Natriuretic peptides are negative modulators of adrenocortical cell function of the eastern fence lizard (Sceloporus undulatus)

Elucidation of the role of natriuretic peptides (NPs) in vertebrate adrenal steroidogenesis has been facilitated by the use of freshly dispersed adrenocortical cells. Our recent characterization of lizard adrenocortical cells [Carsia, R.V., John-Alder, H.B., 2003. Seasonal alterations in adrenocortical cell function associated with stress-responsiveness and sex in the Eastern Fence Lizard (Sceloporus undulatus). Horm. Behav. 43, 408-420] provided the opportunity to examine the influence of atrial natriuretic peptides (ANPs) and related NPs on reptilian adrenal steroidogenesis at the cellular level. In the present report, the action of NPs on lizard adrenal steroidogenesis was investigated using freshly dispersed adrenocortical cells derived from the Eastern Fence Lizard (Sceloporus undulatus). Basal production rates of aldosterone and corticosterone and maximal angiotensin II (ANG II)-induced production rates of these corticosteroids were inhibited with high efficacy (75-90%) by rat ANP at potencies of 0.4-0.7 nM. By contrast, rat ANP had no effect on maximal production rates of these corticosteroids in response to a maximal steroidogenic concentration of adrenocorticotropin (ACTH; 1 nM). However, rat ANP inhibited aldosterone and corticosterone production rates in response to a half-maximal steroidogenic concentration of ACTH (10 pM; ∼50 pg/ml), albeit with less efficacy (∼50%) and potency (∼6 nM) than for ANG II. Rat and eel ANP and rat and chicken brain natriuretic peptide (BNP) were equally efficacious at inhibiting maximal ANG II-induced aldosterone and corticosterone production but with different potencies. The order of inhibitory potency was rat ANP = chicken BNP > eel ANP > rat BNP. However, a specific peptide ligand for the NP clearance receptor was without effect. This study indicates that ANP and related NPs are efficacious inhibitors of lizard adrenal steroidogenesis by acting directly at the level of the adrenocortical cell.     

Effects of food restriction on steroidogenesis in dispersed adrenocortical cells from Yarrow's Spiny Lizard (Sceloporus jarrovii)

Changes in energy balance can lead to functional alterations at all levels of the hypothalamic-pituitary-adrenal (HPA) axis. However, relatively little is known about how energy balance affects functional properties of adrenocortical cells themselves. We investigated effects of restricted food intake on sensitivity to ACTH and rates of steroidogenesis in adrenocortical cells isolated from growing female and male Yarrow's Spiny Lizards (Sceloporus jarrovii). At the end of the feeding regimen, we assayed acute (3h) progesterone (P(4)), corticosterone (B), and aldosterone (ALDO) production in response to ACTH in dispersed adrenocortical cells. Food restriction depressed growth rate by about 50% in both males and females but did not alter baseline plasma B measured at 10weeks in either sex. At the cellular level, food restriction had the following effects: (1) increased basal B production in both sexes and basal ALDO production in males, (2) increased net maximal rates of production of P(4), B, and ALDO in response to ACTH, and (3) no overall effect on adrenocortical cellular sensitivity to ACTH. There were modest sex differences: overall rates of P(4) production were 46% greater in cells from females than from males, and in response to food restriction, the net maximal rate of ALDO production was 50% greater in cells from males than from females. Our results demonstrate that food restriction in S. jarrovii increases adrenocortical cellular rates of steroid production without affecting overall cellular sensitivity to ACTH.     

The influence of sex and gonadectomy on the hypothalamo-pituitary-adrenal axis of the sheep.

