Pituitary and interrenal function in gilthead sea bream (Sparus aurata L., Teleostei) after handling and confinement stress

Dynamics of adrenocorticotropin (ACTH), alpha melanocyte-stimulating hormone (alpha-MSH), N-acetylated-beta-endorphin (N-ac-beta-END), cortisol, and growth hormone (GH) were investigated in gilthead sea bream (Sparus aurata) stressed by handling plus confinement. As indices of the secondary stress response, plasma levels of glucose, lactate, and plasma ions were monitored. Within 1 h, plasma cortisol and ACTH levels increased above the control values but GH levels decreased. Subsequently, at 24 h cortisol and ACTH levels had declined, but were still higher than in controls, whereas GH levels had recovered after 4 h. Regarding the melanotrope peptides, there were no differences in plasma levels of alpha-MSH and N-ac-beta-END, but pituitary stores of these peptides were severely depleted already after 1 h, as were ACTH stores. Pituitary contents of proopiomelanocortin (POMC)-derived hormones did not show significant differences from 72 h onward. Therefore, the results indicate that both handling and confinement affected the corticotropes of the pars distalis and the melanotropes of the neurointermediate lobe but at different magnitudes. The possible involvement of corticotropin-releasing hormone (CRH) in the regulation of pituitary POMC-producing cell types under these conditions was indicated by the in vitro dose-dependent effect of the peptide on release of ACTH, alpha-MSH, and N-ac-beta-END. The corticocotropes appeared more responsive, and approximately 10-fold more sensitive, to CRH compared with the melanotropes. The ACTH-releasing potency of 1 nM CRH was inhibited 75% following pretreatment of the whole pituitary gland with 400 nM of the CRH antagonist alpha-helical CRH(9-41).     

Pituitary proopiomelanocortin-derived peptides and hypothalamus-pituitary-interrenal axis activity in gilthead sea bream (Sparus aurata) during prolonged crowding stress: differential regulation of adrenocorticotropin hormone and alpha-melanocyte-stimulating hormone release by corticotropin-releasing hormone and thyrotropin-releasing hormone

Plasma levels of cortisol, growth hormone (GH), adrenocorticotropin hormone (ACTH), alpha-melanocyte-stimulating hormone (alpha-MSH), N-acetyl-beta-endorphin, in vitro ACTH-stimulated cortisol secretion, and in vitro corticotropin-releasing hormone (CRH)- and thyrotropin-releasing hormone (TRH)-stimulated ACTH and alpha-MSH secretion were investigated in gilthead sea bream exposed to high stocking density (30 kg m(-3)) for 23 days. Within 3 days after the onset of crowding, plasma levels of cortisol, ACTH, alpha-MSH, and N-acetyl-beta-endorphin were above control values. After 7 days, plasma parameters had returned to control levels, but at 23 days, cortisol, alpha-MSH, and N-acetyl-beta-endorphin levels were again elevated over controls, indicating a long-term activation of the melanotrope cells. In contrast, crowding stress elicited a prolonged reduction in plasma GH levels concomitant with the increased hypothalamus-pituitary-interrenal axis (HPI) activation. Crowding stress enhanced cortisol secretory activity of the unstimulated interrenal cells. However, interrenal tissue from crowded fish in vitro displayed an attenuated response to ACTH stimulation compared with tissue from control fish, indicating a desensitization of these cells to ACTH during crowding. The involvement of pituitary proopiomelanocortin-derived peptides in the HPI axis of sea bream is indicated by the observed modulation of the CRH and TRH responsiveness of the corticotropes and melanotropes in crowded fish. At day 1, when there were crowding-induced plasma increases in ACTH and alpha-MSH, there was an attenuated CRH-stimulated but not TRH-stimulated, ACTH release. However, at that time, CRH- and TRH-induced responses of alpha-MSH secretion, and the unstimulated secretory activity of the MSH cells, were enhanced in crowded sea bream. These data provide evidence for stimulatory roles of multiple hypothalamic (CRH and TRH) and pituitary (ACTH and alpha-MSH) peptides in the activation of the hypothalamus-pituitary-interrenal axis under crowding conditions in sea bream.     

The effects of glucose ingestion and fasting on plasma immunoreactive beta-endorphin, adrenocorticotropic hormone and cortisol in obese subjects.

