Psychosocial Crowding Stress-Induced Changes in Synaptic Transmission and Glutamate Receptor Expression in the Rat Frontal Cortex.

This study demonstrates how exposure to psychosocial crowding stress (CS) for 3, 7, and 14 days affects glutamate synapse functioning and signal transduction in the frontal cortex (FC) of rats. CS effects on synaptic activity were evaluated in FC slices of the primary motor cortex (M1) by measuring field potential (FP) amplitude, paired-pulse ratio (PPR), and long-term potentiation (LTP). Protein expression of GluA1, GluN2B mGluR1a/5, VGLUT1, and VGLUT2 was assessed in FC by western blot. The body's response to CS was evaluated by measuring body weight and the plasma level of plasma corticosterone (CORT), adrenocorticotropic hormone (ACTH), and interleukin 1 beta (IL1B). CS 3 14d increased FP and attenuated LTP in M1, while PPR was augmented in CS 14d. The expression of GluA1, GluN2B, and mGluR1a/5 was up-regulated in CS 3d and downregulated in CS 14d. VGLUTs expression tended to increase in CS 7d. The failure to blunt the effects of chronic CS on FP and LTP in M1 suggests the impairment of habituation mechanisms by psychosocial stressors. PPR augmented by chronic CS with increased VGLUTs level in the CS 7d indicates that prolonged CS exposure changed presynaptic signaling within the FC. The CS bidirectional profile of changes in glutamate receptors' expression seems to be a common mechanism evoked by stress in the FC.

Forms of selenium in vitamin-mineral mixes differentially affect the expression of genes responsible for prolactin, ACTH, and α-MSH synthesis and mitochondrial dysfunction in pituitaries of steers grazing endophyte-infected tall fescue.

The goal of this study was to test the hypothesis that sodium selenite (inorganic Se, ISe), SEL-PLEX (organic forms of Se, OSe), vs. a 1:1 blend (MIX) of ISe and OSe in a basal vitamin-mineral (VM) mix would differentially alter pituitary transcriptome profiles in growing beef steers grazing an endophyte-infected tall fescue (E+) pasture. Predominately Angus steers (BW = 183 ± 34 kg) were randomly selected from fall-calving cows grazing E+ pasture and consuming VM mixes that contained 35 ppm Se as ISe, OSe, or MIX forms. Steers were weaned, depleted of Se for 98 d, and subjected to summer-long common grazing of a 10.1 ha E+ pasture containing 0.51 ppm ergot alkaloids. Steers were assigned (n = 8 per treatment) to the same Se-form treatments on which they were raised. Selenium treatments were administered by daily top-dressing 85 g of VM mix onto 0.23 kg soyhulls, using in-pasture Calan gates. As previously reported, serum prolactin was greater for MIX (52%) and OSe (59%) steers vs. ISe. Pituitaries were collected at slaughter and changes in global and selected mRNA expression patterns determined by microarray and real-time reverse transcription PCR analyses, respectively. The effects of Se treatment on relative gene expression were subjected to one-way ANOVA. The form of Se affected the expression of 542 annotated genes (P < 0.005). Integrated pathway analysis found a canonical pathway network between prolactin and pro-opiomelanocortin (POMC)/ACTH/α-melanocyte-stimulating hormone (α-MSH) synthesis-related proteins and that mitochondrial dysfunction was a top-affected canonical pathway. Targeted reverse transcription-PCR analysis found that the relative abundance of mRNA encoding prolactin and POMC/ACTH/α-MSH synthesis-related proteins was affected (P < 0.05) by the form of Se, as were (P ≤ 0.05) mitochondrial dysfunction-related proteins (CYB5A, FURIN, GPX4, and PSENEN). OSe steers appeared to have a greater prolactin synthesis capacity (more PRL mRNA) vs. ISe steers through decreased dopamine type two receptor signaling (more DRD2 mRNA), whereas MIX steers had a greater prolactin synthesis capacity (more PRL mRNA) and release potential by increasing thyrotropin-releasing hormone concentrations (less TRH receptor mRNA) than ISe steers. OSe steers also had a greater ACTH and α-MSH synthesis potential (more POMC, PCSK2, CPE, and PAM mRNA) than ISe steers. We conclude that form of Se in VM mixes altered expression of genes responsible for prolactin and POMC/ACTH/α-MSH synthesis, and mitochondrial function, in pituitaries of growing beef steers subjected to summer-long grazing an E+ pasture.

