Loss of sensitivity to ACTH of adrenocortical cells isolated from maturing domestic fowl

Maturation of domestic fowl corticosteroidogenesis was evaluated using purified adrenocortical cells. Basal corticosterone production decreased steadily from 2 days to 26 weeks after hatching. However, maximally stimulated corticosterone production was not changed. In contrast, the half-maximal steroidogenic concentrations (ED50 values or effective doses for 50% maximal effect) of ACTH analogs increased approximately 40 times by 26 weeks, but the ED50 values of 8-bromo-cyclic AMP and pregnenolone were not changed. This suggests that adrenocortical cell sensitivity to ACTH decreases with maturation of the domestic fowl.     

Ontogenic corticosteroidogenesis of the domestic fowl: response of isolated adrenocortical cells

Ontogenic adrenocortical function of the domestic was investigated using adrenocortical cells isolated from embryonic chicks (18, 19, 20, and 21 days old) and male and female posthatch birds (1 day, 1 week, and 3 weeks old). Production of the predominant corticosteroids secreted by the chicken adrenal gland, corticosterone, cortisol, and aldosterone, was measured by radioimmunoassay after 2-hr incubation of cells with or without steroidogenic agents. Approaching hatch, basal and maximal ACTH-(1-24) (ACTH)-induced corticosteroid production increased steadily and peaked around 1 day posthatch (5-18 times and 3-9 times, respectively, the production values at 18 days embryonic life). Thereafter, corticosteroid production values decreased steadily to 3 weeks posthatch. Corticosterone predominated over the ages studied: Maximal ACTH-induced corticosterone production averaged 52 and 115 times the production values of aldosterone and cortisol, respectively. In addition, maximal ACTH-induced aldosterone production was roughly 2.2 times greater than cortisol production over the ages studied except for a short-lived, disproportionately greater aldosterone production at 1 day posthatch. In addition to perihatch and age-related differences in cellular corticosteroid production, there were also differences in cellular sensitivity to steroidogenic agents as indicated by the differences in half-maximal steroidogenic concentration values (ED50 values) of the steroidogenic agents. Sensitivity to ACTH increased 2.7 times from Day 18 of embryonic life to 1 day posthatch and then decreased steadily to 3 weeks posthatch. In addition, sensitivity to 8-bromo-cAMP (8-Br-cAMP) increased abruptly at 1 day posthatch (nearly 3 times) but then remained constant thereafter. However, a consistent change in cellular sensitivity to 25-hydroxycholesterol was not observed until 3 weeks posthatch (an increase in sensitivity of 3 times that at Day 18 of embryonic life). These data of cellular sensitivity suggest that there were distinct development and maturational alterations in the cellular loci at which ACTH, 8-Br-cAMP, and 25-hydroxycholesterol acted. Thus, during the transition from embryonic to postembryonic life of the domestic fowl, there are alterations in adrenocortical cell steroidogenic capacity and in the function of some cellular loci comprising the corticosteroidogenic pathway.     

Isolated adrenocortical cells of the domestic fowl (Gallus domesticus): steroidogenic and ultrastructural properties

Isolated adrenocortical cells from White Leghorn chickens (Gallus domesticus) were compared to those from rats (Rattus norvegicus). Cells were prepared from collagenase-dispersed adrenal glands of sexually mature male animals. Corticosterone was measured by radioimmunoassay after incubation for 2 h with steroidogenic agents. Of the four ACTH analogues used, three were 6-17 times more potent with rat cells than with fowl cells (potencies were indicated by half-maximal steroidogenic concentrations). However, 9-tryptophan (O-nitrophenylsulfenyl) ACTH was 8 times more potent with fowl cells than with rat cells, thus suggesting that ACTH receptor differences exist between the two cell types. In addition, cAMP analogues were 10 times more potent with rat cells than with fowl cells suggesting that fowl corticosteroidogenesis is less dependent on cAMP than is rat corticosteroidogenesis. At equal cell concentrations, rat cells secreted 20-40 times more corticosterone than did chicken cells when they were maximally stimulated. Although rat cells converted 8 times more pregnenolone to corticosterone than did fowl cells, the half-maximal steroidogenic concentration for pregnenolone-supported corticosterone synthesis was the same for both cell types (about 5 microM). This suggests that fowl cells have lower steroidogenic enzyme content rather than lower steroidogenic enzyme activity. An unusual feature seen in the isolated fowl adrenocortical cells was an abundance of intracellular filaments.     