It has been demonstrated that opioid peptides are involved in the stimulation of food intake in rats and that the circulating beta-endorphin levels are increased in genetically obese rodents. Therefore, to assess whether the changes in food intake may influence circulating beta-endorphin levels in obese subjects, plasma beta-endorphin, ACTH and cortisol concentrations were determined in obese patients after an oral glucose load and during a 7-day total starvation. Baseline plasma beta-endorphin concentrations were significantly higher in obese patients than in control normal-weight subjects, while ACTH and cortisol levels were similar in both groups. Plasma beta-endorphin, ACTH and cortisol concentrations were not affected by the ingestion of 75 g glucose, neither were plasma beta-endorphin concentrations modified during prolonged starvation. Moreover, the lack of nycthemeral variations in beta-endorphin levels, documented before and during starvation while plasma ACTH and cortisol were significantly reduced in the evening, suggests that some extra anterior pituitary sources or some obesity-related changes in beta-endorphin metabolism may contribute to the pool of circulating beta-endorphin in obese subjects. On the other hand, even the extreme changes in nutritional conditions, such as total food deprivation or glucose ingestion, are devoid of any detectable influence on circulating beta-endorphin levels.     

Differences between rainbow trout and brown trout in the regulation of the pituitary-interrenal axis and physiological performance during confinement.

The responses of rainbow trout and brown trout to the same stressor were compared by measuring primary and secondary stress responses during and after a 5.5-h net confinement. Basal levels of adrenocorticotropic hormone (ACTH), alpha-melanocyte-stimulating hormone (alpha-MSH), and glucose were higher in brown trout than in rainbow trout. While confinement induced transient increases in plasma ACTH and cortisol levels in both species, the magnitude of these responses, but not the time course, was greater in brown trout. Brown trout, but not rainbow trout, showed a reduction in plasma alpha-MSH levels after 5.5 h confinement before returning to control values, and the glucose levels in the brown trout were elevated throughout the confinement and recovery periods. Confinement also resulted in increased hematocrit values and reduced plasma sodium and chloride levels in both species. Rainbow trout appeared to recover faster from the confinement, as glucose and hematocrit values in the brown trout remained elevated and ionoregulatory disturbances persisted even after 46 h. During recovery effects on the immune system were more pronounced in brown trout than in rainbow trout. Circulating white blood cell numbers were reduced in both species following 23 h recovery, but remained low in the brown trout. Elevated alternative complement activity and oxygen radical production were found after 23 h recovery, and reduced lysozyme activity was found after 46 h, in brown trout only. Results indicate that differences in the stress response of these closely related species, as illustrated by the intensity of the cortisol response, originate at the level of the pituitary and are also manifested through secondary stress responses.     

alpha-MSH acetylation in the pituitary gland of the sea bream (Sparus aurata L.) in response to different backgrounds, confinement and air exposure

MSH is a pituitary hormone derived by post-translational processing from POMC and involved in stress and background adaptation. N-terminal acetylation of MSH to monoacetyl alpha-MSH or diacetyl alpha-MSH increases the bioactivity of the peptide. The aim of this study was to characterize alpha-MSH acetylation in the sea bream (Sparus aurata L.) pituitary gland in response to the stressors air exposure and confinement, as well as in fish adapted for 15 days to a white, gray or black background. Pituitary homogenates were purified by reversed-phase HPLC (RP-HPLC). The alpha-MSH content of fractions was measured by RIA. Immunoreactive RP-HPLC fractions were further analyzed by electrospray mass spectrometry and the peptide sequence determined as SYSMEHFRWGKPV-NH2. In the pituitary gland of sea bream, des-, mono- and diacetyl alpha-MSH were identified. Then plasma alpha-MSH levels were measured in sea bream adapted to different backgrounds. Surprisingly, we found the highest plasma alpha-MSH levels in white-adapted as compared with black-adapted sea bream with intermediate values for gray-adapted fish. This observation is in contrast with results that have been obtained in eel, trout or terrestrial vertebrates. Next, des-, mono- and diacetyl alpha-MSH forms were measured in homogenates of the pituitary gland and in plasma of sea bream exposed to air, to confinement, or to different backgrounds. Monoacetyl alpha-MSH was the predominant form in all control and experimental groups. The lowest content of monoacetyl alpha-MSH relative to des- and diacetyl alpha-MSH was found in white-adapted fish. Levels of des- and diacetyl alpha-MSH forms were similar under all conditions. We observed that monoacetyl alpha-MSH is the most abundant isoform in the pituitary gland after background adaptation, confinement and air exposure, in sea bream. These data indicate that the physiologically most potent isoform of alpha-MSH may vary from species to species.     