Hypothalamic and pituitary clusterin modulates neurohormonal responses to stress.

Clusterin is a sulfated glycoprotein abundantly expressed in the pituitary gland and hypothalamus of mammals. However, its physiological role in neuroendocrine function is largely unknown. In the present study, we investigated the effects of intracerebroventricular (ICV) administration of clusterin on plasma pituitary hormone levels in normal rats. Single ICV injection of clusterin provoked neurohormonal changes seen under acute stress condition: increased plasma adrenocorticotropic hormone (ACTH), corticosterone, GH and prolactin levels and decreased LH and FSH levels. Consistently, hypothalamic and pituitary clusterin expression levels were upregulated following a restraint stress, suggesting an involvement of endogenous clusterin in stress-induced neurohormonal changes. In the pituitary intermediate lobe, clusterin was coexpressed with proopiomelanocortin (POMC), a precursor of ACTH. Treatment of clusterin in POMC expressing AtT-20 pituitary cells increased basal and corticotropin-releasing hormone (CRH)-stimulated POMC promoter activities and intracellular cAMP levels. Furthermore, clusterin treatment triggered ACTH secretion from AtT-20 cells in a CRH-dependent manner, indicating that increased clusterin under stressful conditions may augment CRH-stimulated ACTH production and release. In summary, hypothalamic and pituitary clusterin may function as a modulator of neurohormonal responses under stressful conditions.

Fetal glucocorticoid synthesis is required for development of fetal adrenal medulla and hypothalamus feedback suppression.

During pregnancy, fetal glucocorticoid is derived from both maternal supply and fetal secretion. We have created mice with a disruption of the Cyp11a1 gene resulting in loss of fetal steroid secretion but preserving the maternal supply. Cyp11a1null embryos have appreciable although lower amounts of circulating corticosterone, the major mouse glucocorticoid, suggesting that transplacental corticosterone is a major source of corticosterone in fetal circulation. These embryos thus provide a means to examine the effect of fetal glucocorticoids. The adrenal in Cyp11a1 null embryos was disorganized with abnormal mitochondria and oil accumulation. The adrenal medullary cells did not express phenylethanolamine N-methyltransferase and synthesized no epinephrine. Cyp11a1 null embryos had decreased diencephalon Hsd11b1, increased diencephalon Crh, and increased pituitary Pomc expression, leading to higher adrenocorticotropin level in the plasma. These data indicate blunted feedback suppression despite reasonable amounts of circulating corticosterone. Thus, the corticosterone synthesized in situ by the fetus is required for negative feedback suppression of the hypothalamus-pituitary-adrenal axis and for catecholamine synthesis in adrenal medulla.

The effects of ghrelin/GHSs on AVP mRNA expression and release in cultured hypothalamic cells in rats.

Ghrelin, the endogenous ligand for growth hormone secretagogues (GHSs) receptor (GHS-R), increases adrenocorticotropin (ACTH) and cortisol (corticosterone) as well as GH secretion in humans and animals. However, the site of GHSs action to induce ACTH secretion is not fully understood. To clarify the mechanisms of the action of ghrelin/GHSs on ACTH secretion, we analyzed the effects of KP-102 and ghrelin on the mRNA expression and release of corticotropin releasing factor (CRF) and arginine vasopressin (AVP), ACTH secretagogues, in monolayer-cultured hypothalamic cells of rats. Incubation of cells with KP-102 for 4h and 8h and with ghrelin for 4h significantly increased AVP mRNA expression and release without changing CRF mRNA expression. CRF levels in culture media were undetectable. Suppression of GHS-R expression by siRNA blocked ghrelin- and KP-102-induced AVP mRNA expression and release. NPY significantly increased AVP mRNA expression and release. Furthermore, treatment of cells with anti-NPY IgG blocked KP-102-induced AVP mRNA expression and release. We previously reported that KP-102 significantly increases NPY mRNA expression in cultured hypothalamic cells. Taken together, these results suggest that ACTH secretion by ghrelin/GHSs is induced mainly through hypothalamic AVP, and that NPY mediates the action of ghrelin/GHSs.

