Early life corticosteroid overexposure: Epigenetic and fetal origins of adult diseases.

The relationship between events occurring during intrauterine development and later-life predisposition to long-term disease, has been described. The fetus responds to excess intrauterine exposure to high levels of corticosteroids, modifying their physiological development and stopping their growth. Fetal exposure to elevated levels of either endogenous (alterations in fetal hypothalamic-pituitary-adrenal axis) or synthetic corticosteroids, is one model of early-life adversity; to developing adult disease. At the molecular level, there are transcriptional changes in metabolic and growth pathways. Epigenetic mechanisms participate in transgenerational inheritance, not genomic. Exposures that change 11ß-hydroxysteroid dehydrogenase type 2 enzyme methylation status in the placenta can result in transcriptional repression of the gene, causing the fetus to be exposed to higher levels of cortisol. More precise diagnosis and management of antenatal corticosteroids for preterm birth, would potentially decrease the risk of long-term adverse outcomes. More studies are needed to understand the potential roles of factors to alter fetal corticosteroid exposure. Long-term infant follow-up is required to determine whether methylation changes in placenta may represent useful biomarkers of later disease risk. This review, summarize recent advances in the programming of fetal effects of corticosteroid exposure, the role of corticosteroids in epigenetic gene regulation of placental 11ß-hydroxysteroid dehydrogenase type 2 enzyme expression and transgenerational effects.

The Role of the Adrenal-Gut-Brain Axis on Comorbid Depressive Disorder Development in Diabetes.

Diabetic patients are more affected by depression than non-diabetics, and this is related to greater treatment resistance and associated with poorer outcomes. This increase in the prevalence of depression in diabetics is also related to hyperglycemia and hypercortisolism. In diabetics, the hyperactivity of the HPA axis occurs in parallel to gut dysbiosis, weakness of the intestinal permeability barrier, and high bacterial-product translocation into the bloodstream. Diabetes also induces an increase in the permeability of the blood-brain barrier (BBB) and Toll-like receptor 4 (TLR4) expression in the hippocampus. Furthermore, lipopolysaccharide (LPS)-induced depression behaviors and neuroinflammation are exacerbated in diabetic mice. In this context, we propose here that hypercortisolism, in association with gut dysbiosis, leads to an exacerbation of hippocampal neuroinflammation, glutamatergic transmission, and neuronal apoptosis, leading to the development and aggravation of depression and to resistance to treatment of this mood disorder in diabetic patients.

Patterns of poverty across adolescence predict salivary cortisol stress responses in Mexican-origin youths.

Poverty is a chronic stressor associated with disruptions in psychophysiological development during adolescence. This study examined associations of chronic poverty and income changes experienced from pre- to mid-adolescence with hypothalamic-pituitary-adrenal (HPA) axis stress responses in late adolescence. Participants (N = 229) were adolescents of Mexican-origin (48.7% female). Household income (converted to income-to-needs ratios) was assessed annually when children were 10-16 years old. At 17 years, adolescents completed Cyberball, a social exclusion simulation task while undergoing a functional magnetic resonance imaging scan. Saliva samples were collected prior to and five times over a 50-minute period following the scan, from which salivary cortisol was assayed. Results showed that differential trajectories of poverty from ages 10-16 predicted HPA axis activity at age 17. Relative to others, distinct HPA suppression (hyporeactivity) was demonstrated by youth who started adolescence in deep poverty and were still living in poverty at age 16 despite experiencing some income gains. Youth from more economically secure families evinced typical cortisol increases following the lab stressor. These results suggest that subsequent HPA functioning varies as a function of economic status throughout adolescence, and that efforts to increase family income may promote healthy HPA functioning for youths in the most impoverished circumstances.

Nociceptin/orphanin FQ receptor system blockade as an innovative strategy for increasing resilience to stress.