Metabolic regulation of steroid steroidogenesis in isolated adrenal and adrenocortical carcinoma cells of rat. The incorporation of (20S)-20-(7- 3 H)hydroxycholesterol into deoxycorticosterone and corticosterone

Previous studies from these laboratories have demonstrated the following: (1) that cyclic 3′5′-AMP (c-AMP) phosphodiesterase activity of adrenocortical carcinoma 494 is only 20% of that found in the normal adrenal; (2) that corticosteroido-genesis in the isolated tumor cells is inhibited by ACTH, and (3) that the normal adrenal biosynthetic pathway from pregnenolone to corticosterone is intact but less active in the tumor. The present studies show that both the isolated adrenocortical carcinoma cell and the normal isolated adrenal cell of the rat have the capacity to transform (20S)-20-hydroxycholesterol into deoxycorticosterone and corticosterone. It is, therefore, proposed that the lack of stimulation by ACTH of corticosterone synthesis of the tumor cells cannot be explained by the absence of enzymes cleaving the cholesterol side chain. It is, therefore, postulated that a modified protein kinase may be present in the tumor which is not stimulated by c-AMP.     

Development of the ACTH and corticosterone response to acute hypoxia in the neonatal rat

Acute episodes of severe hypoxia are among the most common stressors in neonates. An understanding of the development of the physiological response to acute hypoxia will help improve clinical interventions. The present study measured ACTH and corticosterone responses to acute, severe hypoxia (8% inspired O(2) for 4 h) in neonatal rats at postnatal days (PD) 2, 5, and 8. Expression of specific hypothalamic, anterior pituitary, and adrenocortical mRNAs was assessed by real-time PCR, and expression of specific proteins in isolated adrenal mitochondria from adrenal zona fascisulata/reticularis was assessed by immunoblot analyses. Oxygen saturation, heart rate, and body temperature were also measured. Exposure to 8% O(2) for as little as 1 h elicited an increase in plasma corticosterone in all age groups studied, with PD2 pups showing the greatest response ( approximately 3 times greater than PD8 pups). Interestingly, the ACTH response to hypoxia was absent in PD2 pups, while plasma ACTH nearly tripled in PD8 pups. Analysis of adrenal mRNA expression revealed a hypoxia-induced increase in Ldlr mRNA at PD2, while both Ldlr and Star mRNA were increased at PD8. Acute hypoxia decreased arterial O(2) saturation (SPo(2)) to approximately 80% and also decreased body temperature by 5-6 degrees C. The hypoxic thermal response may contribute to the ACTH and corticosterone response to decreases in oxygen. The present data describe a developmentally regulated, differential corticosterone response to acute hypoxia, shifting from ACTH independence in early life (PD2) to ACTH dependence less than 1 wk later (PD8).     

Glucocorticoid enhancement of glucocorticoid production by cultured ovine adrenocortical cells

The present study examines the effect of chronic treatment with glucocorticoids on the steroidogenic activity of ovine adrenocortical cells in vitro. Cells cultured in the presence of 10(-9) to 10(-5) M dexamethasone produced more glucocorticosteroids in response to ACTH1-24, forskolin or 8 BrcAMP than did control cells. Such an enhancing effect required more than 5 h of treatment and was maximal at 30 h; it was both concentration-dependent and steroid-specific. The maximal secretion of corticosteroids was observed when cells were exposed to 10(-7) M dexamethasone; with higher concentrations the response to ACTH1-24 decreased steadily; the ED50 was 2.8 +/- 0.8 nM. Cortisol and corticosterone enhanced ACTH1-24-induced steroidogenesis to the same extent as dexamethasone, but at concentrations roughly 100-fold higher than for dexamethasone. Testosterone and 17 beta-oestradiol had no enhancing effect. Dexamethasone not only enhanced the maximal steroidogenic response to ACTH1-24 but also decreased its ED50 3-fold. Treatment of cultures with the antiglucocorticoid RU 38486 resulted in a dose-dependent, time-dependent, decrease in ACTH1-24-induced corticosteroid output. Moreover, RU 38486 antagonized the enhancing effect of dexamethasone. The production of corticosteroids by dexamethasone-treated cells incubated in the presence of 22(R)-hydroxycholesterol or of exogenous pregnenolone was similar to that of control cells. The enhancing effect of dexamethasone was also observed when cultures were performed in the absence of insulin and/or in serum-free media. These data suggest that chronic exposure to glucocorticoids is necessary for the full steroidogenic activity of ovine adrenocortical cells. Moreover, they indicate that glucocorticoids exert their effect at least at two different levels in the cell: (i) on the adenylate cyclase system and (ii) at step(s) beyond cAMP but before pregnenolone formation.     