Stress decreases the ability to resist smoking and potentiates smoking intensity and reward.

We have developed a novel human laboratory model to examine two primary aspects of stress-precipitated tobacco relapse: (1) Does stress reduce the ability to resist the first cigarette? (2) Once the first cigarette is initiated, does stress facilitate subsequent smoking? Using a within-subject design, daily smokers (n = 37) who were nicotine deprived overnight received a personalized imagery induction (stress or neutral) on two separate days, and then had the option of initiating a tobacco self-administration session or delaying initiation for up to 50 min in exchange for three levels of monetary reinforcement. Subsequently, the tobacco self-administration session entailed a 1-hour period in which subjects could choose to smoke using a smoking topography system. Following the stress induction, subjects were less able to resist smoking, smoked more intensely (increased puffs, shorter inter-puff interval, and greater peak puff velocity), and perceived greater satisfaction and reward from smoking. Stress significantly increased hypothalamus-pituitary-adrenal (HPA) axis reactivity, tobacco craving, negative emotion, and physiologic reactivity relative to the neutral condition. In addition, increased cortisol, ACTH, and tobacco craving were associated with reduced ability to resist smoking following stress. These findings have implications for understanding the impact of stress on smoking relapse and model development to assess smoking lapse behavior.

Impact of loudness dependency of auditory evoked potentials on the panic response to CCK-4.

RATIONALE: Experimental panic induction with cholecystokinin-tetrapeptide (CCK-4) has been established as a model to study the pathophysiology of panic disorder. In line with the serotonin (5-HT)-hypothesis of panic disorder it has been suggested that the panicogenic effects of CCK-4 are mediated in part through the 5-HT system. The analysis of the loudness dependency of the auditory evoked potentials (LDAEP) is a valid non-invasive indicator of central serotonergic activity. METHODS: We investigated the correlation between LDAEP and behavioral, cardiovascular and neuroendocrine panic responses to CCK-4in 77 healthy volunteers and explored whether differences in LDAEP paralleled subjective panic severity. Behavioral panic responses were measured with the panic symptom scale (PSS). Heart rate and ACTH/cortisol plasma concentrations were assessed concomitantly. RESULTS: LDAEP did not differ between panickers and nonpanickers. Furthermore, LDAEP did not correlate with the behavioral panic response. However, a significant positive correlation between LDAEP and CCK-4 induced HPA-axis activation, which was uniform in panickers and nonpanickers, could be detected. CONCLUSIONS: The psychological effects of CCK-4 rather are mediated by neurotransmitters others than the endogenous 5-HT system. However, the extent of the neuroendocrine activation related to the CCK-4 panic provocation was correlated with the LDAEP, thereby suggesting that central 5-HT mechanisms are involved in the HPA-axis activation during this challenge paradigm.

[Adrenopause]. / Adrenopauza.

The hypothalamic-pituitary-adrenal axis activity in the aging people is characterised by an unexplained reduction of dehydroepiandrosterone (DHEA) and its sulfate (DHEAS) secretion while ACTH and cortisol production remains relatively unchanged. This decline in the biological activity of the zona reticularis, referred to as 'adrenopause', may contribute to the physiology of human aging. The reduced endogenous concentrations of DHEA and DHEAS found in advancing age have been correlated with a constellation of health problems. Because these steroids seem to play a role in the maintenance of immunity, musculoskeletal integrity, and cardiovascular health, age-associated declines in adrenal androgen production may lead to decreased immune function, osteoporosis, and atherosclerosis. Despite clear benefits of DHEA administration in patients with adrenal insufficiency, the results of DHEA supplementation in healthy euadrenal subjects are not so clear-cut. Studies assessing its action on sexual function, metabolism and cardiovascular functions have provided conflicting results. This paper summarises the present state of knowledge on the age-related changes in adrenal androgen production and discusses pros and cons of DHEA use in older people.