The ability to successfully cope with stress is known as 'resilience', and resilient individuals are less prone to develop psychopathologies. Understanding the neurobiological mechanisms of resilience may be instrumental to improve current therapies and benefit high-risk subjects. This review summarizes the complex interplay that exists between physiological and pathological responses to stressful events and the nociceptin/orphanin FQ (N/OFQ) - N/OFQ receptor (NOP) system, including: the effects of stress in regulating N/OFQ release and NOP expression; the ability of the N/OFQ-NOP system to modulate the hypothalamic-pituitary-adrenal axis; behavioral studies; and evidence in humans correlating this peptidergic system with psychopathologies. Available findings support the view that N/OFQ signaling stimulates the hypothalamic-pituitary-adrenal axis, thus increasing stress circulating hormones and corticotropin-releasing factor signaling. Additionally, activation of the NOP receptor inhibits monoamine transmission, including 5-HT, and this may contribute to maladaptive outcomes of stress. Ultimately, the N/OFQ system seems to have an important role in stress vulnerability, and blockade of NOP signaling may provide an innovative strategy for the treatment of stress related psychopathologies.

The influence of hypertensive environment on adipose tissue remodeling measured by fluorescence lifetime imaging in spontaneously hypertensive rats.

There is a lack of information correlating low adiposity with hypertension experienced by Spontaneous Hypertensive Rats (SHR) or overweight and normotension in Wistar-Kyoto (WKY). We aimed to investigate this lipodystrophy phenomenon by measuring fluorescence lifetime (FLIM), optical redox ratio (ORR), serum levels of hypothalamic-pituitary-adrenal (HPA) and/or hypothalamic-pituitary-thyroid (HPT) hormones axes between Wistar, WKY and SHR before and after establishment of hypertension. Under high blood pressure, we evaluated serum adipokines. Brown adipose tissue was characterized as lower ORR and shorter FLIM compared to white adipose tissue. HPT axis showed a crucial role in the SHR adipose tissue configuration by attenuating whitening. The increased adiposity in WKY may act as a preventive agent for hypertension, since SHR, with low adiposity, establishes the disease. The hypertensive environment can highlight key adipokines that may result in new therapeutic approaches to the treatment of adiposity dysfunctions and hypertension.

The NR3C1 gene expression is a potential surrogate biomarker for risk and diagnosis of posttraumatic stress disorder.

Posttraumatic Stress Disorder (PTSD) is an anxiety disorder which occurs after a traumatic event. The NR3C1 gene codes for the Glucocorticoid Receptor, which participate in the Hypothalamic-Pituitary-Adrenal (HPA) axis and is altered in PTSD patients. To evaluate whether the NR3C1 gene expression in peripheral blood could be useful as a diagnosis biomarker, a total of 32 PTSD patients and 59 healthy controls were analyzed with quantitative RT-PCR. Also, to assess if NR3C1 dysregulation is associated with hypocortisolism in PTSD patients, serum cortisol was quantified by ELISA in a subset of these samples. Significant NR3C1 over-expression was found in PTSD patients compared with controls, and this was higher in patients with acute PTSD. The Area Under the Curve (AUC) of NR3C1 gene expression was 0.797. The sensibility and specificity of NRC1 gene expression to diagnose PTSD was 62.5% and 89.8%, respectively. We also found that an up-regulation of NR3C1 increased the risk for being diagnosed with PTSD (OR= 12.8, 95%, CI 4-41.4). Finally, the NR3C1 gene expression was inversely related with serum cortisol in PTSD patients. The present results suggest that NR3C1 gene expression could be a promising biomarker for PTSD diagnosis and estimate the risk for disease development.

Differential antipredatory responses in the tuco-tuco (Ctenomys talarum) in relation to endogenous and exogenous changes in glucocorticoids.

Glucocorticoids participate in the behavioral and physiological responses generated under stressful circumstances coming from different sources-physical and/or psychological. In mammals, the increases of these hormones are mediated by the activation of the hypothalamic-pituitary-adrenal axis. This response occurs after exposure to novel and unpredictable situations that lead to the loss of homeostasis, for example, a direct encounter with predators or their cues. However, the relationship between the physiological and behavioral responses is still a complex issue in vertebrates. We evaluate the effects of an experimental manipulation of glucocorticoid levels on the generation of the behavioral and physiological response to stress by predation in the subterranean rodent C. talarum. We found that when tuco-tucos encountered predator cues-fur odor, and largely, immobilization-they responded physiologically by secreting cortisol. This response was accompanied by an associated behavioral response. However, when the increase in plasma cortisol originated exogenously by the injection of cortisol, a behavioral response was not observed. Finally, inhibition of glucocorticoids' synthesis was effective in weakening the behavioral effects produced by immobilization. In conclusion, in tuco-tucos, predator cues act as stress factors that trigger differential increases in plasma cortisol and a behavioral response associated with the appearance of anxiety states.