Influence of triiodothyronine and growth hormone on growth of dwarf and normal chickens: interactions of hormones and genotype

The effects of dietary triiodothyronine (T3), injections of a preparation of growth hormone (GH) (purified from chicken pituitary tissue) and their combination on growth were investigated in three lines of chickens. The three lines were the Cornell K strain (K) (a single Comb White Leghorn strain), the Cornell K strain hemizygous for the sex-linked dwarfing gene (SLD), and the Cornell K strain homozygous recessive for the autosomal dwarfing gene (ADW). A dietary T3 treatment by genotype interaction was observed. Dietary T3 (0.1 ppm) decreased growth in the K line, tended to decrease growth in the ADW line while it tended to increase growth in the SLD line. Chicken growth hormone (100 micrograms/kg body wt) alone did not affect growth in any of the lines studied. There was, however, a GH treatment by T3 treatment interaction. Chicken GH overcame the growth-depressing effects of T3 in the K and ADW lines while it tended to promote growth in T3 treated SLD birds. Dwarf (SLD) chickens had higher basal circulating GH concentrations, lower circulating immunoreactive somatomedin C concentrations, and lower circulating T3 concentrations than the K or ADW chickens.     

Adrenal adenylate cyclase and steroidogenic activities of 63 day old ovine fetuses: in vitro effects of ACTH1-24 and forskolin

This study examines the activity of the adenylate cyclase system and that of some enzymes of the steroidogenic pathway of adrenal cells from 62-63 day old ovine fetuses. Synthetic corticotropin (ACTH1-24), cholera toxin and forskolin stimulated both cAMP and corticoid productions by freshly isolated adrenal cells. The cAMP response to ACTH1-24 was lower than that to forskolin. However, forskolin-induced steroidogenesis was significantly lower than the ACTH1-24-induced steroid output. Freshly isolated cells metabolized quickly [14C]-labeled pregnenolone mainly through the 17-deoxy pathway. The amounts of cortisol and of corticosterone formed, in the presence of exogenous pregnenolone, were roughly 15-fold higher than under maximal stimulation by ACTH1-24. When the cells were cultured for 6 days in the absence or presence of ACTH1-24 (10(-8) M) or forskolin (10(-5) M), a small development of the cAMP response to these factors was observed in the course of the experiment. However, the mechanism of this development appeared different, according to the conditions of culture. The amounts of corticosterone secreted on day 6 by ACTH1-24- or forskolin-treated cells were 2- to 4-fold higher than on day 1, whereas cortisol outputs were much lower on day 6 than on day 1. The response to ACTH1-24 of cells maintained in ACTH-free media decreased dramatically during the culture in terms of both cortisol and of corticosterone. On day 6 of the experiment, the metabolism of [14C]pregnenolone was lower than on day 1 under all 3 conditions of culture. Only the 3 beta-hydroxysteroid dehydrogenase/isomerase activity could be maintained by continuous treatment with forskolin. However, both ACTH1-24 and forskolin enhanced the production of pregnenolone from an endogenous substrate. In conclusion, these results present evidence that: 1) the adenylate cyclase system is not a bottleneck in the steroidogenic response to ACTH1-24 of freshly isolated adrenal cells from 62-63 day old ovine fetuses; 2) the main rate-limiting step for steroidogenesis by these cells is the availability of pregnenolone; 3) neither ACTH1-24 nor forskolin is able to maintain the activity of most enzymes involved in the metabolization of pregnenolone by cultured cells while increasing pregnenolone availability; 4) some inhibiting factors are involved in the loss of adrenal cells responsiveness to ACTH between days 50 and 100 of gestation, and they probably act mainly on the adenylate cyclase system.     