Effects of neuropeptides B and W on the rat pituitary-adrenocortical axis: in vivo and in vitro studies.

Neuropeptides (NP) B and W are hypothalamic peptides involved in the regulation of feeding and neuro-endocrine axes. Evidence has been provided that NPB and NPW act on both the central and the peripheral branches of the rat hypothalamic-pituitary-adrenocortical axis, and we carried out in vivo and in vitro studies to gain insight into this topic. Reverse transcription-polymerase chain reaction showed the expression of NPB, NPW and their receptors in both adrenal cortex (zonae glomerulosa and fasciculata-reticularis) and adrenal medulla, where immunocytochemistry also detected the presence of abundant NPB- and NPW-immunoreactivity. The acute subcutaneous administration of NPB (0.5 or 1.5 nmol/100 g) did not alter ACTH plasma concentration, while that of NPW (1.5 nmol/100 g) decreased it. Neither NPB nor NPW affected the blood level of aldosterone, while both peptides (0.5 nmol/100 g) raised that of corticosterone. NPB (10(-6) M) lowered ACTH-stimulated aldosterone secretion, and basal and ACTH-stimulated corticosterone production from adrenal quarters containing both cortical and medullary tissues. NPW (10(-6) M) enhanced basal aldosterone secretion from adrenal quarters, and the effect was suppressed by the beta-adrenoceptor antagonist l-alprenolol (10(-5) M). NPW did not affect corticosterone production. Collectively, our findings allow us to draw the following tentative conclusions: i) ACTH-independent extra-adrenal mechanism(s) are operative in vivo, by which NPB and NPW stimulate adrenal glucocorticoid, but not mineralocorticoid secretion; ii) in vitro the interaction of NPB with adrenal medulla activates unknown mechanism(s) hampering adrenocortical steroidogenic machinery; and iii) NPW stimulates in vitro aldosterone secretion by enhancing the release of medullary catecholamines, which in turn activate beta-adrenoceptors located on zona glomerulosa cells.

Effect of prolonged exposure to low concentrations of formaldehyde on the corticotropin releasing hormone neurons in the hypothalamus and adrenocorticotropic hormone cells in the pituitary gland in female mice.

We examine the effect on the hypothalamus-pituitary-adrenal gland (HPA) axis of prolonged exposure to low levels of formaldehyde in female C3H/He mice, using immunocytochemical and RT-PCR methods. Two groups of female mice were exposed to differing concentrations (0, 80, 400, 2000 ppb) of formaldehyde inhalation for 16 h/day, 5 days/week, for 12 weeks. The corticotropin releasing hormone (CRH)-immunoreactive (ir) neurons in the hypothalamus were then examined, together with the adrenocorticotropin hormone (ACTH)-ir cells and ACTH mRNA in the pituitary. One group comprised sham control mice. The other group was made allergic by injection of ovalbumin (OVA) and alum prior to exposure to formaldehyde, since most sick building syndrome (SBS) sufferers are women with allergic disease. These animals were further exposed to aerosolized OVA as a booster four times during the exposure period. Our results showed a dose-dependent increase in the number of CRH-ir neurons in the non-allergy (NAG) group. A similar pattern was found in ACTH-ir cells and ACTH mRNA. The allergy (AG) model group showed an increase in basal levels of all markers of HPA activity. Moreover, the AG mice appeared to respond to the lowest concentration of formaldehyde, and all indices of HPA activity were reduced at the highest concentrations of formaldehyde. These results relate to an important clinical issue and also have implications in the broader area of HPA regulation. We conclude that our experimental system may be a suitable animal model for SBS and/or multiple chemical sensitivity (MCS).