Physiological responses of dogs to different housing systems

Background: The hypothalamic-pituitary-adrenal (HPA) axis undergoes adaptations throughout housing system that mightcontribute to the avoidance of adverse effects of welfare status in dogs housed in a shelter. Nevertheless, the influence ofhousing systems and stabling time on glucose and PCV changes is little known. The purpose of the present study was toevaluate the patterns of cortisol, glucose and PCV in dogs housed in a kennel and normal environments, evaluating thedifferences between housing systems, by taking into account the different stabling time and sex.Materials, Methods & Results: The study comprised 98 cross-breed dogs, aged 4 ± 1.5 years, lodged in a kennel (observational group I: N=61, 29 females and 27 males), in paired household dogs (control group II: N=25, 13 females and 12males) and in unpaired household dogs (control group III: N=12, 6 females and 6 males). Females of both groups werespayed. The subjects were studied on the basis of different stabling times, ranged among <1 year, 2 years and 4 years, anddifferent sex.Discussion: This observational study showed that kennelled males lodged for 2 (P < 0.01) and 4 (P < 0.001) years showedlower cortisol concentrations than males lodged <1 year; males lodged for <1 year (P < 0.001) showed higher cortisol concentrations than females; males lodged for 4 year showed lower cortisol concentrations (P < 0.01) than females. Kennelledfemales lodged for 4 year showed higher PCV values (P < 0.001) than females lodged for <1 year. Paired and unpairedhousehold females and males lodged for 4 years showed lower cortisol concentrations (P < 0.01) than 2 years and <1 year.Paired and unpaired household females and males lodged for short-, medium- and long-term times showed higher glucoseconcentrations (P < 0.001) than kennelled dogs. This study showed significant changes of circulating cortisol, glucoseand PCV...(AU)

Clinical and biochemical characteristics of high-intensity functional training (HIFT) and overtraining syndrome: findings from the EROS study (The EROS-HIFT).

The metabolic and hormonal consequences of high-intensity functional training regimens such as CrossFit® (CF) are unclear. Little is known about the triggers and clinical and biochemical features of CF-related overtraining syndrome (OTS). The EROS study compared endocrine and metabolic responses, and eating, social, psychological and body characteristics of OTS-affected (OTS) and healthy athletes (ATL), and non-physically active controls (NPAC). The current study is a post-hoc analysis of the CF subgroups of the EROS study, to evaluate specific characteristics of CF in ATL and OTS. Parameters were overall and pairwise compared among OTS-affected (CF-OTS) and healthy (CF-ATL) athletes that exclusively practiced CF, and NPAC. CF-ATL yielded earlier and enhanced cortisol, GH, and prolactin responses to an insulin tolerance test (ITT), increased neutrophils, lower lactate, increased testosterone, improved sleep quality, better psychological performance, increased measured-to-predicted basal metabolic rate (BMR) ratio and fat oxidation, and better hydration, when compared to NPAC. Conversely, more than 90% of the adaptive changes in CF were lost under OTS, including an attenuation of the hormonal responses to an ITT, increased estradiol, decreased testosterone, and decreased BMR and fat oxidation; the most remarkable trigger of OTS among "HIFT athletes" was the long-term low carbohydrate and calorie intake.

Glucocorticoids and sympathetic neurotransmitters modulate the acute immune response to Trypanosoma cruzi.

Evidence suggests that natural and adaptive immune responses can trigger neuroendocrine responses. Here, we discuss changes in the activity of the hypothalamus-pituitary-adrenal axis and in autonomic nerves, predominantly of the sympathetic nervous system, in a mouse model of acute infection with Trypanosoma cruzi. The endocrine response includes a marked increased release of glucocorticoid and a decrease of immune-stimulatory hormones, such as dehydroepiandrosterone sulfate, prolactin, and growth hormone during infection. These endocrine changes result in reduced proinflammatory cytokine production, increased regulatory/effector T cell ratio, and thymus atrophy. The sympathetic activity in the spleen of infected mice is also markedly reduced. However, the residual sympathetic activity can modulate the immune response to the parasite, as shown by increased mortality and production of proinflammatory cytokines in sympathetically denervated, infected mice. The outcome of the neuroendocrine response is the moderation of the intensity of the immune response to the parasite, an effect that results in delayed mortality in susceptible mice, and favors the course toward chronicity in more resistant animals.