There is a sex difference in the hypothalamo-pituitary-adrenal (HPA) axis of many species, although there are sparse data on the sheep. In the present study we have compared the HPA axes of intact and gonadectomised adult male and female sheep at the level of the median eminence, pituitary and adrenal glands using a variety of in vitro approaches. The concentration of arginine vasopressin (AVP) was higher (P<0.01) in the median eminence of male than female sheep, and was also elevated by gonadectomy of either sex (P<0.01). The concentration of corticotrophin-releasing factor (CRF) in the median eminence did not differ between the sexes, but was also elevated in both sexes following gonadectomy (P<0.01). Anterior pituitary pro-opiomelanocortin mRNA concentrations were higher (P<0.05) in intact male sheep than in intact females, with the levels in gonadectomised animals of both sexes being intermediate. In contrast to this finding, basal ACTH secretion from anterior pituitary cells was higher (P<0.05) in cultures derived from female sheep than those from males, but gonadectomy was without effect. There was no effect of sex or gonadectomy on in vitro ACTH secretion in response to AVP, CRF or the combination of AVP and CRF, and in all cases the combination of AVP and CRF generated greater (P<0.0001) ACTH secretion than AVP alone. AVP alone was more effective (P<0.01) than CRF alone as an ACTH secretagogue. The adrenal glands were larger (P<0.05) in female than male sheep, with no effect of gonadectomy. Basal cortisol production was greatest (P<0.05) in cultures of adrenal cells from intact male sheep, though ACTH- and 8BrcAMP-induced cortisol production was greater in the cultures of cells from females (P=0.05); there were no effects of gonadectomy. Cultures of adrenocortical cells from male sheep had greater (P<0.05) basal cAMP production, but ACTH-stimulated cAMP production did not differ between any of the groups of animals. These findings show a range of differences in the HPA axis of male and female sheep. Furthermore, they suggest that the heightened activity of the axis in the female occurs primarily due to differences at the level of the adrenal gland, and that greater adrenal responsiveness of female animals is due to differences in the latter stages of steroidogenesis, rather than an effect on ACTH signal transduction at its receptor.     

Effects of prenatal ethanol exposure on basal limbic-hypothalamic-pituitary-adrenal regulation: role of corticosterone

BACKGROUND: Rats prenatally exposed to ethanol (E) exhibit hypothalamic-pituitary-adrenal (HPA) hyperresponsiveness and changes in central HPA regulation following exposure to stressors. Whether ethanol-induced alterations in basal HPA regulation play a role in mediating HPA hyperresponsiveness remains unclear. We utilized adrenalectomy (ADX), with or without corticosterone (CORT) replacement, to investigate basal HPA function and the role of CORT in mediating ethanol-induced alterations. METHODS: Adult males and females from prenatal E, pair-fed (PF), and ad lib-fed control (C) groups were terminated at the circadian peak, 7 days following sham surgery or ADX, with or without CORT replacement. Plasma levels of CORT and adrenocorticotropin (ACTH), and mRNA levels of corticotropin-releasing hormone (CRH) and arginine vasopressin (AVP) in the paraventricular nucleus, CRH Type 1 receptor (CRH-R1) and pro-opiomelanocortin (POMC) in the anterior pituitary, and mineralocorticoid (MR) and glucocorticoid (GR) receptors in the dorsal hippocampus were determined. RESULTS: Adrenalectomy resulted in significantly greater plasma ACTH elevations in E and PF males, and parallel CRH mRNA elevations in both E and PF males and females compared with their C counterparts. In contrast, pituitary CRH-R1 mRNA levels were lower in E compared with C males, with no differences in POMC. In addition, in response to ADX, E females showed a greater MR mRNA response, and E males showed a greater GR mRNA response compared with their C counterparts, and CORT replacement was ineffective in normalizing ADX-induced alterations in ACTH levels in E and PF females, hippocampal MR mRNA levels in E males, and AVP mRNA levels in PF males and females. CONCLUSIONS: Together, these data indicate that the prenatal ethanol exposure induces HPA dysregulation under basal conditions at multiple levels of the axis, resulting in alterations in both HPA drive and feedback regulation and/or in the balance between drive and feedback. While some effects may be nutritionally mediated, it appears that the mechanisms underlying basal HPA dysregulation may differ between E and PF animals rather than occurring along a continuum of effects on the same pathway. Altered basal HPA tone may play a role in mediating the HPA hyperresponsiveness to stressors observed in E offspring.     

Does adrenal responsiveness vary with sex and reproductive status in Egernia whitii, a viviparous skink?

In mammals, oestrogens generally stimulate adrenal function whilst androgens are inhibitory, and gestating females down-regulate their acute response to stressors in order to protect current reproductive investment. This study aimed to determine if adrenocortical function is similarly modulated by sex and reproductive status in the viviparous lizard, Egernia whitii. We compared the adrenocortical response to acute capture stress in female E. whitii during active (post-ovulatory and gestating) and quiescent (post-partum) phases of their reproductive cycle. We also compared the responses of reproductively quiescent males and females to acute stress and ACTH challenge to determine if there are sex-related differences in HPA axis activity when the influence of reproductive hormones is minimal. The females' responses to acute capture stress varied significantly with reproductive stage, and quiescent females displayed the strongest immediate response, with a rapid and sustained increase in plasma corticosterone (CORT) concentrations. Post-ovulatory females showed the most conservative adrenocortical response and while gestating females showed a large immediate response, this was not as prolonged as in quiescent females. Reproductively quiescent males and females exhibited similar responses to acute stress, and also responded similarly to ACTH injection, with plasma CORT reaching maximal concentrations of 52.1 and 59.4 ng/mL, respectively. Reproductively quiescent females treated with oestrogen exhibited greater responsiveness to ACTH than control females, although basal plasma CORT concentrations were unaltered: these results suggest that the attenuation of the acute stress response observed in reproductively active females of E. whitii may be regulated upstream of ACTH secretion. Our results demonstrate that the activity of the HPA axis is modulated by reproductive status in this viviparous reptile, and that gestating females are able to buffer their embryos from the potentially adverse effects of elevated plasma corticosteroids.     