[Effect of jingui shenqi pill on pituitary adrenocorticotropic hormone gene expression in shen-yang deficiency rats].

OBJECTIVE: To explore the effect of Jingui Shenqi pill (JGSQP) with various concentrations at different time points on pituitary adrencorticotropic hormone (ACTH) gene expression level in Shen-Yang deficiency rats. METHODS: The Shen-Yang deficiency rats were randomly divided into the model control group and the high, medium and low dosage of JGSQP groups. Reverse transcriptase polymerase chain reaction was used to observe the effect of JGSQP on the ACTH mRNA of pituitary tissue in rats treated at different time points (10 d, 20 d, 30 d). RESULTS: As compared with that in the model group, the ACTH gene expression level was significantly higher in the high dose JGSQP group (P < 0.05), and the increment in the medium dosage group was significantly higher in comparing with that in the high and low dosage groups (P < 0.05 or P < 0.01). CONCLUSION: Through up-regulation on ACTH gene expression is possibly one of the mechanisms of JGSQP in treating Shen-Yang deficiency.

Effect of Hypericum extract on the hypothalamic-pituitary-adrenal system in rats.

We studied the effect of Hypericum extract on activity of the hypothalamic-pituitary-adrenal system in rats. In rats exposed to stress after a 30-day daily oral treatment with Hypericum extract, the weight of the adrenals and ACTH concentration were lower than in controls. Hence, treatment with Hypericum extract improved resistance to stress and prevented exhausting of the hypothalamic-pituitary-adrenal system.

T-lymphocyte activation increases hypothalamic and amygdaloid expression of CRH mRNA and emotional reactivity to novelty.

Stimulation of T-cells with staphylococcal enterotoxin B (SEB) significantly elevates interleukin-2 (IL-2) and contemporaneous activation of the hypothalamic-pituitary-adrenal (HPA) axis and c-fos in the paraventricular nucleus (PVN) of BALB/cByJ mice. Such neural signaling may promote cognitive and emotional adaptation before or during infectious illness. Because corticotropin-releasing hormone (CRH) is an anxiogenic neuropeptide that may mediate the stressor-like effects of immunological stimuli, we measured neuronal CRH mRNA alterations in mice challenged with SEB. Increased CRH mRNA levels were observed in the PVN and central nucleus of the amygdala (ceA) 4-6 hr after SEB administration. This was associated with plasma ACTH increases, which could be abrogated by the systemic administration of anti-CRH antiserum. Additional experiments did not support a role for IL-2 or prostaglandin synthesis in activating the HPA axis. Behavioral experiments testing for conditioned taste aversion did not confirm that SEB challenge promotes malaise. However, consistent with the notion that central CRH alterations induced by SEB may affect emotionality (e.g., fear), SEB challenge augmented appetitive neophobia in a context-dependent manner, being marked in a novel and stressful environment. It is hypothesized that immunological stimuli generate a cascade of events that solicit integrative neural processes involved in emotional behavior. As such, these data support the contention that affective illness may be influenced by immunological processes and the production of cytokines and are consistent with other evidence demonstrating that autoimmune reactivity is associated with enhanced emotionality.

Hypothalamic oxytocin in the developing ovine fetus: interaction with pituitary-adrenocortical function.