Physiological responses of dogs to different housing systems

Background: The hypothalamic-pituitary-adrenal (HPA) axis undergoes adaptations throughout housing system that mightcontribute to the avoidance of adverse effects of welfare status in dogs housed in a shelter. Nevertheless, the influence ofhousing systems and stabling time on glucose and PCV changes is little known. The purpose of the present study was toevaluate the patterns of cortisol, glucose and PCV in dogs housed in a kennel and normal environments, evaluating thedifferences between housing systems, by taking into account the different stabling time and sex.Materials, Methods & Results: The study comprised 98 cross-breed dogs, aged 4 ± 1.5 years, lodged in a kennel (observational group I: N=61, 29 females and 27 males), in paired household dogs (control group II: N=25, 13 females and 12males) and in unpaired household dogs (control group III: N=12, 6 females and 6 males). Females of both groups werespayed. The subjects were studied on the basis of different stabling times, ranged among <1 year, 2 years and 4 years, anddifferent sex.Discussion: This observational study showed that kennelled males lodged for 2 (P < 0.01) and 4 (P < 0.001) years showedlower cortisol concentrations than males lodged <1 year; males lodged for <1 year (P < 0.001) showed higher cortisol concentrations than females; males lodged for 4 year showed lower cortisol concentrations (P < 0.01) than females. Kennelledfemales lodged for 4 year showed higher PCV values (P < 0.001) than females lodged for <1 year. Paired and unpairedhousehold females and males lodged for 4 years showed lower cortisol concentrations (P < 0.01) than 2 years and <1 year.Paired and unpaired household females and males lodged for short-, medium- and long-term times showed higher glucoseconcentrations (P < 0.001) than kennelled dogs. This study showed significant changes of circulating cortisol, glucoseand PCV...

Growth hormone and cortisol secretion in the elderly evaluated using the glucagon stimulation test.

OBJECTIVES: The glucagon stimulation test is a reliable alternative test to assess growth hormone and cortisol secretion, but has not been widely used in the elderly population. The aim of this study was to evaluate growth hormone and cortisol secretion using the glucagon stimulation test in an elderly population without known hypothalamic-pituitary disease and to correlate growth hormone and cortisol peaks with age (less than or greater than 80 years) and body mass index. METHODS: Forty-two subjects (67-88 years) from the geriatric ambulatory unit were submitted and 41 subjects completed the glucagon stimulation test. RESULTS: Median growth hormone peak was 5.99 µg/L and median cortisol peak was 21.6 µg/dL. Growth hormone peak was >3 µg/L in 73.2%, and cortisol peak was >18 µg/dL in 65.8% of patients. There was a statistically significant positive correlation between the growth hormone peak and the cortisol peak. The cortisol peak was significantly different between subjects stratified by growth hormone peak of < or >3 µg/L (15.7 and 21.8 µg/dL, respectively). There was a statistically significant difference in cortisol peak according to age < or > 80 years (22.4 and 18.5 µg/dL, respectively). Considering lower cut-offs recently proposed for growth hormone peak (1.0 µg/L for overweight subjects) and cortisol peak (9.1 µg/dL), only two patients had a growth hormone peak below this value, and all patients had preserved cortisol secretion. CONCLUSIONS: We did find a positive correlation between growth hormone and cortisol peaks in the glucagon stimulation test in the elderly, confirming the capacity of the glucagon stimulation test to stimulate both axes. According to the new proposed cut-points for growth hormone and cortisol, we had 95% of normal growth hormone and 100% of normal cortisol responses.

Dopamine D2-like receptors modulate freezing response, but not the activation of HPA axis, during the expression of conditioned fear.