Osmoregulatory action of PRL,GH, and cortisol in the gilthead seabream (Sparus aurata L)

The osmoregulatory actions of ovine prolactin (oPRL), ovine growth hormone (oGH), and cortisol were tested in the euryhaline gilthead seabream Sparus aurata. Acclimated to sea water (SW, 40 ppt salinity, 1000 mOsm/kg H(2)O) or brackish water (BW, 5 ppt, salinity, 130 mOsm/kg H(2)O), injected every other day for one week (number of injections, 4) with saline (0.9% NaCl), oPRL (4 microg/g body weight), oGH (4 microg/g body weight) or cortisol (5 microg/g body weight), and transferred from SW to BW or from BW to SW 24h after the last injection. Fish were sampled before and 24h after transfer. Gill Na(+), K(+)-ATPase activity, plasma osmolality, plasma ions (sodium and chloride), plasma glucose, and muscle water moisture were examined. SW-adapted fish showed higher gill Na(+), K(+)-ATPase activity, plasma osmolality, and plasma ions levels than BW-adapted fish. Transfer from SW to BW decreased plasma osmolality and ions levels after 24h, while transfer from BW to SW increased these parameters, whereas gill Na(+),K(+)-ATPase activity was unaffected. oPRL treatment significantly decreased gill Na(+),K(+)-ATPase activity and increased plasma osmolality and ions in SW- and BW-adapted fish. This treatment minimizes loss of osmolality and ions in plasma after transfer to BW and increased these values after transfer to SW. No significant changes were observed in gill Na(+),K(+)-ATPase activity, plasma osmolality, and plasma ions in oGH-treated group with respect to saline group before or after transfer from SW to BW or from BW to SW. Treatment with cortisol induced, in SW-adapted fish, a significant increase of gill Na(+),K(+)-ATPase activity and decrease of plasma osmolality and plasma ions. In BW-adapted fish this treatment induced a significant increases in gill Na(+),K(+)-ATPase activity, plasma osmolality, and plasma ions. After transfer to SW cortisol-treated fish had higher plasma osmolality than the saline group. Our results support the osmoregulatory role of PRL in the adaptation to hypoosmotic environment in the gilthead seabream S. aurata. Further studies will be necessary to elucidate the osmoregulatory role of GH in this species. Cortisol results suggest a "dual osmoregulatory role" of this hormone in S. aurata.     

Interrenal Stress Responsiveness of Tilapia (Oreochromis mossambicus) Is Impaired by Dietary Exposure to PCB 126

Activation of the hypothalamus–pituitary–interrenal (HPI) axis is characteristic of stress responses, which may result from a variety of environmental challenges. To investigate whether the stress response, and in particular the HPI axis, in tilapia (Oreochromis mossambicus) is compromised by short-term exposure to PCB 126, fish of both sexes were fed diets containing PCB 126 (50 μg/kg fish · day) for 5 days. In the first approach, which was performed twice, fish were acutely stressed for periods varying between 1 and 30 min at the end of the exposure period; in the second approach fish were sampled at the end of the exposure period either at rest or after 2 h of stress (confinement). After 5 days, the body weights in all experiments were significantly lower in PCB-fed fish than in control fish. There were no changes in basal plasma glucose levels, plasma ion concentrations, or branchial, renal, and intestinal Na,K-ATPase activity following PCB exposure. In the first experimental approach, in which fish experienced acute sampling stress, plasma cortisol levels reached lower levels in PCB-fed fish than in controls. This suggests an impaired ability to acutely activate interrenal steroidogenesis in PCB-treated tilapia. Adrenocorticotropic hormone (ACTH)- and cAMP-stimulatedin vitrocortisol release from superfused head kidneys was lower in tissues from tilapia exposed to PCB 126 than in tissues from control animals. This effect persisted after 24 hin vitro,which, together with the high PCB 126 concentrations measured in the head kidneys of PCB-fed fish, may indicate direct toxic effects on the interrenal cells. The second experimental approach demonstrated that basal plasma cortisol and ACTH levels were not influenced by PCB treatment, but that the basal ACTH content of the rostral pars distalis (RPD) of the pituitary gland of PCB-fed fish was lower than that of control fish. After 2 h confinement, plasma cortisol levels and ACTH content of the RPD rose to similar values in both groups, whereas plasma ACTH levels were higher in confined PCB-fed fish than in confined controls. PCB-fed fish showed a lower hyperglycemic response to confinement than control fish. Confinement resulted in similarly elevated renal and intestinal Na,K-ATPase activities in both PCB-fed and control fish; branchial enzyme activities were not affected. Since PCB did not affect Na,K-ATPase activities and plasma ion concentrations, it is concluded that the effects of PCB 126 on the HPI axis in tilapia are not secondary to ionoregulatory dysfunction.     