Extraction of corticosterone from cell homogenates and subcellular fractions of the rat adrenal cortex. III. ACTH-induced temporal subcellular redistributions of steroid precursors to corticosterone

Cholesterol, pregnenolone, progesterone, 11-deoxycorticosterone (11-DOC) and corticosterone were quantitated in subcellular fractions isolated from in vivo adrenocorticotropin (ACTH)-stimulated rat adrenal zona fasciculata/reticularis. Six adrenal subcellular fractions separated by discontinuous sucrose gradient centrifugation (lipid, 0.125 M sucrose, cytosolic, microsomal, mitochondrial and nuclear) were extracted with alkaline ether/ethanol and assayed by high pressure liquid chromatography (HPLC). Lipid fractions contained the major cholesterol stores, while most pregnenolone and progesterone was found in lipid, microsomal and mitochondrial fractions. The 0.125 M sucrose and cytosol fractions together contained approximately 75% of the total 11-DOC and corticosterone. The five steroids were only present in small amounts in organelle fractions containing steroidogenic enzymes. Homogenate and lipid fraction cholesterol decreased between 10 and 15 min and again 30 min after ACTH injection. In the homogenate, lipid, microsomal and mitochondrial fractions, pregnenolone and progesterone were increased after ACTH injection; peak pregnenolone and progesterone concentrations were often measured in adrenal gland sucrose, cytosolic, microsomal and mitochondrial fractions 15 to 20 min after rats were injected with ACTH. Although ACTH increased 11-DOC and corticosterone in all but the mitochondrial and nuclear fractions, the sucrose, cytosolic and microsomal 11-DOC, and cytosolic corticosterone increased most dramatically. In many fractions, peak 11-DOC and corticosterone concentrations were most often observed between the 10 and 15 min periods and again at 30 min.     

Effects of triiodothyronine treatments on body and organ growth and the development of immune function in dwarf chickens

The effects of triiodothyronine (T3) treatments on general body growth, long bone growth, primary lymphoid organ development, antibody production, and serum growth hormone (GH) and thyroid hormone levels were examined in two dwarf strains (sex-linked dwarf--SLD, and autosomal dwarf--ADW) and in a normal-growing control strain (K) of White Leghorn chickens. One-day-old male chicks from each of these strains were assigned to either an untreated control group or to one of the groups receiving a T3 supplement ranging from 0.01 to 1.0 ppm. General body growth and long bone growth were significantly (P less than 0.05) stimulated only within the SLD strain by the intermediate T3 dosages. The 1.0-ppm T3 dosage level resulted in depressed body weights within both the K and ADW strains but produced no significant changes within the SLD strain. Thymic growth was significantly stimulated due to treatments of 0.1 ppm T3 in the SLD strain (P less than 0.05) and 1.0 ppm T3 in both the SLD and ADW strains (P less than 0.001 and P less than 0.05, respectively). Bursal growth was significantly depressed (P less than 0.05) at all T3 dosage levels within the SLD strain while 0.01 and 0.1-ppm T3 treatments resulted in significant bursal growth stimulation in the K and ADW strains, respectively. Concomitant with the depressed bursal growth, antibody production was significantly depressed (P less than 0.05) within the SLD strain at the 1.0-ppm T3 dosage level. Antibody production was not significantly affected by any of the T3 treatments within the control K or ADW strains. Serum T3 levels were significantly increased in all strains by the T3 supplementation but thyroxine (T4) serum levels were affected only within the SLD strain. The 0.01-ppm T3 treatment resulted in a significant increase (P less than 0.05) in serum T4 levels within this strain and treatment group. The only increase (P less than 0.05) in GH levels due to T3 treatments occurred within the same SLD treatment group. The higher T3 treatments resulted in serum GH levels being severely depressed (P less than 0.01) in all strains.     