Effects of breeding season, testosterone and ACTH on the corticosterone response of free-ranging male fence lizards (Sceloporus undulatus)

An attenuated stress response during the breeding season has been reported for several vertebrate species, but the underlying physiological mechanism has received little attention, particularly in reptiles. Modulation could involve changes in the capacity of the adrenal gland to secrete glucocorticoids in addition to upstream changes in the pituitary or hypothalamus. In this study the magnitude of the corticosterone response to capture and confinement was compared between the breeding and postbreeding season in adult male eastern fence lizards, Sceloporus undulatus. Males were captured in both seasons and subjected to the identical stressor of 4 h of confinement. Plasma corticosterone levels in response to confinement were significantly lower in the breeding than the postbreeding season. The effect of testosterone on the stress response was tested by experimentally elevating plasma testosterone levels via silastic implants in free-living males during the postbreeding season. Males with experimentally elevated testosterone exhibited significantly weaker corticosterone responses to 1 h of confinement than sham-implanted males. Finally the capacity of the adrenal glands to secrete corticosterone during the breeding season was tested by challenging males with adrenocorticotropin (ACTH) injections. In spite of naturally suppressed corticosterone responses during the breeding season, males nonetheless responded robustly to ACTH. Altogether these results suggest that modulation resides upstream of the adrenal gland, as has been shown in some arctic-breeding avian species, and likely involves seasonal changes in testosterone levels.     

Gonadectomy-induced reduction of antihypertensive action of angiotensin converting enzyme inhibitor and its role of central renin activity in spontaneously hypertensive rats]

Gonadectomy induced a significant retardation of systolic blood pressure (BP) in male and female spontaneously hypertensive rats (SHR). Captopril (5 mg/kg, i.p.), an angiotensin converting enzyme (ACE) inhibitor, significantly decreased BP in all groups, except that the antihypertensive action was significantly inhibited by gonadectomy in both sexes. The plasma ACE activity was increased in orchiectomized males, but this effect did not appear in females. Neither plasma renin activity nor its concentration were changed by gonadectomy in either sex. Aorta ACE activity was not changed by gonadectomy in either sex, while the renin activity was increased only in gonadectomized females. Diencephalon ACE activity was not changed by gonadectomy in either sex, but the renin activity was decreased in these groups. These results suggested that the decrease of BP by gonadectomy in both sexes SHR was closely related to the decrease of renin activity in diencephalon by androgen deprivation with gonadectomy.     

Corpora lutea and oviposition in the lizard Sceloporus undulatus

Gravid Sceloporus undulatus were either deluteinized or sham-operated. Some of each group were then treated with mammalian FSH or saline. FSH stimulated similar ovarian follicular growth irrespective of the presence or absence of corpora lutea. Deluteinization alone tended to reduce follicular growth and also elicited early oviposition.     

Effect of orchiectomy and testosterone on the early stages of azaserine-induced pancreatic carcinogenesis in the rat

The incidence of carcinoma of the pancreas is higher among men than women. It is also higher among male than female carcinogen-treated rats. The role of testosterone in this preferential induction of pancreatic cancer was evaluated in a rat model of pancreatic carcinogenesis. Two-week-old Lewis rats were treated with a single injection of azaserine. At weaning (3 weeks), rats were divided into five groups as follows: females; intact males; sham-operated males; orchiectomized males; and orchiectomized males plus testosterone. Four months after administration of azaserine, quantitative histologic analysis of atypical acinar cell foci and nodules of the pancreas showed that in female and orchiectomized male rats, foci and nodules were smaller and less numerous than in intact males. Testosterone treatment partly reversed the effect of orchiectomy. This suggests that the susceptibility of male rats to induction of pancreatic carcinomas by azaserine is at least partially mediated by testosterone. Estrogen and testosterone receptors were assayed, but high-affinity receptors characteristic of gonadal tissues were not detected in normal pancreas or in a transplantable azaserine-induced acinar cell carcinoma. Thus, the effect of testosterone in the pancreas may depend on steroid-binding proteins of another type, or may be indirectly mediated.     