Impaired beta-endorphin response to human corticotropin-releasing hormone in obese children

In order to evaluate the secretion of beta-endorphin in obese children and adolescents, we measured plasma beta-endorphin, ACTH and cortisol levels before and following administration of CRH (1 microgram/kg). Fourteen normal weight and 22 obese subjects (weight excess ranging from 30 to 98%) were studied. Plasma hormone levels were measured by radioimmunoassay directly in plasma (cortisol, ACTH) and after silicic acid extraction and Sephadex G-75 column chromatography (beta-endorphin). Basal beta-endorphin levels in obese children were significantly higher than in controls (14.7 +/- 1.8 vs 6.0 +/- 0.6 pmol/l; mean +/- SEM). No differences were found in basal ACTH and cortisol levels. CRH administration significantly increased beta-endorphin, ACTH and cortisol levels in normal subjects and ACTH and cortisol levels in obese subjects. Plasma beta-endorphin levels in obese children and adolescents did not show any significant increment. These data confirm the higher than normal beta-endorphin plasma levels in obese subjects in childhood and demonstrate that CRH is unable to increase beta-endorphin levels, suggesting an impairment of the hypothalamo-pituitary control mechanisms or an extra-anterior pituitary source.     

The effects of stress on plasma ACTH, alpha-MSH, and cortisol levels in salmonid fishes

Handling and confinement caused a steady increase in the plasma ACTH level in both coho salmon and rainbow trout. Within 2 min plasma ACTH levels had increased significantly, and by 30 min they were 5- to 8-fold higher than the basal ACTH level in unstressed fish. This type of stress also caused a pronounced elevation in plasma cortisol, which lagged behind the ACTH increase, although the degree of change was greater, the level rising between 20- and 50-fold. The plasma alpha-MSH level was unaffected by handling and confinement stress. A second series of experiments assessed the effects of a more severe stress, which consisted of 5 min out of water, during which the fish were restrained, followed by 25-min confinement in a small volume of water. This caused a very rapid, pronounced increase in the plasma ACTH level of sterile rainbow trout, the level reaching a peak at 5 min, and remaining elevated for the next 25 min. Plasma cortisol levels, which were low at the beginning of the experiment, remained so for the first 5 min, and rose thereafter. This type of stress also caused a rapid and pronounced elevation of the plasma alpha-MSH level. It rose in a very similar way, and at the same time, as the plasma ACTH level, but instead of remaining elevated it fell during the 25 min of confinement which followed the 5 min of restraint, to finish one-third of the peak value reached after 5 min.     

Effects of acute behavioral stress on plasma and cerebrospinal fluid ACTH and beta-endorphin in rhesus monkeys

To elucidate the effect of acute behavioral stress on plasma and cerebrospinal fluid (CSF) concentrations of adrenocorticotropic hormone (ACTH) and beta-endorphin, rhesus monkeys were subjected to 30 min of confinement stress. Simultaneous plasma and CSF samples revealed no significant change in CSF ACTH or beta-endorphin up to 120 min after the onset of the stress despite significant elevations in plasma cortisol, ACTH, and beta-endorphin. It is suggested that acute behavioral stress does not alter CSF ACTH or beta-endorphin, and that this information may be clinically useful for future human studies of CSF ACTH and beta-endorphin in neuropsychiatric illnesses.     

Plasma cortisol concentrations in Atlantic salmon, Salmo salar: episodic variations, diurnal change, and short term response to adrenocorticotrophic hormone

Short term episodic peaks 1-2 hr in duration in plasma cortisol levels were observed in adult Atlantic salmon. These results support the concept that episodic secretion of corticosteroids may be a characteristic feature of the vertebrate adrenal/interrenal regulatory system and provide an explanation for the variability frequently observed in plasma cortisol concentrations in salmonids. Plasma cortisol levels also varied throughout the light-dark cycle and mean concentrations were higher during the night. High frequency variations in plasma cortisol levels (with a time scale of minutes rather than hours) were not observed. Porcine adrenocorticotrophic hormone (ACTH) and ACTH1-24 but not alpha-melanocyte-stimulating hormone (alpha-MSH) increased plasma cortisol levels 10 min after injection. The change in plasma cortisol levels after injection of 1 mU/kg porcine ACTH was of similar magnitude and duration as endogenous episodic variations. Plasma cortisol levels during episodic peaks, circadian changes, or after ACTH injection were less than the binding capacity of the "high" affinity protein binding system for cortisol. Hence, the unbound cortisol concentration is a linear function of the total cortisol concentration. Whereas episodic release of cortisol is indicated here, the physiological importance of this mode of hormone secretion is unclear. Plasma glucose levels were unaffected up to 2 hr after injection of ACTH and did not show close temporal correlation with endogenous episodic peaks in plasma cortisol.     

