechanism that underlies the early inhibitory effects of glucocorticoids during stress in various types of cell.
S Hundertmark, H Buhler, M Rudolf, HK Weitzel and V Ragosch
This in vitro study on MCF-7 and ZR-75-1 breast cancer cells showed that the antiproliferative action of glucocorticosteroids (GCS) on breast cancer cells is weakened by a high oxidative activity of 11 beta-hydroxysteroid dehydrogenase (11 beta-HSD; EC 18.104.22.168): both endogenic as well as synthetic GCS (dexamethasone, prednisolone) were metabolised to hormonally inactive 11-dehydro metabolites. This enzymatic shield protected the breast cancer cells from the antiproliferative action of GCS. Continuous exposure of breast cancer cells to GCS resulted in enhanced 11 beta-HSD activity. The intracellular GCS concentration was further reduced by this feedback and thus the antiproliferative effect was additionally weakened. These mechanisms of GCS deactivation could be influenced by inhibiting 11 beta-HSD with the liquorice compound glycyrrhetinic acid (GLY). In MCF-7 and ZR-75-1 cultures the antiproliferative effect of GCS was significantly increased by GLY.
J R Seckl and T Olsson
Increased glucocorticoid secretion is a key feature of the stress response, serving to mobilise energy substrates, inhibit non-vital processes and restore stress effector systems. However, chronic glucocorticoid excess (in Cushing's disease or during pharmacotherapy) is associated with a broad spectrum of deleterious effects including diabetes mellitus, reproductive failure, hypertension, osteoporosis, immunosuppression, myopathy, growth impairment and, not least, affective and cognitive dysfunction. Clearly therefore, the autoregulatory (negative feedback) actions of glucocorticoids upon the hypothalamic-pituitary-adrenal (HPA) axis are of crucial importance.
Glucocorticoids act, in large part, by binding to intracellular receptors. There are two types, mineralocorticoid (MR, type I) and glucocorticoid (GR, type II) (McEwen et al. 1986) receptors. Ligand-activated receptors function as nuclear transcription factors, attaching to specific DNA sequences and regulating target gene expression. Interactions with other nuclear factors, notably AP-1 (Pfahl 1993) and cyclic AMP response element binding protein (Stauber et al. 1992), may also occur and modulate
M. ZANISI, M. MOTTA and L. MARTINI
It is now well established that testosterone can be converted intracellularly into 5α-androstan-17β-ol-3-one (dihydrotestosterone, DHT), 5α-androstan-3β,17β-diol (3β-diol) and 5α-androstan-3α,17β-diol (3α-diol) in order to become fully active on androgen-dependent peripheral structures (Robel, 1971). Massa, Stupnicka, Kniewald & Martini (1972) have shown that testosterone is also converted into DHT, 3α-diol and 3β-diol in the anterior pituitary and in the hypothalamus, i.e. in those structures on which androgens exert their feedback control on gonadotrophin secretion. The experiments to be described here were designed to discover whether: (a) the three 5α-reduced metabolites of testosterone mentioned above might exert an inhibitory action on gonadotrophin secretion; (b) the feedback activity of these steroids might be correlated with their androgenic potencies.