Seasonal alterations in adrenocortical cell function associated with stress-responsiveness and sex in the eastern fence lizard (Sceloporus undulatus)

We characterized steroidogenic properties of dispersed adrenocortical cells from field-active male and female eastern fence lizards (Sceloporus undulatus) to investigate whether alterations in cell function could, in part, explain seasonal variation in baseline and stress-induced plasma corticosterone (B). Lizards were collected during the breeding and postbreeding seasons and shortly prior to hibernation. Dispersed cells in vitro produced B, aldosterone (ALDO), and progesterone in response to 8-Br-cAMP, 25-(OH)cholesterol, adrenocorticotropin (ACTH; as little as 100 fM), and angiotensin II. Maximal progesterone, B, and ALDO responses to ACTH were roughly 1000%, 500%, and 100% greater than corresponding basal values. Angiotensin II was an effective steroidogenic stimulant but much less so than ACTH. Corticosteroid production exhibited considerable steroid-specific variation among seasons. Maximal ACTH-induced B production was lower in the postbreeding season than at either of the other two measurement points, essentially opposite to the pattern for ALDO. Males and females generally produced B at similar rates, but ALDO and progesterone showed numerous sex differences that usually covaried between the two steroids. Cellular sensitivity to 25-(OH)cholesterol and angiotensin II showed few sex differences or seasonal changes. In contrast, sensitivity to ACTH decreased markedly from the breeding to the postbreeding season in males, corresponding to the decrease in stress-responsiveness, and in both sexes was considerably lower prior to hibernation than during the breeding season. Under some conditions, plasma B may be limited by the production capacity of adrenocortical cells. In summary, seasonal variations in body condition, reproductive activity, and baseline and stress-induced plasma B may be attributed at least in part to alterations in adrenocortical cell steroidogenic function.     

Enhanced 17 alpha-hydroxylation of pregnenolone and increased androgen production by rabbit adrenocortical cells stimulated chronically with corticotropin

The postulated chronic stimulatory effect of corticotropin (ACTH) on pregnenolone production and on 17 alpha-hydroxylase activity was evaluated on adrenocortical cells obtained from control and chronically ACTH-treated rabbits. The cells were incubated with various concentrations of ACTH added alone or together with trilostane, so as to inhibit further conversion of pregnenolone and dehydroepiandrosterone. The maximal steroidogenic effect of ACTH (determined in the absence of trilostane) was increased 2-fold in adrenocortical cells from ACTH-treated animals; furthermore, cortisol production was increased whereas that of corticosterone decreased. While the generation of pregnenolone was of comparable magnitude for cells from both experimental groups, chronic in vivo treatment with ACTH was followed by a 40-fold enhancement in 17-hydroxypregnenolone production. Concomitantly, maximal DHEA production documented in the presence of ACTH and trilostane was enhanced more than 200-fold, from 0.45 +/- 0.20 pmol in control rabbits to 147 +/- 67 pmol in cells from ACTH-treated animals. The corresponding values of DHEA-sulphate production were 0.86 +/- 0.12 and 432 +/- 334 pmol, respectively. Thus, a prolonged stimulatory effect of ACTH on rabbit adrenocortical cells consists in an enhancement of the capacity to generate pregnenolone, and to convert this compound into 17-hydroxylated steroids.     

The effects of ACTH, phytoestrogens and estrogens on corticosterone secretion by gander adrenocortical cells in breeding and nonbreeding seasons

The aim of this study was to investigate the effects of ACTH, phytoestrogens (genistein, daidzein, biochanin A and coumestrol), and animal estrogens (estradiol and estrone) on corticosterone secretion by isolated adrenocortical cells of the ganders in breeding (April) and nonbreeding seasons (July). ACTH stimulated corticosterone output in the breeding season. In July (photorefractoriness and postbreeding molt) ACTH had no effect on corticosterone production. Coumestrol reduced corticosterone secretion by the cells obtained in nonbreeding season. Other examined phytoestrogens did not affect corticosterone production. Estrogens showed differentiated effects. Estradiol stimulated the corticosterone output in breeding season; estrone inhibited corticosterone release in July. The season can probably affect sensitivity of isolated gander adrenal cells, especially to ACTH. It seems that goose adrenocortical cells, in contrast to the mammalian cells, can be weakly sensitive to phytoestrogens.     

Chronic administration of the non-peptide CRH type 1 receptor antagonist antalarmin does not blunt hypothalamic-pituitary-adrenal axis responses to acute immobilization stress.