Effects of mammalian ACTH on potential fuels and gluconeogenic substrates in the plasma of the spiny dogfish shark (Squalus acanthias).

The effects of mammalian ACTH on plasma beta-hydroxybutyrate, glucose, alanine, and lactate levels were determined in the unfed spiny dogfish shark (Squalus acanthias). Serial blood samples were collected from a cannula implanted in the dorsal aorta. The metabolite levels were estimated by standard enzymatic procedures for 4 days before treatment and for 4 days after treatment. Plasma glucose, alanine, and lactate, but not beta-hydroxybutyrate, declined after the surgery, but the metabolites were relatively stable for at least 48 hr before treatment. ACTH (40 units/kg) or the control solution were infused via the cannula after the morning blood sample on postoperative Day 4 and again after the 12-hr blood sample. Control plasma beta-hydroxybutyrate, glucose, and lactate did not change significantly, but alanine levels increased approximately 29% by 96 hr when sampling ended. Plasma beta-hydroxybutyrate levels were not significantly changed by ACTH treatment. Plasma glucose levels increased approximately 36% by 3 hr after ACTH infusion, remained elevated following the second ACTH treatment, and then declined to approximately the initial levels by 96 hr. Plasma alanine levels increased approximately 53% by 0.75 hr after ACTH treatment, were still approximately 15% greater than the initial levels by 12 hr, rose again after the second ACTH infusion at 12 hr, and then declined to near the control levels by 96 hr. Plasma lactate levels increased approximately 107% by 1.5 hr after ACTH infusion and then decreased to approximately 22% greater than the initial levels by 12 hr. Lactate levels increased after the second ACTH infusion and remained approximately 58% greater than the initial levels when sampling ended at 96 hr. The results indicate that the pituitary-adrenocortical axis hormones do not directly influence plasma beta-hydroxybutyrate levels. However, the data support the suggestion that the hyperglycemic action of the pituitary-adrenocortical axis hormones is mediated, at least in part, by the provision of alanine and lactate substrates for gluconeogenesis.     

Plasma beta-endorphin levels in primary aldosteronism

Excessive production of an as yet unidentified aldosterone-stimulating factor may cause idiopathic hyperaldosteronism (IHA). This putative factor may be related to proopiomelanocortin-derived peptides, some of which have aldosterone-stimulating properties. The present study evaluated plasma beta-endorphin, ACTH, cortisol, and aldosterone levels in patients with IHA (n = 10), aldosterone-producing adenomas (n = 4), essential hypertension (n = 11), and normal subjects (n = 10). Plasma and urinary hormone measurements were obtained at timed intervals during an isocaloric, fixed electrolyte intake (Na+, 128 meq/day; K+, 80 meq/day) in a metabolic unit. Plasma for beta-endorphin assay was preincubated with sepharose-bound anti-beta-lipotropin to remove beta-lipotropin that cross-reacted with the beta-endorphin RIA. Mean +/- SE plasma beta-endorphin levels at 0800 h were elevated in IHA patients (47 +/- 13 fmol/ml) compared to those in aldosterone-producing adenoma (25 +/- 9), essential hypertension (16 +/- 1), and normal control (20 +/- 2; P less than 0.05) subjects. Plasma ACTH, plasma cortisol, and urinary cortisol levels were not different in these four groups. These data support the hypothesis that excess production of either beta-endorphin or related proopiomelanocortin-derived peptides may function as aldosterone secretogogue(s) in IHA.     