Considering the complexity of aversive information processing and defensive response expression, a combined action of stress modulators may be required for an optimal performance during threatening situations. Dopamine is now recognized as one of the most active modulators underlying states of fear and anxiety. On the other hand, activation of hypothalamic-pituitary-adrenocortical (HPA) axis, which leads to the release of corticosterone in rodents, has been considered a key part of the stress response. The current study is an extension of prior work investigating modulatory effects of dopamine and corticosterone on conditioned fear expression. We have showed that corticosterone, acting through mineralocorticoid receptors in the ventral tegmental area (VTA), upregulates dopaminergic system in the basolateral amygdala (BLA), enabling the expression of conditioned freezing response. The novel question addressed here is whether VTA-BLA dopaminergic signaling is necessary for increases in corticosterone during conditioned fear expression. Using site-specific treatment with D2-like agonist quinpirole (VTA) and D2-like antagonist sulpiride (BLA), we evaluated freezing and plasma corticosterone in rats exposed to a light used as aversive conditioned stimulus (CS). Intra-VTA quinpirole and intra-BLA sulpiride significantly decreased freezing expression in the conditioned fear test, but this anxiolytic-like effect of the dopaminergic drugs was not associated with changes in plasma corticosterone concentrations. Altogether, data suggest that interferences with the ability of the CS to activate the dopaminergic VTA-BLA pathway reduce the expression of freezing, but activation of the HPA axis seems to occur upstream of the recruitment of dopaminergic mechanisms in conditioned fear states.

The Role of the Microbiome in the Relationship of Asthma and Affective Disorders.

The effect of stress, anxiety and other affective states on inflammatory conditions such as asthma is well documented. Although several immune pathway mechanisms have been proposed and studied, they cannot fully explain the relationship. In this chapter we present a new perspective on asthma development and exacerbation that integrates findings on the role of psychological factors in asthma with the microbiome and the hygiene hypothesis in asthma development.

Chlorella vulgaris reduces the impact of stress on hypothalamic-pituitary-adrenal axis and brain c-fos expression.

Predominantly emotional stressors activate a wide range of brain areas, as revealed by the expression of immediate early genes, such as c-fos. Chlorella vulgaris (CV) is considered a biological response modifier, as demonstrated by its protective activities against infections, tumors and stress. We evaluated the effect of acute pretreatment with CV on the peripheral and central responses to forced swimming stress in adult male rats. Pretreatment with CV produced a significant reduction of stress-related hypothalamic-pituitary-adrenal activation, demonstrated by decreased corticotrophin releasing factor gene expression in the hypothalamic paraventricular nucleus (PVN) and lower ACTH response. Hyperglycemia induced by the stressor was similarly reduced. This attenuated neuroendocrine response to stress occurred in parallel with a diminished c-fos expression in most evaluated areas, including the PVN. The data presented in this study reinforce the usefulness of CV to diminish the impact of stressors, by reducing the HPA response. Although our results suggest a central effect of CV, further studies are necessary to understand the precise mechanisms underpinning this effect.

Positive modulation of a neutral declarative memory by a threatening social event.

Memories can be altered by negative or arousing experiences due to the activation of the stress-responsive sympatho-adrenal-medullary axis (SYM). Here, we used a neutral declarative memory that was acquired during multi-trial training to determine the effect of a threatening event on memory without emotional valence. To this end, participants received a new threatening social protocol before learning pairs of meaningless syllables and were tested either 15 min, 2 days or 8 days after acquisition. We first demonstrated that this threatening social situation activates not only the SYM axis (Experiment 1) and the hypothalamus-pituitary-adrenal axis (HPA; Experiment 2), but also, it improves the acquisition or early consolidation of the syllable pairs (Experiment 3). This improvement is not a transient effect; it can be observed after the memory is consolidated. Furthermore, this modulation increases the persistence of memory (Experiment 4). Thus, it is possible to affect memories with specific events that contain unrelated content and a different valence.

Food restriction negatively affects multiple levels of the reproductive axis in male house finches, Haemorhous mexicanus.

Nutrition influences reproductive functions across vertebrates, but the effects of food availability on the functioning of the hypothalamic-pituitary-gonadal (HPG) axis in wild birds and the mechanisms mediating these effects remain unclear. We investigated the influence of chronic food restriction on the HPG axis of photostimulated house finches, Haemorhous mexicanus. Food-restricted birds had underdeveloped testes with smaller seminiferous tubules than ad libitum-fed birds. Baseline plasma testosterone increased in response to photostimulation in ad libitum-fed but not in food-restricted birds. Food availability did not, however, affect the plasma testosterone increase resulting from a gonadotropin-releasing hormone-I (GnRH) or a luteinizing hormone (LH) challenge. The number of hypothalamic GnRH immunoreactive (ir) but not proGnRH-ir perikarya was higher in food-restricted than in ad libitum-fed finches, suggesting inhibited secretion of GnRH. Hypothalamic gonadotropin-inhibitory hormone (GnIH)-ir and neuropeptide Y (NPY)-ir were not affected by food availability. Plasma corticosterone (CORT) was also not affected by food availability, indicating that the observed HPG axis inhibition did not result from increased activity of the hypothalamic-pituitary-adrenal (HPA) axis. This study is among the first to examine multilevel functional changes in the HPG axis in response to food restriction in a wild bird. The results indicate that food availability affects both hypothalamic and gonadal function, but further investigations are needed to clarify the mechanisms by which nutritional signals mediate these effects.