Female Rats Release More Corticosterone Than Males in Response to Alcohol: Influence of Circulating Sex Steroids and Possible Consequences for Blood Alcohol Levels

The hypothalamic-pituitary-adrenal (HPA) axis of female rats is more responsive to a variety of stimuli than that of males. Proestrous females are also reported to release more ACTH and corticosterone in response to restraint stress than females at other stages of the estrous cycle. Finally, blood alcohol levels (BALs) reached in response to a standard dose of alcohol also indicate the presence of a gender specificity, with females exhibiting higher BALs than males. The aim of this study was therefore 2-fold: first, we investigated the influence of gender on the ability of alcohol to increase plasma ACTH and corticosterone secretion in the rat. Second, we tested the hypothesis that corticosterone alters alcohol metabolism and asked whether this might represent a mechanism underlying the sex difference in BALs. We observed that compared with intact males, intact females taken at random stages of the estrous cycle secreted significantly (p < 0.01) more ACTH and corticosterone in response to alcohol (0.2-1.8 g/kg). Within females, the intraperitoneal administration of alcohol was followed by higher plasma ACTH and corticosteroids levels during proestrus and estrus, compared with diestrus. Removal of circulating sex steroids abolished the gender difference in terms of ACTH secretion, but ovariectomized females still released more corticosterone than castrated males in response to 0.6 and 1.8 g alcohol/kg. This difference could not be explained by a sex-related component of pituitary responsiveness to corticotropin-releasing factor. These results demonstrate the existence of a sex-specific activation of the HPA axis in response to alcohol, and suggest that sex steroids exert an activational influence on ACTH and corticosterone release in response to ethanol. The possible influence of corticosteroids on the pharmacokinetics of alcohol was investigated in intact males, intact females, and adrenalectomized (ADX) males bearing 15 mg (low dose) or 150 mg (high dose) corticosterone pellets. Following the intraperitoneal injection of a standard dose of ethanol (1.5 g/kg), ADX animals with low corticosterone therapy had higher BALs than either intact rats, or ADX animals with high corticosterone replacement. Intact females exhibited the highest corticosterone levels of all groups of animals, but showed BALs that were intermediate between those of intact males and ADX rats with the low dose corticosterone pellets. Thus, although we cannot exclude an influence of body water content in ADX rats replaced with various regimens of corticosteroids, our results indicate that corticosteroids may modulate alcohol metabolism in the rat. In summary, we have shown that, in the rat, alcohol induces a gender-specific pattern of ACTH and corticosterone secretion that appears to be at least in part dependent on circulating sex steroids. Our results also suggest that although corticosteroids may play a role in regulating the rate of alcohol metabolism, this effect cannot account for the higher BALs measured in female rats.     

Gonadal and sex steroid feedback regulation of gonadotrophin mRNA levels and secretion in neonatal male and female rats

Serum and pituitary LH and FSH, and their pituitary mRNA levels, were measured in neonatal male and female rats after gonadectomy and after gonadectomy with sex steroid replacement. The animals were gonadectomized on day 3 of life, and those given sex steroid replacement were implanted with silicone elastomer capsules containing testosterone for males and diethylstilboestrol for females. Sham-operated rats served as controls. The animals were killed 4 or 8 days latter and the sera and pituitaries collected. Pituitary contents of mRNAs for the alpha subunit, FSH-beta and LH-beta were determined by blot hybridization using corresponding cDNAs. Distinct sex differences were found in the mRNA responses to gonadectomy and steroid replacement. In the males, gonadectomy increased all mRNA levels at 7 days of age. In the females, a rise on day 7 was detected only for FSH-beta; the other mRNAs were increased on day 11 of age. The steroid replacements reversed all the post-gonadectomy increases of mRNAs in both sexes. Moreover, the common alpha and LH-beta mRNAs of the male animals were consistently suppressed below control levels. The serum concentrations of gonadotrophins increased after gonadectomy on day 7 in the males but only on day 11 in the females. The steroid replacements also suppressed the post-gonadectomy increases in serum gonadotrophins, but only the serum concentration of FSH in the females was reduced below controls. Pituitary gonadotrophin concentrations were not affected by gonadectomy, but the steroids suppressed LH in the males and FSH in the females.(ABSTRACT TRUNCATED AT 250 WORDS)     