Divergence of endocrine and metabolic responses to stress in two rainbow trout lines selected for differing cortisol responsiveness to stress

Rainbow trout (Oncorhynchus mykiss) of two lines selected for low (LR) and high (HR) cortisol stress-responsiveness were subjected to confinement for a period of 336 h. Endocrine (plasma cortisol, hepatic cortisol binding) and metabolic (plasma glucose, lactate, amino acids; hepatic glycogen and alanine aminotransferase levels) indices of stress were measured at intervals in confined and unconfined fish of both lines. During confinement plasma cortisol concentration reached maximum values earlier in HR fish (2h) than in LR fish (6h) returning to control values within 336 h in both lines. Paradoxically, although both HR and LR lines displayed a characteristic metabolic stress response, these changes were more pronounced in LR fish. Plasma glucose and lactate levels increased during confinement in both lines but to a significantly greater extent in LR fish. Confinement significantly elevated plasma amino acids to a greater extent in LR fish than in HR fish. Liver glycogen concentration was depleted most rapidly in LR fish but was significantly higher in confined fish of both lines than controls at the end of the experiment. No significant changes were observed in hepatic alanine aminotransferase activity during confinement. Confined fish of both lines displayed a decrease in hepatic cortisol receptor abundance within 24h and this was more sustained in HR fish. The more pronounced disturbance of a broad range of indicators of stress in confined LR fish, compared to HR fish, throws doubt on the magnitude of the cortisol response being the primary driver of these differences.     

Ontogeny of corticotropin-releasing factor and of hypothalamic-pituitary-interrenal axis responsiveness to stress in tilapia (Oreochromis mossambicus; Teleostei)

The ontogeny of the corticotropin-releasing factor (CRF) system and of the ability of the hypothalamic-pituitary-interrenal (HPI) axis to respond to stressors (capture or confinement), or to cortisol treatment was investigated in tilapia (Oreochromis mossambicus). In 2 days post hatching (dph) larvae, the first developmental stage used for immunohistochemistry, CRF-immunoreactivity (ir) was observed in the nucleus preopticus (npo), and in two hypothalamic nuclei (nlt and nrl). In this stage, CRF- and AVT-ir was found in the neural part of the pituitary, and endocrine cells in the pars distalis and pars intermedia contained POMC-derived peptides. In the ventral telencephalon, CRF-ir cells were first observed 5 dph, whereas projections from these cells into the anterior part of the latero-dorsal telencephalon (Dla) from 7 dph onwards. CRF, ACTH, alpha-MSH, and cortisol were quantified by radioimmunoassays in homogenates of the anterior-cranial region of the larvae containing brain, pituitary, and headkidneys. CRF contents increased from 43 +/- 3 to 1070 +/- 70 pg/larvae between 5 and 110 dph. Larvae of age 5, 12, 24, and 42 dph were captured sequentially from a group. All life stages were able to rapidly increase their cortisol content in response to this stressor (ANOVA: P < 0.001). Overall, the developmental stage affected cortisol content (ANOVA: P < 0.001), but developmental stage did not influence the cortisol reaction to stress (ANOVA: P > 0.162). Whole brain CRF content did not change during the 20 min stress period and the relationship between CRF-producing neurons and the initial HPI stress response in early life stages remains to be established. Cortisol feeding of 18 and 29 dph larvae for periods ranging from 2 to 24 days resulted in elevations of the CRF content (P < 0.003) in comparison to controls. In 18 dph larvae cortisol feeding abolished the cortisol response to capture stress as observed in control fed larvae (P < 0.008). We propose that cortisol induced upregulation of CRF takes place in the telencephalon and is restricted to a time period during larval development, characterised by the absence of glucocortoid receptor (GR) expression in the telencephalic Dm region in these larvae. Finally, the stress response to 24 h confinement was compared between saltwater adapted and freshwater adapted juveniles (age 77 dph). Confinement stress (24 h) affected cortisol and CRF content (ANOVA: P < 0.001, P < 0.008, respectively), but not ACTH content. Interactions were observed between salinity and confinement regarding cortisol and alpha-MSH contents (ANOVA: P < 0.02), but not regarding CRF and ACTH contents. The increase in cortisol levels induced by confinement was remarkably high in freshwater adapted larvae (five times higher than in saltwater adapted larvae). Regarding the cortisol response it is concluded that during and after the period of mouth breeding tilapia larvae respond to capture stress in a similar fashion (onset and height) as adults. Previously, we reported that the initial plasma cortisol response to capture stress in adult tilapia occurred independently from changes in plasma ACTH levels. The current finding that also brain CRF contents do not alter during the initial cortisol response in larvae further indicates that the initial cortisol response in this species may be regulated independently from CRF and ACTH.     