Antalarmin is a pyrrolopyrimidine compound that antagonizes corticotropin-releasing hormone (CRH) type 1 receptors (CRHR1). In order to assess the effects of antalarmin treatment on hypothalamic-pituitary-adrenal (HPA) function we measured the plasma concentrations of adrenocorticotropic hormone (ACTH) and corticosterone in animals treated with either antalarmin or vehicle for 1 week or for 8 weeks. We found that antalarmin treatment for 1 week did not affect basal concentrations of ACTH or corticosterone. In contrast, treatment for 8 weeks significantly lowered basal ACTH and corticosterone concentrations and also significantly decreased the basal corticosterone to ACTH ratio, indicating decreased basal adrenocortical responsiveness to ACTH. However, immobilization stress resulted in ACTH and corticosterone concentrations that were the same in animals treated with vehicle or antalarmin for either 1 or 8 weeks. We conclude that even though 8-week antagonism of CRHR1 by the non-peptide antalarmin blunts basal concentrations of ACTH and corticosterone, and affects the adrenal responsiveness to ACTH, it does not blunt the HPA response to acute stress, and it does not appear to cause stress-induced adrenal insufficiency.     

Adrenal splanchnic innervation contributes to the diurnal rhythm of plasma corticosterone in rats by modulating adrenal sensitivity to ACTH

Activity of the hypothalamic-pituitary-adrenal axis is characterized by a diurnal rhythm with an AM nadir and PM peak. Splanchnic nerve transection disrupts the diurnal rhythm in plasma corticosterone; however, there is a controversy as to whether the nerve-mediated effect is 1) via inhibition in the AM vs. excitation in the PM, or 2) involves changes in adrenal sensitivity to ACTH. The present studies were designed to address these issues. Adult male rats were anesthetized and underwent bilateral transection of the thoracic splanchnic nerve or sham transection. One week after surgery, rats were killed in the AM or PM with collection of nonstress plasma for measurement of corticosterone and ACTH. Plasma corticosterone was increased in the PM relative to the AM; however, plasma corticosterone in the PM was attenuated by splanchnic nerve transection, without affecting plasma ACTH. This decrease in PM plasma corticosterone after nerve-transection was 1) associated with decreased adrenal responsivity to ACTH, 2) associated with decreased adrenal cAMP content, 3) prevented by adrenal demedullation, and 4) not affected by removal of adrenal capsaicin-sensitive afferent fibers. Repeated serial blood sampling from individual rats confirmed the excitatory effect of splanchnic innervation in the PM. These results support the hypothesis that the adrenal splanchnic innervation modulates the diurnal rhythm in plasma corticosterone by increasing adrenal responsivity to ACTH and augmenting steroidogenesis in the PM and suggest that alterations in adrenal corticosterone secretion obscured by pulsatile secretion are more clearly revealed with repeated serial blood sampling.     

Effect of the growth hormone-secreting tumor StW5 on pituitary and adrenal gland function in rats

A growth hormone-secreting tumor (StW5 was implanted into male rats and resulted in a tripling of adrenal weight concomitant with a 30% decrement in pituitary weight. Plasma concentrations of corticosterone in tumor-bearing (TB) rats were significantly elevated at rest or after ACTH injections or the stress of either anesthesia. The rise in plasma concentrations of corticosterone was due mainly to the large increment in adrenal size although a significant increase in adrenal responsiveness to ACTH was demonstrated in vitro. In addition, plasma corticosterone concentrations were higher in TB rats despite both a doubling of the blood volume and a 50% increase in liver capacity to metabolize corticosterone. Pituitary ACTH content was significantly lower in TB rats, but these pituitary glands could still release near-normal quantities of ACTH as shown both by in vitro incubations and adrenal corticosterone output following ether stress.     

Mechanism of glucocorticoid enhancement of the responsiveness of ovine adrenocortical cells to adrenocorticotropin

The present studies were undertaken to precise the mechanism through which glucocorticoids enhance the responsiveness of ovine adrenocortical cells to ACTH. Experiments using intact cells and crude adrenal membranes have shown that, at the level of the adenylate cyclase system, dexamethasone increases the number of ACTH receptors without modification of the catalytic subunit or of the GTP binding regulatory components Gs and Gi. Cells cultured with dexamethasone secreted more pregnenolone and more corticosteroids in response to 8-BrcAMP than did control cells. By contrast, dexamethasone did not increase corticosterone secretion by cells incubated in the presence of 22-(R)-hydroxycholesterol or of exogenous pregnenolone. Dexamethasone neither affected the incorporation of [14C] acetate into cellular cholesterol nor the amount of cholesterol present in mitochondria of unstimulated cells. However, dexamethasone-treated cells incubated in the presence of both 8-BrcAMP and aminoglutethimide exhibited higher amounts of mitochondrial cholesterol than control cells. These data indicate that dexamethasone enhances the number of cellular ACTH receptors together with increasing the cAMP-induced translocation of cholesterol from the cytoplasm into mitochondria and/or mitochondrial storage of cholesterol.     