Postnatal handling does not attenuate hypothalamic-pituitary-adrenal hyperresponsiveness after prenatal ethanol exposure

BACKGROUND: Prenatal ethanol exposure results in hypothalamic-pituitary-adrenal (HPA) hyperresponsiveness to stress in the adult animal. In contrast, an early environmental manipulation, termed "postnatal handling," has been shown to result in decreased and/or less prolonged HPA activity in response to moderate stressors throughout the lifespan of the animal. The effects of both prenatal ethanol exposure and postnatal handling on HPA activity may be mediated by altered feedback regulation of the HPA axis. The present study tested the hypothesis that postnatal handling could attenuate the impact of prenatal ethanol exposure on hormonal responses to stressors. METHODS: Male and female Sprague Dawley rats from prenatal ethanol (E), pair-fed (PF), and ad libitum-fed control (C) groups were either handled (H) or nonhandled (NH) during the preweaning period and were tested at 4 to 5 months of age. Animals were subjected to a 60 min restraint stress, 3 hr after intraperitoneal injection with either saline (SAL) or a synthetic glucocorticoid, dexamethasone-21-phosphate (DEX), in order to examine HPA responsiveness after DEX blockade of endogenous HPA activity. Blood samples were collected via jugular cannulae immediately before restraint (0 min), during restraint (10, 30, and 60 min), and 30 min after the termination of restraint (90 min). RESULTS: For both males and females, DEX administration significantly reduced plasma adrenocorticotropic hormone (ACTH) and corticosterone (CORT) concentrations compared with SAL administration. H animals showed greater suppression of HPA activity (i.e., lower ACTH and/or CORT levels) than NH animals regardless of prenatal group. In addition, E females from both the H and NH treatments showed elevated ACTH and CORT after both SAL and DEX administration, whereas H and NH E males showed elevations in ACTH and CORT only after SAL, compared with their PF and C counterparts. CONCLUSIONS: These data extend results from previous studies that demonstrated HPA hyperresponsiveness in E animals. The finding that E females but not males exhibit elevated ACTH and CORT after DEX administration suggests that prenatal ethanol exposure results in sex-specific alterations of HPA feedback. Consistent with previous data, handling in itself reduced the HPA response to restraint stress. However, handling did not attenuate either HPA hyperresponsiveness or feedback deficits in E animals.     

Effect of periconceptional undernutrition and gender on hypothalamic-pituitary-adrenal axis function in young adult sheep

Glucocorticoids are proposed to act as intermediary factors that transcribe the developmental programming sequelae of maternal nutrient restriction (NR). Periconceptional under-nutrition of sheep markedly activates fetal hypothalamic-pituitary-adrenal (HPA) axis activity leading to preterm birth, while transient undernutrition during late gestation in sheep programs adult HPA axis function. To date, no study has examined resting or stimulated HPA axis function in young adult offspring following a periconceptional nutritional challenge. In the present study, 20 ewes were either periconceptionally undernourished (50% metabolisable energy requirements from days 1 to 30 gestation; NR, n = 8) or fed to control levels (100% requirement; controls, n = 12) to term (147 days gestation). Ewes were blood sampled remotely at 2 and 30 days using automated blood sampling equipment. Thereafter, offspring (controls, n = 6/6 males/females; NR, n = 4/4 males/females) were reared to 1 year of age and on separate days received either an i.v. corticotrophin-releasing hormone (CRH; 0.5 microg/kg) and vasopressin (AVP; 0.1 microg/kg) challenge or a synthetic ACTH i.v. bolus (Synacthen; 1.25 microg/kg), and blood samples were taken (manually and remotely) at appropriate intervals for measurement of plasma ACTH and cortisol accordingly. Resting plasma cortisol, assessed remotely, was similar in ewes during undernutrition (control 18.3 +/- 1.4 vs NR 23.4 +/- 1.9 nmol/l) and in offspring at 4 months of age (control male 17.6 +/- 2.9; control female 17.2 +/- 0.4, NR male 16.5 +/- 3.1, NR female 21.7 +/- 4.0 nmol/l). At 12 months of age, however, resting plasma cortisol was significantly increased in NR females (control male 28.0 +/- 1.5, control female 32.9 +/- 9, NR male 32 +/- 7, NR female 53 +/- 10 nmol/l, F 5.7, P = 0.02) despite no difference in plasma ACTH concentration. There was an interaction between nutritional group and gender for both the pituitary and adrenal responses to CRH and AVP, i.e. for controls, females exhibited increased plasma ACTH or cortisol relative to males but for NR this trend was either not present or reversed. The adrenocortical response to synthetic ACTH was gender-dependent only, being greater in female offspring. Combined CRH and AVP provoked a transient hypertension and marked bradycardia in all animals, irrespective of dietary group or gender and could be effectively reproduced by an AVP bolus alone. In conclusion, the present study has shown that periconceptional undernutrition of sheep has only a minor influence on HPA axis function in their young adult offspring when considered alongside the effect of gender per se.     