Oxytocin (OT) stimulates corticotroph function in adult sheep, however, there is little information on OT synthesis and its potential involvement in hypothalamo-pituitary-adrenal (HPA) function in the fetus. The objectives of this study were to examine developmental changes in hypothalamic OT synthesis and to investigate the actions of OT on fetal corticotroph function. Hypothalami were removed at various stages of pre- and post-natal development. OT mRNA levels were measured using in situ hybridization. For in vitro studies, fetal pituitaries were removed on days 129 and 138 of gestation. Anterior pituitary cells were dispersed and cells were treated with different concentrations and combinations of OT, corticotrophin-releasing hormone (CRH), vasopressin (AVP) and cortisol. OT mRNA was present in the paraventricular nucleus (PVN) and supraoptic nucleus (SON) by day 60 of gestation, and levels significantly increased at term. OT mRNA was present in parvocellular and magnocellular fields of the PVN. In vitro, OT stimulated adrenocorticotropin (ACTH) output in a dose-dependent fashion, but had no effect on cellular pro-opiomelanocortin (POMC) mRNA levels. There was no significant difference in corticotroph responsiveness to secretagogues between cells harvested at gestation day 129 or gestation day 138. Simultaneous exposure to CRH and OT stimulated increases in ACTH output that were significantly greater than for OT or CRH alone. However, no similar synergistic interaction existed between OT and AVP. Cortisol attenuated OT-stimulated ACTH output. In conclusion, hypothalamic OT mRNA increases at term and OT can stimulate ACTH output from fetal corticotrophs. Together, these data indicate that OT may be involved in the regulation of ACTH secretion in fetal sheep in late gestation.

Modulation of adrenal cell functions by cadmium salts. 4. Ca(2+)-dependent sites affected by CdCl2 during basal and ACTH-stimulated steroid synthesis.

In previous studies, nonlethal CdCl2 concentrations apparently inhibited basal Y-1 mouse adrenal tumor cell endogenous mitochondrial cholesterol conversion to pregnenolone. In addition, CdCl2 inhibited all agents stimulating both plasma membrane-dependent cAMP synthesis and 20 alpha-hydroxy-4-pregnen-3-one (20DHP) secretion. Bypassing the plasma membrane using dibutyryl-cAMP (dbcAMP) stimulated cytoplasmic cholesterol metabolism and 20DHP secretion in the presence of CdCl2. Since CdCl2 competed at metabolic steps requiring Ca2+ in other tissues, experiments were designed to examine Cd2+ competition with Ca2+ during steroidogenesis. Sets of cells incubated with either medium or adrenocorticotropin (ACTH) with or without CdCl2 were also treated with 0, 1.0, 5.0 or 10.0 mmol/L CaCl2 in the presence or absence of EGTA, a relatively specific Ca2+, but not Cd2+, chelating agent. Another experimental cell set incubated with either medium or ACTH, with or without CdCl2, was treated with or without 1 mmol/L A23187, an ionophore specifically facilitating extracellular Ca2+ transfer across plasma membranes. Besides determining Ca2+ involvement in steroidogenesis using steroid secretion as an endpoint, we directly measured Ca2+ concentrations using intracellular fura-2 fluorescence. Following loading with 2 mumol/L fura-2, cells remained untreated or medium was infused with CdCl2, ACTH, ACTH/CdCl2 or ACTH followed after 50 s by CdCl2. Using Ca(2+)-supplemented media, we observed that Cd2+ inhibition of ACTH-stimulated 20DHP secretion was completely reversed. Standard Ca(2+)-containing medium supplemented with Ca2+ also enhanced maximally stimulated 20DHP secretion by ACTH. 20DHP secretion by ACTH-treated and ACTH/Cd(2+)-treated cells was only reduced by EGTA, when Ca2+ was not supplemented. The ionophore A23187 increased basal and ACTH-stimulated 20DHP secretion by Cd(2+)-treated cells, suggesting that extracellular Ca2+ resources may compete against Cd2+ effects on plasma membrane cAMP synthesis and on basal cholesterol metabolism by mitochondria. No time-dependent change in Ca2+ concentrations occurred within untreated cell suspensions. ACTH stimulation caused a 25 s burst in Ca2+ concentrations before returning to basal, steady-state levels. Cd2+ also stimulated intracellular fura-2 fluorescence. Untreated cell suspensions infused with Cd2+ exhibited a continuous rise in intracellular fluorescence. ACTH/CdCl2-treated cells exhibited a hyperbolic rise in intracellular fluorescence over the 300 s study period. Cells treated with Cd2+ 50 s after ACTH treatment initially exhibited the 25 s fluorescence burst followed by a Cd(2+)-induced hyperbolic rise in intracellular Cd2+. These fluorescence measurements suggested that cytoplasmic Ca2+ changes do not appear to be necessary for basal 20DHP synthesis and secretion; only a 25 s burst in intracellular Ca2+ is necessary to a slightly higher plateau level for stimulated 20DHP synthesis and secretion. Cd2+ freely enters the cell under basal conditions and Cd2+ entry is accelerated by ACTH stimulation. Data were consistent with Ca2+ being required for optimal stimulated steroid production and Cd2+ probably competing with Ca2+ during basal mitochondrial cholesterol metabolism and plasma membrane ACTH-stimulated cAMP generation.