Photoperiodically-induced changes in hypothalamic-pituitary-adrenal axis sensitivity in captive house sparrows (Passer domesticus)

We used captive house sparrows (Passer domesticus) to identify regulatory mechanisms underlying seasonal (mimicked by changes in photoperiod) and diel differences in corticosterone output. Corticosterone responses were measured during three simulated seasons: short-day and long-day photoperiods and while birds underwent a pre-basic molt. Under all three conditions we tested for adrenal sensitivity by injecting exogenous ACTH, for pituitary sensitivity by injecting corticotropin-releasing factor (CRF) and arginine vasotocin (AVT), and for diel changes by repeating the injections during the day and at night. The daytime adrenal sensitivities were greatest on long days, lower on short days, and lowest during molt. These data suggest that reductions in either adrenal sensitivity to ACTH and/or capacity to secrete corticosterone could explain lowered endogenous corticosterone titers during molt. Furthermore, adrenal sensitivity to ACTH and pituitary sensitivity to AVT appeared to be greatest at night. This suggests that both the adrenal's sensitivity to the ACTH signal and the pituitary's capacity to secrete ACTH might provide a mechanism allowing for diel changes in corticosterone titers. This differs substantially from what is known about diel regulation in rodents. Taken together, these data provide further evidence that there are complex regulatory mechanisms controlling diel and seasonal changes in corticosterone titers in birds.     

A correlated stereological and functional studies on the long-term effects of ACTH on rat adrenal cortex

The aim of the study was to investigate the effect of prolonged ACTH administration on quantitative structural changes of the rat adrenal cortex and on function of its cells with particular emphasis on correlation of the results of biochemical estimations with stereologic parameters. Daily injections of 20 micrograms ACTH (Synacthen, Depot) for 35 days results in a marked enlargement of the cortex due to an increase in the volume of all the zones. This increase depends upon hypertrophy and hyperplasia of parenchymal cells. At day 21 of experiment the number of parenchymal cells markedly decreased if compared with day 14, the lost of cells being observed mainly in the zona reticularis. Prolonged ACTH treatment only insignificantly changed serum corticosterone concentration and--if calculated per mg of adrenal weight--did not change adrenal corticosterone concentration and 11 beta-hydroxylase activity and decreased corticosterone output by adrenal homogenate. If expressed per adrenocortical cell or per pair of glands, ACTH increased corticosterone concentration and 11 beta-hydroxylase activity while the drop in corticosterone output occurred only at days 28 and 35 of experiment. The striking differences in and among various functional parameters depicting adrenal steroidogenesis show for necessity--in case of long-term experiments leading to hypertrophy or atrophy of the gland--of using coupled stereologic and biochemical techniques which better evaluate the cytophysiological state of adrenocortical cells.     

Effect of synthetic atrial natriuretic polypeptide on hemorrhage-induced adrenocorticotropin secretion of the rat

The effect of synthetic alpha-human atrial natriuretic polypeptide (alpha-hANP) on the in vivo and in vitro release of ACTH and corticosterone was examined. In the in vivo study ACTH and corticosterone responses to rapid 2-ml/rat hemorrhage were measured in sixteen conscious rats after alpha-hANP administration. The hemorrhage increased plasma ACTH and corticosterone concentrations in the control group of rats (p greater than 0.01). ANP inhibited hemorrhage-induced ACTH secretion (p less than 0.05), but the plasma corticosterone response was not affected. In the in vitro study a high concentration of ANP (1 microM) reduced basal corticosterone secretion from the isolated rat adrenal gland (p less than 0.05), but the response to ACTH (10 ng/ml) and dibutyryl cyclic AMP (0.5 mM, 5.0 mM) was not affected. Our data suggest that ANP inhibits hemorrhage-induced ACTH secretion from the anterior pituitary but inhibits corticosterone secretion from the adrenal gland very weakly.