The control of adrenocortical secretion in the brush-tailed possum, Trichosurus vulpecula

Circulating levels of corticosteroids and androgens have been studied in the brush-tailed possum (Trichosurus vulpecula) under conditions of tropic hormone stimulation. In both males and anestrous females, levels of androgen (considered to consist largely of testosterone) were elevated by 3 days of treatment with PMS. Further treatment with ACTH (Synacthen) subsequently reduced the level of testosterone in the females to values significantly below the control values, and in the male to a value intermediate between control and PMS stimulated levels. Levels of corticosteroid (considered to consist largely of cortisol) were unaffected by gonadotropin treatment, but significantly increased with ACTH. Dexamethasone treatment greatly decreased cortisol levels, but androgen levels were unaffected in both sexes.In vitro androgen production by the definitive adrenal cortex of the female was significantly decreased by dexamethasone pretreatment. Androgen production by a special hypertrophied adrenocortical zone was unaffected. Corticosteroid formation was reduced in both types of tissue.It is concluded that the adrenal cortex contributes to circulating testosterone levels in the female possum. This is supported by gonadotropin, whereas ACTH has a double effect of stimulating overall steroidogenesis, but switching the relative proportions of the steroids by decreasing the synthesis of androgen and stimulating the corticosteroid pathway. The possible physiological role for this phenomenon and the function of the special hypertrophied zone in the female adrenal cortex are discussed.     

2型糖尿病患者皮质醇分泌和调节变化

目的探讨2型糖尿病（T2DM）患者皮质醇分泌和下丘脑-垂体-肾上腺（HPA）轴活性的改变。方法选取2型糖尿病患者114例（T2DM组）和17例单纯向心性肥胖患者、正常对照80名（NC组）,T2DM组再按腰围分为肥胖组和非肥胖组,比较4组皮质醇分泌水平和HPA轴活性。结果4组患者皮质醇基础水平无差异;用地塞米松后T2DM＋OB组女性患者皮质醇被抑制程度较NC组少（P〈0．05）;与T2DM＋OB组、NC组比较,T2DM组患者肾上腺被兴奋程度最高,尤其在男性（P〈0．05）。结论不同体脂分布、不同性别T2DM患者存在HPA轴活性的差异。     

Variations in Corticosterone Feedback Do Not Reveal Differences in HPA Activity After Prenatal Ethanol Exposure

BACKGROUND: Prenatal ethanol exposure results in hypothalamic-pituitary-adrenal (HPA) hyperresponsiveness to stressors in adult animals. Possible mechanisms mediating this alteration in HPA responsiveness include stress-associated changes in corticosterone (CORT) feedback signals, alterations in CORT signals under basal conditions, and CORT-independent mechanisms. METHODS: We examined the effects of adrenalectomy (ADX) and CORT replacement with a constant, low-level CORT signal via CORT/cholesterol pellets on HPA responses to restraint stress. Adult Sprague-Dawley rats from prenatal ethanol (E), pair-fed (PF), and ad libitum-fed control (C) groups underwent sham ADX (sham), ADX without CORT replacement, or ADX with CORT replacement. Animals were tested during the trough of the circadian rhythm. RESULTS: In the sham condition, E females showed increased adrenocorticotropin hormone (ACTH) and CORT responses to restraint stress compared with C females. Basal and stress-induced ACTH levels were significantly increased in ADX compared with sham animals across all prenatal groups. Constant CORT replacement reduced basal ACTH levels compared with levels in the ADX group, although levels were still increased compared with those observed in the sham group. CORT replacement was minimally effective at reducing ACTH levels during stress. CONCLUSIONS: Although the effects of ADX may have masked possible influences of circadian drive or prenatal group, these findings suggest that in the absence of a CORT feedback signal or in the presence of a constant, low-level CORT feedback signal, E, PF, and C animals do not differ in their abilities to regulate ACTH secretion during the trough of the circadian rhythm.     

