Corticotropin-releasing hormone microinfusion in the central amygdala diminishes a cardiac parasympathetic outflow under stress-free conditions

The central nucleus of the amygdala (CeA) is known to be involved in the regulation of autonomic, neuroendocrine and behavioural responses in stress situations. The CeA contains large numbers of corticotropin-releasing hormone (CRH) cell bodies. Neuroanatomical studies revealed that the majority of the CRH fibres from the CeA have direct connections with autonomic regulatory nuclei in the brainstem. In the present study, the effects of locally infused CRH (30 ng) into the CeA, in freely moving male Wistar rats under stress-free conditions, were examined. Heart rate, endocrine parameters and behavioural activity were repeatedly measured before, during and after local administration of CRH, pretreated with either artificial CSF or the CRH-receptor antagonist, α-helical CRH (α-CRH). CRH infusion alone caused a long-lasting increases in heart rate without affecting plasma adrenaline and noradrenaline as indicators of sympathetic activity. This CRH-induced tachycardia was effectively blocked by pretreatment with a high dose (1 μg) α-hCRH locally into the CeA, while the pretreatment with low dose (0.1 μg) of the α-hCRH caused a minor blockade of the CRH-induced tachycardia. The results suggest that CRH mechanisms in the CeA regulate the autonomic changes probably only by affecting parasympathetic but not sympathetic output systems. Because CRH is given at the level of the cell body of the CRH neurons in the CeA, we suggest that the reduction of the parasympathetic output may be explained as an autoreceptor-mediated inhibition of CRH neurons from the CeA with parasympathetic-regulating brainstem nuclei.     

Aminoglutethimide, a corticosteroid synthesis inhibitor, facilitates brain stimulation reward in food-restricted rats: an investigation of underlying mechanisms

 It was previously observed that the corticosteroid synthesis inhibitor, aminoglutethimide (AG), markedly facilitates lateral hypothalamic self-stimulation (LHSS) in food-restricted rats. This effect is not present 30 min after injection when plasma corticosterone levels are suppressed, but rather at 2 h when corticosterone has recovered from suppression. In experiment 1, it was confirmed that AG (50.0 mg/kg, SC) lowers the threshold for LHSS in food-restricted rats but not in control rats that have ad libitum access to food. This effect occurred independently of whether food restriction, by itself, lowered threshold. Experiment 2 examined whether the facilitation of LHSS coincides with biosynthetic rebound of corticosteroid precursors. While a pregnenolone surge was demonstrated by radioimmunoassay, dose-response testing with exogenous pregnenolone and progesterone (0.1, 1.0 and 10.0 mg/kg, SC) failed to confirm the prediction that one of these precursors facilitates reward. Therefore, a general test of the involvement of adrenocortical biosynthetic events was conducted in experiment 3 where rats were adrenalectomized (ADX) or sham-operated prior to food restriction. Surprisingly, ADX did not diminish the effect of AG. This finding raises the possibility of a CNS, rather than adrenal, site of action. AG is known to penetrate the blood-brain barrier and exert weak anticonvulsant effects. The facilitation of reward may result from central inhibitory effects of the drug and share a common basis with the enhanced reinforcing potency of other CNS depressants in food- restricted rats. Received: 25 March 1997 / Final version: 2 June 1997     

Role of δ-opioid receptor subtypes in anxiety-related behaviors in the elevated plus-maze in rats

Rationale Recent studies have shown that endogenous opioid systems are associated with the regulation of emotional responses. In particular, it has been reported that δ-opioid receptors act naturally to inhibit stress and anxiety. Objective The present study was designed to examine the possible involvement of opioid δ-receptor subtypes in the anxiety-related behavior in the elevated-plus-maze test. Methods Six-week-old male Lewis rats were used. The total numbers of visits to the closed and open arms and the cumulative time spent and visits in the open arms were determined. Plasma corticosterone levels were measured by enzyme immunoassay. Results Naltrindole (NTI), a δ-opioid receptor antagonist (3 mg/kg s.c.), induced a significant decrease in the percentages of time spent and visits in the open arms. Naltriben (NTB), a δ₂-opioid receptor antagonist (3 mg/kg s.c.), but not 7-benzylidenenaltrexone, a δ₁-opioid receptor antagonist, produced similar anxiety-related behaviors in the elevated plus-maze. These effects of NTI and NTB were antagonized by pretreatment with (+)-4-[(aR)-a-((2S,5R)-4-allyl-2,5-dimethyl-1piperazinyl)-3-methoxybenzyl]-N,N-diethylbenzamide (SNC80), a δ-opioid receptor agonist. Furthermore, after exposure to the elevated plus-maze, the maximal increase in the plasma corticosterone level in NTI-treated rats was clearly higher than that in vehicle-treated rats. However, when NTI and SNC80 were coadministered, higher levels of plasma corticosterone were not seen after exposure to the elevated plus-maze. Conclusion These results suggest that endogenous δ₂-opioid-receptor-mediated systems are involved in the regulation of anxiety-related behaviors and might play a physiologically important role in the regulation of adrenocortical activity.     