Neuroendocrine and viral correlates of premature immunosenescence.

Aging continuously remodels the immune system, a process known as immunosenescence. Here, we review evidence of premature immunosenescence in younger individuals under conditions of chronic psychological stress, chronic inflammation, or exposure to certain persistent viral infections. Chronic stress may accelerate various features of immunosenescence by activating key allostatic systems, notably the hypothalamic-pituitary-adrenal axis and increased cortisol levels. Chronic stress is associated with thymic involution, blunted T cell proliferation, increased serum proinflammatory markers, and shorter telomere lengths. Human cytomegalovirus (CMV) infection has been implicated in accelerating immunosenescence by shrinking the T cell receptor repertoire and causing clonal expansion of senescent CD8(+) CD28(-) T cells with a proinflammatory profile. These factors increase inflammation associated with aging, or "inflammaging," particularly as it relates to etiology of several age-related diseases and increased mortality. Patients with rheumatoid arthritis have been shown to have several signatures of premature immunosenescence, including expansion of senescent T cells associated with cognitive impairment. We end by speculating that bipolar disorder can be considered as a model of accelerated aging because it has been associated with shortened telomeres, higher CMV IgG titers, and expansion of senescent and regulatory T cells.

Regulation of TRH neurons and energy homeostasis-related signals under stress.

Energy homeostasis relies on a concerted response of the nervous and endocrine systems to signals evoked by intake, storage, and expenditure of fuels. Glucocorticoids (GCs) and thyroid hormones are involved in meeting immediate energy demands, thus placing the hypothalamo-pituitary-thyroid (HPT) and hypothalamo-pituitary-adrenal axes at a central interface. This review describes the mode of regulation of hypophysiotropic TRHergic neurons and the evidence supporting the concept that they act as metabolic integrators. Emphasis has been be placed on i) the effects of GCs on the modulation of transcription of Trh in vivo and in vitro, ii) the physiological and molecular mechanisms by which acute or chronic situations of stress and energy demands affect the activity of TRHergic neurons and the HPT axis, and iii) the less explored role of non-hypophysiotropic hypothalamic TRH neurons. The partial evidence gathered so far is indicative of a contrasting involvement of distinct TRH cell types, manifested through variability in cellular phenotype and physiology, including rapid responses to energy demands for thermogenesis or physical activity and nutritional status that may be modified according to stress history.

Morning serum Basal cortisol levels are affected by age and pubertal maturation in school-aged children and adolescents.

BACKGROUND: The same reference values for cortisol have been used for adults and children, but laboratory results obtained from an adult population might not be suitable for pediatric patients. AIMS: To determine morning serum basal cortisol levels in children and adolescents. METHODS: The study was conducted on 120 suitable reference individuals, healthy Brazilian children and adolescents from both genders aged 4-19 years old. The method used for cortisol assessment was the chemiluminescent enzyme immunoassay (VITROS® 5600 MicroWell; Johnson & Johnson, High Wycombe, UK, 2009), and the kit reference interval was 4.46-22.7 µg/dl (122.7-626.2 nmol/l). The setting limits were calculated according to the Clinical Laboratory Standards Institute guidelines. RESULTS: The morning serum basal cortisol levels increased with age and pubertal maturation, but there were no differences based on gender. The cortisol reference values were established based on the 2.5th and 97.5th percentiles as 2.97 µg/dl [81.9 nmol/l, 90% confidence interval (CI) 1.44-3.69 µg/dl] and 23.4 µg/dl (645.5 nmol/l; 90% CI 16.3-26.4 µg/dl), respectively. CONCLUSIONS: The assessment of the morning serum basal cortisol levels showed a unique pattern, with a different lower limit for the cohort compared to current values established for adult subjects.