β-Endorphin and adrenocortical function in obesity

OBJECTIVE: To test the hypothesis that the hyperendorphinaemia in obesity originates from outside the pituitary. DESIGN: Intravenous administration of corticotrophin-releasing hormone (CRH) after overnight suppression with 2 mg of dexamethasone in normal-weight controls and in obese subjects before and after weight reduction. PATIENTS: Eleven obese females, age (mean +/- SEM) 30 +/- 2.1 years, body mass index (BMI) 41.2 +/- 1.9 kg/m2. Eight normal-weight females served as controls, age 26 +/- 2.1 years, BMI 21.4 +/- 0.5 kg/m2. Five obese subjects were also studied after weight loss of 18.4 +/- 1.0% of original weight. MEASUREMENTS: Plasma beta-endorphin, ACTH and cortisol. Cortisol production rate in 24-hour urine. Basal (without dexamethasone suppression) plasma beta-endorphin levels. RESULTS: Basal (without dexamethasone suppression) beta-endorphin levels were 7.7 +/- 0.8 pmol/l in the obese and 3.8 +/- 0.5 pmol/l in the control subjects (P less than 0.005). The degree of suppression of beta-endorphin after dexamethasone was similar in the obese (23.2 +/- 3.7%) and in the control subjects (28.2 +/- 0.12%). Administration of CRH following dexamethasone suppression resulted in a small but significant increase of plasma beta-endorphin in both obese (from 1.55 +/- 0.12 to 2.32 +/- 0.28 pmol/l) and control subjects (from 0.98 +/- 0.24 to 1.69 +/- 0.33 pmol/l). The groups did not differ regarding this response, nor regarding the release of ACTH and cortisol after CRH. Cortisol production rate was higher (P less than 0.001) in the obese (68.7 +/- 3.3 mumol/24 h) than in the controls (40.0 +/- 3.0 mumol/24 h). No correlation between cortisol production rate and basal beta-endorphin levels was found. Weight loss appeared to have no influence on cortisol production rate, basal beta-endorphin levels, or on the responses to dexamethasone or CRH. CONCLUSIONS: Plasma beta-endorphin in obese subjects can be affected by manipulations of the hypothalamic-pituitary-adrenocortical axis; the hypothesis that the hyperendorphinaemia of obesity originates from outside the pituitary cannot be confirmed.     

The acute stress response of red porgy, Pagrus pagrus, kept on a red or white background

The skin colour of red porgy, Pagrus pagrus, can be modified by exposure to different background colours. Red and white background colours brighten the dark skin colour that develops under common culture conditions in red porgy. To assess whether skin colour is also modified by aquaculture related handling stress, we subjected red porgy to 5 min of netting stress combined with air exposure. Fish kept on a white background have: (1) a lighter skin colour, which is not influenced by an acute stressor, (2) a less saturated red colour, which significantly decreases 24h post-handling, and (3) a similar hue as fish kept on a red background. The first plasma parameters to rise after application of the stressor are cortisol, lactate and Na(+); then, glucose levels rose. Other plasma ions (Ca(2+), Cl(-), K(+)) were not affected up to 2h post-stressor, but had decreased at 8 and 24h after handling. Plasma pH decreased over the first 2h post-handling, indicative of plasma acidosis upon air exposure. The acidosis then coincided with increases in plasma lactate levels. As alphaMSH levels were not significantly affected by the stressor while cortisol levels showed a five to tenfold increase, we suggest that following acute stress in red porgy, plasma cortisol release is controlled by ACTH, perhaps in combination with a sympathic stimulation.     

Infusion of beta-endorphin has no suppressive effect on the releasing hormone-stimulated pituitary-adrenal-axis of normal human subjects

The existence of a short-loop feedback inhibition of pituitary ACTH release by administration of beta-endorphin was postulated. However, data on the effect of peripherally administered beta-endorphin in humans are highly controversial. We infused human synthetic beta-endorphin at a constant rate of 1 microgram.kg-1.min-1 or normal saline to 7 normal volunteers for 90 min. Thirty min after starting the beta-endorphin or placebo infusion, releasing hormones were injected as a bolus iv (oCRH and GHRH 1 microgram/kg, GnRH 100 micrograms, TRH 200 micrograms) and blood was drawn for measurements of beta-endorphin immunoreactivity, all other pituitary hormones, and cortisol. Infusion of beta-endorphin resulted in high beta-endorphin plasma levels with a rapid decrease after the infusion was stopped. During the control infusion, beta-endorphin plasma levels rose in response to CRH. Plasma ACTH and serum cortisol levels in response to the releasing hormone were not different in subjects infused with beta-endorphin or placebo. The PRL response to TRH was significantly higher after beta-endorphin than after placebo (area under the stimulation curve 1209 +/- 183 vs 834 +/- 104 micrograms.l-1.h). There was no difference in the response of all other hormones measured. Our data on ACTH and cortisol secretion do not support the concept of a short-loop negative feedback of beta-endorphin acting at the site of the pituitary.     