Atrial natriuretic peptide and feeding activity patterns in rats.

This review presents historical data about atrial natriuretic peptide (ANP) from its discovery as an atrial natriuretic factor (ANF) to its role as an atrial natriuretic hormone (ANH). As a hormone, ANP can interact with the hypothalamic-pituitary-adrenal axis (HPA-A) and is related to feeding activity patterns in the rat. Food restriction proved to be an interesting model to investigate this relationship. The role of ANP must be understood within a context of peripheral and central interactions involving different peptides and pathways.

Atrial natriuretic peptide and feeding activity patterns in rats

This review presents historical data about atrial natriuretic peptide (ANP) from its discovery as an atrial natriuretic factor (ANF) to its role as an atrial natriuretic hormone (ANH). As a hormone, ANP can interact with the hypothalamic-pituitary-adrenal axis (HPA-A) and is related to feeding activity patterns in the rat. Food restriction proved to be an interesting model to investigate this relationship. The role of ANP must be understood within a context of peripheral and central interactions involving different peptides and pathways.

Daily cycle of fos expression within hypothalamic POMC neurons of the male rat.

The activity of hypothalamic pro-opiomelanocortin (POMC) neurons is known to display a circadian cycle. We hypothesized that the existence of a c-Fos responsive element (AP-1 site) within the POMC gene sequence might reflect the ability of POMC neurons to express c-fos proto-oncogene during circadian increase of their neuronal activity. To this aim, adult male rats previously kept under a controlled 12 h light/12 h dark schedule were sacrificed every 4 h throughout the 24 h cycle and their brains processed for Fos and/or POMC immunocytochemistry. Here we show that, specifically during the dark period of the cycle, the mediobasal hypothalamic area spontaneously exhibits a strong Fos immunoreactivity, whereas very low Fos labelling was detected during the light period. As postulated, the simultaneous visualisation of both Fos and POMC antigens allowed us to show that this nocturnal induction of Fos occurs almost exclusively at the nuclear level of POMC-producing neurons. These results not only highlight the mechanisms underlying the physiological functioning of the hypothalamic POMC system, but also demonstrate the feasibility of using c-fos expression as a useful tool to assess the pharmacological effect of drugs on the activity of POMC neurons as is the case for many other neuronal systems. Such drugs might be relevant in the treatment of psychosis since an alteration of POMC-related peptide transmission has been reported in the brains of both schizophrenic and depressive patients.

The tyrosine hydroxylase-human growth hormone (GH) transgenic mouse as a model of hypothalamic GH deficiency: growth retardation is the result of a selective reduction in somatotrope numbers despite normal somatotrope function.