Effects of prenatal ethanol exposure on hypothalamic-pituitary-adrenal responses to chronic cold stress in rats

Animals prenatally exposed to ethanol typically exhibit hypothalamic-pituitary-adrenal (HPA) hyperresponsiveness to stressors. In contrast to previous studies that have investigated effects of prenatal ethanol exposure on HPA responses to acute or intermittent stressors, our study investigated HPA responses to a chronic continuous stressor, cold stress (4 degrees C for 0, 1, or 3 days). We tested the hypothesis that prenatal ethanol exposure would result in increased plasma corticosterone (CORT) and adrenocorticotropin (ACTH) responses and increased peptide [corticotropin-releasing factor and vasopressin] mRNA levels in the paraventricular nucleus (PVN) of the hypothalamus compared to that in control animals. In addition, CORT and ACTH responses were measured after exposure to an acute stressor (i.p. isotonic saline injection), superimposed during chronic cold exposure, to examine possible sensitization of the HPA response to the acute stress. Thus, blood samples were collected at the end of each of the three periods of cold exposure, either before (0 min) or 15 min after acute stress. The subjects were adult male and female Sprague-Dawley rat offspring from prenatal ethanol (E), pair-fed (PF), and ad libitum-fed control (C) treatment groups. Exposure to cold stress resulted in significant body weight loss in E males at 1 day and in both males and females of all prenatal treatment groups by 3 days of cold stress. Males in all prenatal groups also exhibited significant increases in adrenal weight:body weight ratios. Cold stress alone (0 min condition) increased CORT levels in E males and overall ACTH levels in E males and females compared to controls. ACTH levels were also higher overall in E compared to control males after acute stress (15 min condition). Sensitization of the CORT response to acute stress was observed in males but not females across all prenatal treatment groups. Corticotropin-releasing factor and vasopressin mRNA levels in the PVN were not significantly affected by prenatal treatment or chronic cold stress in either males or females. In contrast, both males and females displayed increases in PVN thyrotropin-releasing hormone (TRH) mRNA levels after cold stress. These data support and extend previous work demonstrating differential effects of prenatal ethanol exposure on HPA responsiveness of male and female offspring, and suggest that E males may be more vulnerable to the effects of chronic cold stress than E females.     

Effects of fetal ethanol exposure on pituitary-adrenal sensitivity to secretagogues

BACKGROUND: Rodents prenatally exposed to ethanol demonstrate hormonal hyper-responsiveness to stressors in adulthood. The present study examined the hypothesis that an increased sensitivity of the adrenal to ACTH and/or the pituitary to corticotropin releasing hormone (CRH) after dexamethasone suppression, may play a role in the hormonal hyper-responsiveness seen in fetal ethanol-exposed rats. METHODS: Sprague-Dawley males and females from prenatal ethanol-exposed (E), pair-fed (PF), and ad libitum-fed control (C) groups were tested in adulthood (90-120 days). Testing was done in a series of two experiments carried out during the trough of the corticosterone rhythm, the time of greatest sensitivity to feedback inhibition. Twenty-four to 48 hr before testing, jugular cannulae were implanted for hormone infusion and blood sample collection. In both experiments, animals were injected intraperitoneally with dexamethasone-21-phosphate (DEX) (15 microg/100 g body weight for males or 30 microg/100 g body weight for females) 3 hr before testing to suppress endogenous hypothalamic-pituitary-adrenal (HPA) activity. Animals were given a bolus infusion of ACTH (0-0.10 mg/rat) and blood samples (0.2 cc) were drawn at 60-min intervals over 240 min for determination of plasma corticosterone (CORT) levels (Experiment 1), or were given a bolus infusion of CRH (0-20 microg/kg body wt) and samples drawn at 0, 5, 15, and 30 min for determination of plasma ACTH and CORT levels (Experiment 2). RESULTS: As expected, sex differences in adrenal response to ACTH and pituitary response to CRH were observed; females had higher CORT and ACTH levels than males at all concentrations of ACTH and CRH. In addition, dose-response relationships between exogenously administered ACTH or CRH and plasma CORT were demonstrated; increasing concentrations of secretagogues resulted in higher and/or more prolonged CORT responses in both males and females. There were no significant differences among E, PF, and C males or females in adrenal sensitivity to ACTH. However, prenatal ethanol exposure altered pituitary sensitivity to CRH in both males and females. E and PF males demonstrated increased plasma ACTH but not CORT compared with C males, whereas E females demonstrated increased plasma ACTH and CORT levels compared with PF and C females after CRH infusion. CONCLUSIONS: Together these data suggest that (1) E animals do not show increased adrenal sensitivity to ACTH compared with controls; (2) the insult of prenatal ethanol exposure may result in altered pituitary sensitivity to CRH after DEX suppression; and (3) there may be a sex-specific difference in sensitivity of the mechanism(s) underlying HPA hyper-responsiveness.     