Immunoreactive alpha-MSH and ACTH levels in rat plasma and pituitary.

A radioimmunoassay is described for the measurement of alpha-melanocyte-stimulating hormone (alpha-MSH). The antibody was produced in rabbits by immunization with alpha-MSH coupled to bovine serum albumin with carbodiimide. The antibody did not react significantly with ACTH, beta-MSH, or 6 fragments of ACTH. The sensitivity and reliability of the assay were improved by employing a simple plasma extraction procedure. When applied to a 2 ml plasma sample, the detection limit of the radioimmunoassay was 6 pg/ml. ACTH was measured with a sensitive and specific radioimmunoassay previously described for humans and adapted for the rat. The anti-ACTH serum cross-reacted with the biologically active portion of alpha-p ACTH and not with alpha-MSH, beta-MSH or the alpha-p 17-39 and alpha-p 25-39 fragments of ACTH. The detection limit was 20 pg/ml. Plasma and pituitary alpha-MSH and ACTH had the same immunoreactivity as synthetic alpha-MSH and ACTH. alpha-MSH and ACTH contents of the rat neurointermediate lobe were 1398 +/- 360 (SE) ng and 28.2 +/- 2.9 ng, respectively, while in the anterior lobe they were 102 +/- 31 ng and 551 +/- 36 ng, respectively. The plasma alpha-MSH concentration at 8 AM in male rats was 64 +/- 8 pg/ml when the plasma ACTH concentration was 92 +/- 15 pg/ml. Over a 24-hour period two peaks of plasma alpha-MSH were observed, one at 4 AM (142 +/- 35 pg/ml) and the other at 4 PM (139 +/- 26 pg/ml). Plasma ACTH was higher at noon (151 +/- 43 pg/ml) and 4 PM (130 +/- 48 pg/ml). Short-term exposure to ether induced a transient increase in alpha-MSH level 5 min later and a rapid return to normal levels. Plasma ACTH increased significantly 2.5 min after the onset of ether stress and remained high for 30 min. Two hours' exposure to ether did not change plasma alpha-MSH, although a 3-fold increase in plasma ACTH was observed. Haloperidol injection was followed by a large increase in plasma alpha-MSH, whereas ACTH levels increased similarly after saline and Haloperidol injection. Corticoid administration reduced ACTH, but not alpha-MSH. Three weeks after adrenalectomy, alpha-MSH levels had not changed but ACTH levels had increased ten-fold. These data indicate that alpha-MSH is secreted in the rat, and that the regulation of its secretion is different from that of ACTH.     

Plasma profiles of adrenocorticotropic hormone, cortisol, alpha-melanocyte-stimulating hormone, and growth hormone in dogs with pituitary-dependent hyperadrenocorticism before and after hypophysectomy

The 6-h plasma profiles of adrenocorticotropic hormone (ACTH), cortisol, alpha-melanocyte-stimulating hormone (alpha-MSH), and GH were studied in 17 dogs with pituitary-dependent hyperadrenocorticism (PDH) before and after hypophysectomy. The aim of the study was to investigate the relation between the hormone profile characteristics and recurrence of PDH after surgery. The hormones were secreted in a pulsatile fashion. The basal plasma cortisol concentration and area under the curve (AUC) for cortisol were significantly higher in the PDH cases than in eight controls. The characteristics of the plasma profiles of ACTH and alpha-MSH were not significantly different between the PDH cases and the controls. In the PDH cases, less GH was secreted in pulses than in the controls, but the difference was not significant. The basal plasma cortisol concentration, the AUC for ACTH and cortisol, and the pulse frequency of ACTH and cortisol decreased significantly after hypophysectomy for the group of PDH cases. The basal plasma concentrations of ACTH and alpha-MSH, the AUC for alpha-MSH, and the characteristics of the plasma GH profiles of the PDH cases remained unchanged after hypophysectomy. No pulses of alpha-MSH were observed after hypophysectomy. The co-occurrence between the ACTH and cortisol pulses decreased significantly with hypophysectomy. The postoperative pulse frequency of ACTH was the only characteristic with predictive value for the recurrence of PDH after hypophysectomy. The results of this study demonstrate that ACTH, cortisol, alpha-MSH, and GH are secreted in a pulsatile fashion in dogs with PDH. Hypophysectomy effectively reduces the secretion of ACTH and cortisol. The presence of ACTH pulses after hypophysectomy is a risk factor for the recurrence of hyperadrenocorticism.