Dwarf tyrosine hydroxylase-human GH (TH-hGH) transgenic mice carrying the hGH reporter gene targeted by the TH promoter express hGH in those regions of the hypothalamus responsible for regulation of pituitary GH secretion. Central expression of the hGH gene decreases GH-releasing hormone (GHRH) and increases somatostatin, which ultimately impacts on pituitary function by reducing the overall amount of GH produced. In the present study, we sought to determine if the reduction of pituitary GH in TH-hGH mice could be attributed to a decrease in somatotrope cell numbers and/or an impairment of somatotrope function. Pituitaries from TH-hGH or wild-type (WT) male and female mice were enzymatically dispersed, counted, and immunostained for GH, PRL, TSH, and ACTH. The total number of pituitary cells recovered from TH-hGH pituitaries was approximately one-half of that from WT controls. However, the proportion of cells that stained for GH and PRL were virtually identical (males, GH-TH-hGH, 58.1 +/- 1.0% [mean +/- SEM] vs. WT, 60.7 +/- 1.0%; PRL-TH-hGH, 43.4 +/- 2.2% vs. WT, 43.1 +/- 0.7%; females, GH-TH-hGH, 47.9 +/- 2.3% vs. WT, 41.5 +/- 3.5%; PRL-TH-hGH, 43.3 +/- 3.2% vs. WT, 47.1 +/- 3.3%). In contrast, percentages of both TSH- and ACTH-containing cells were increased in TH-hGH pituitaries relative to controls (males, TSH-TH-hGH, 15.1 +/- 2.3% vs. WT, 9.6 +/- 1.5%; ACTH-TH-hGH, 24.5 +/- 2.5% vs. WT, 10.9 +/- 0.9%; females: TSH-TH-hGH, 11.3 +/- 0.7% vs. WT, 7.5 +/- 0.6%; ACTH-TH-hGH, 19.8 +/- 1.6% vs. WT, 9.3 +/- 0.8%; P < 0.05). Calculation of the absolute number of each cell type per pituitary demonstrated TH-hGH mice to have about one-half the number of GH and PRL cells, whereas TSH and ACTH cell populations were comparable with that of their WT counterparts. Immunocytochemical localization of GH cells within pituitary sections from TH-hGH mice revealed that somatotropes were confined primarily to the lateral wings of the adenohypophysis, in contrast to the heterogeneous distribution of GH-immunostained cells in WT pituitaries. To assess the functional capacity of the somatotrope populations, pituitary cells from TH-hGH and WT mice were challenged with mouse GHRH (0.01-10 nM). The quantity of GH released (as assessed by both RIA and reverse hemolytic plaque assay) under basal and stimulated conditions did not differ among TH-hGH and WT pituitary cell cultures. Similarly, GHRH induced intracellular cAMP levels were comparable. These results indicate that proliferation of pituitary somatotropes and lactotropes is much more sensitive to changes in GHRH input than is the capability of developing regulated GH secretory function.

Modulation of immune responses by anabolic androgenic steroids.

Anabolic androgenic steroids (AS) have recently been placed on the Food and Drug Administration's (FDA's) list of controlled substances, because of the adverse effects seen in athletes taking accelerated dosages in attempts to enhance performance. Reported deleterious effects on abusers include sterility, gynecomastia in males, acne, balding, psychological changes, and increased risks of heart disease and liver neoplasia. Considering the roles of the immune and neuroendocrine systems and their interactions in many of these pathologies, it is important to determine the effects of these derivitized androgens on this connection. Little is known in this respect. We therefore determined the effects of anabolic steroids on certain immune responses and their effects on the extrapituitary production of corticotropin by lymphocytes. We present evidence that (1) both 17-beta and 17-alpha esterified AS, nandrolone decanoate and oxymethenelone, respectively, significantly inhibited production of antibody to sheep red blood cells in a murine abuse model; (2) the control androgens testosterone and dehydroepian-drosterone (DHEA) or sesame seed oil vehicle had no significant effects on antibody production; (3) nandrolone decanoate and oxymethenelone directly induced the production of the inflammatory cytokines IL-1 beta and TNF-alpha from human peripheral blood lymphocytes but had no effect on IL-2 or IL-10 production; (4) control androgens had no direct cytokine inducing effect; (5) nandrolone decanoate significantly inhibited IFN production in human WISH and murine L-929 cells; and (6) nandrolone decanoate significantly inhibited the production of corticotropin in human peripheral blood lymphocytes following viral infection. These data indicate that high doses of anabolic steroids can have significant effects on immune responses and extrapituitary production of corticotropin. Furthermore, the mouse model should provide an effective means by which to study other deleterious effects of anabolic steroid abuse in humans.