Altered ACTH and Corticosterone Responses to Interleukin-1β in Male Rats Exposed to an Alcohol Diet: Possible Role of Vasopressin and Testosterone

We have previously shown that female rats exposed to an alcohol (ethanol, E) diet exhibited a blunted ACTH response to systemically administered interleukin-1β (IL-1β). Because of the presence of gender differences in the activity of the hypothalamic-pituitary-adrenal (HPA) axis, and of the possible role played by sex steroids in modulating the inhibitory influence of E in females, we studied the ability of a 10-day E diet to alter ACTH and corticosterone secretion of intact or castrated male rats injected with IL-1β or endotoxin, a releaser of endogenous cytokines. Pituitary responsiveness to secretagogues that mediate the endocrine effects of IL-1β, namely corticotropin-releasing factor (CRF) and vasopressin (VP), was also investigated. The ACTH responses of animals fed ad libitum (C group) or pair-fed (PF group) to the intravenous administration of IL-1β or endotoxin were not statistically different (p > 0.05); therefore, results from these two groups were combined in the initial experiments. Subsequent experiments only used E and C animals. When compared with this latter group, intact E males showed a significant (p < 0.01) decrease in ACTH levels measured 30 and 60 min after the intravenous injection of IL-β or endotoxin. In contrast, E rats released as much corticosterone as C rats in response to IL-10, but significantly (p < 0.05) more following administration of endotoxin (lipopolysaccha-ride). The stimulatory effect of VP on ACTH release was also measurably blunted by alcohol, whereas that of CRF was not. In none of these experiments were any significant differences observed between C and PF rats. As E rats had markedly lowered circulating testosterone (T) concentrations, and as androgens exerted a restraining effect on the activity of the HPA axis, we then tested the hypothesis that these decreased T values might interact with the inhibitory influence of alcohol. Intact, castrated, or castrated/T-replaced rats fed a control or an alcohol diet all showed comparable pituitary responses to CRF. Alcohol decreased ACTH released in response to IL-β in control and castrated rats. Removal of androgens increased the relative responsiveness of the HPA axis of castrated rats to IL-1β, whether or not they were fed alcohol. In contrast, rats administered T exogenously showed an even larger blunting of their ACTH response to VP, compared with castrated animals without androgen therapy. These results indicate that, as previously shown for females, exposure of male rats to an alcohol diet blunts ACTH released by IL-1β or endotoxin, but not CRF. In addition, we show that activation of the males' pituitary by VP was also blunted by alcohol. Decreased plasma T levels caused by alcohol may participate in the blunted response of the HPA axis to IL-1β, but not to VP. Finally, despite the fact that E rats showed lower plasma ACTH levels than control animals in response to IL-1β, their corticosterone secretion was not proportionally altered. Indeed, adrenal responsiveness of rats injected with endotoxin appeared to be increased by alcohol because similar findings were not obtained upon VP administration. It is possible that, in males, alcohol facilitates a direct stimulatory influence of cytokines on adrenal activity.     

Effect of gonadectomy on HCC development in HBV transgenic mice

Background and aimsEpidemiological data demonstrate that HCC is prevalent in men compared to women. Herein, we examined the effect of gonadectomy in a murine model that spontaneously develops HCC.Animals and methodsThirty-two male and 26 female HBV transgenic mice [Tg (Alb-1 HBV) Bri 44] underwent surgical castration or sham operation. At the 18th month, serum samples were collected and all mice were sacrificed. Liver weight and volume were evaluated, each liver was cut into 1.5-mm-thick consecutive slices and nodules were examined on freshly isolated tissue. Consecutive histological sections obtained from each liver slice were evaluated to confirm the diagnosis of HCC.ResultsSham-operated females showed a significantly lower neoplastic growth compared to sham-operated males. This difference disappeared when females underwent gonadectomy. In males, neoplastic growth was not influenced by gonadectomy. Testosterone and estradiol levels were profoundly modified by gonadectomy in both males and females. The testosterone/estradiol ratio in gonadectomized females increased 4.5-fold compared to that in sham-operated females, becoming more similar to the ratio observed in castrated and sham-operated male mice.ConclusionsHCC growth in our experimental model was not simply influenced by the levels of testosterone or estradiol, taken singularly, but depended on their ratio.