Adult male Sprague—Dawley rats, castrated 3 weeks before use, were each given one single subcutaneous injection of 2 mg of testosterone, DHT, 3α-diol or 3β-diol. All steroids were used in the free-alcohol form. Animals
V Viau and MJ Meaney
Hypothalamic-pituitary-adrenal (HPA) activity is governed by glucocorticoid negative feedback and the magnitude of this signal is determined, in part, by variations in plasma corticosteroid-binding globulin (CBG) capacity. Here, in gonadectomized male rats we examine the extent to which different testosterone replacement levels impact on CBG and HPA function. Compared with gonadectomized rats with low testosterone replacement ( approximately 2 ng/ml), plasma adrenocorticotropin and beta-endorphin/beta-lipotropin responses to restraint stress were reduced in gonadectomized rats with high testosterone replacement ( approximately 5 ng/ml). Plasma CBG levels also varied negatively as a function of testosterone concentration. Moreover, glucocorticoid receptor binding in the liver was elevated by higher testosterone replacement, suggesting that testosterone acts to enhance glucocorticoid suppression of CBG synthesis. Since pituitary intracellular CBG (or transcortin) is derived from plasma, this prompted us to examine whether transcortin binding was similarly responsive to different testosterone replacement levels. Transcortin binding was lower in gonadectomized rats with high plasma testosterone replacement ( approximately 7 ng/ml) than in gonadectomized rats with low testosterone replacement ( approximately 2 ng/ml). This testosterone-dependent decrease in pituitary transcortin was associated, in vitro, with an enhanced nuclear uptake of corticosterone. These findings indicate that the inhibitory effects of testosterone on corticotrope responses to stress may be linked to decrements in plasma and intrapituitary CBG. This could permit greater access of corticosterone to its receptors and enhance glucocorticoid feedback regulation of ACTH release and/or proopiomelanocortin processing.
A V Sirotkin
The reciprocal control of nonapeptide hormone (oxytocin, vasopressin) and cyclic nucleotide (cAMP, cGMP) release by porcine granulosa cells was studied. In particular, the influence of vasopressin and oxytocin treatment (10–10 000 ng/ml) on basal and LH-induced cAMP and cGMP output, as well as the effects of dibutyryl cAMP (dbcAMP; cAMP analogue) and forskolin (a stimulator of cAMP formation; 0·1–1000 ng/ml) on vasopressin and oxytocin secretion by cultured porcine granulosa cells were examined.
It was observed that the addition of arginine-8-vasopressin or oxytocin stimulated both cAMP and cGMP output from granulosa cells. Moreover, both vasopressin and oxytocin also increased LH-stimulated cAMP and cGMP release. On the other hand, both dbcAMP and forskolin decreased vasopressin secretion. Oxytocin release was stimulated under the influence of dbcAMP. The same stimulating effect occurred with forskolin given at a low dose (1 ng/ml), whilst higher doses of forskolin (10 or 1000 ng/ml) were inhibitory.
The present observations demonstrate the reciprocal influence of nonapeptide hormones and cyclic nucleotides in porcine ovarian cells. Oxytocin and vasopressin, like LH, exert their action on the ovary via the activation of cAMP- and cGMP-dependent intracellular mechanisms. cAMP in turn inhibits vasopressin release through a negative feedback mechanism. On the other hand, a reciprocal stimulation of oxytocin and cAMP output in granulosa cells is suggested. Thus, cyclic nucleotides can be both regulators of nonapeptide hormone secretion and mediators of their action within porcine ovaries.
Journal of Endocrinology (1996) 150, 343–348
S. W. Manley, D. S. Rose, G. J. Huxham and J. R. Bourke
The calcium ionophore A23187 (0·1–1 μmol/l) inhibited membrane electrical polarization, uptake of 125I, fluid transport and TSH-stimulated release of radioiodine from the organic pool in follicular cultures of porcine thyroid cells. At higher concentrations (1–30 μmol/l), A23187 promoted release of radioiodine from the organic pool. Stimulation of release of radioiodine from the organic pool by veratridine (a sodium channel agonist, 0·4–1 mmol/l) and A23187 was dependent on the calcium concentration of the medium, while TSH action was independent. Incubation in medium of very low calcium concentration (0·0177 mmol/l) resulted in enhanced release from the organic pool, which was inhibited by TSH (256 μU/ml), A23187 (25 μmol/l) or veratridine (0·5 mmol/l). These data therefore do not support the hypothesis that calcium acts as a mediator of the secretomotor action of TSH, but suggest the possibility of a TSH-induced increase in intracellular calcium as a regulatory negative-feedback mechanism.
J. Endocr. (1988) 116, 373–380