Plasma corticosterone in the rat in response to nicotine and saline injections in a context previously paired or unpaired with nicotine

Rationale Following repeated injections with nicotine paired with a distinctive environment, some studies have reported that the distinctive context becomes a conditioned stimulus (CS) capable of eliciting conditioned corticosterone (CORT) release. Conversely, other studies have found that exposure to the CS results in conditioned attenuation of nicotine-induced CORT release.Objective The present study was designed to examine whether these sets of separate findings could be replicated in animals exposed to the same experimental procedures within the same study.Methods CORT assessments were conducted in a distinctive context after independent groups of animals were injected with either saline or nicotine (1 mg/kg, salt) after five or ten nicotine injections either explicitly paired or unpaired with a distinctive context. The design also included groups of nicotine-naïve rats exposed to the experimental procedures and assessed for CORT levels following either nicotine or saline injections during their first, and after their fifth and tenth context exposures.Results CORT levels were higher after nicotine than after saline, and higher in the paired than in the unpaired condition. Exposure to the context without nicotine produced conditioned CORT release and exposure to the context did not attenuate nicotine-induced CORT release.Conclusions The results supported the notion that a CS associated with nicotine effects elicit a conditioned response (CR) in the form of CORT release. Future research will be needed to examine whether conditioned CORT release can explain the context-dependent attenuations of nicotine-induced CORT.     

Exposure to forced swim stress alters morphofunctional characteristics of the rat thymus

The aim of this study was to investigate whether chronic stress, induced by repeated daily swimming during 21 days, alters the morphofunctional parameters in the thymus of adult rats. Our results showed that chronic stress reduced thymus mass, total number of thymocytes, volume of the thymus compartments and numerical density of thymocytes within thymus inner cortex and medulla. However, the percentage of apoptotic cells and the level of corticosterone were significantly increased. The percentages of CD4-CD8-TCRβ^low/high and CD4-CD8+TCRβ^-thymocytes were significantly increased, while the percentage of the least mature CD4+CD8-SP TCRβ⁻ thymocytes was significantly decreased.

These results show that recurred swimming procedure induces thymus hypotrophy and elevated percentage of DN TCRβ⁺ cells.     

Glucocorticoid regulation of peptide genes in neuroendocrine CRH neurons: a complexity beyond negative feedback

This review will examine our current knowledge of a fundamental property of CRH neuroendocrine neurons: how the major endpoint of the HPA axis--adrenal glucocorticoids--interacts with the mechanisms controlling the expression of the genes that encode ACTH secretogogues. A great deal of work over the past 25 years has led to the notion that this question has an ostensibly simple answer: glucocorticoids inhibit peptide gene expression using "negative feedback" at the CRH neuron and elsewhere. However, closely examining how glucocorticoids act in different physiological circumstances reveals a much more complex set of answers, particularly if we consider how the processes that control peptide synthesis and release are coupled. Out of this examination emerges a more flexible and complex framework for examining the integrative mechanisms controlling the CRH neuron. Although we will mostly focus on the Crh gene, relevant aspects of the vasopressin (Avp) and pro-enkephalin (pEnk) gene regulatory mechanisms will also be discussed.     

Brain catalase activity inhibition as well as opioid receptor antagonism increases ethanol-induced HPA axis activation

BACKGROUND: Growing evidence indicates that brain catalase activity is involved in the psychopharmacological actions of ethanol. Recent data suggest that participation of this enzymatic system in some ethanol effects could be mediated by the endogenous opioid system. The present study assessed whether brain catalase has a role in ethanol-induced activation of the HPA axis, a neuroendocrine system modulated by the endogenous opioid neurotransmission. METHODS: Swiss male mice received an intraperitoneal injection of the catalase inhibitor 3-amino-1,2,4-triazole (AT; 0-1 g/kg), and 0 to 20 hr after this administration, animals received an ethanol (0-4 g/kg; intraperitoneally) challenge. Thirty, 60, or 120 min after ethanol administration, plasma corticosterone levels were determined immunoenzymatically. In addition, we tested the effects of 45 mg/kg of cyanamide (another catalase inhibitor) and 0 to 2 mg/kg of naltrexone (nonselective opioid receptor antagonist) on ethanol-induced enhancement in plasma corticosterone values. RESULTS: The present study revealed that AT boosts ethanol-induced increase in plasma corticosterone levels in a dose- and time-dependent manner. However, it did not affect corticosterone values when measured after administration of saline, cocaine (4 mg/kg, intraperitoneally), or morphine (30 mg/kg, intraperitoneally). The catalase inhibitor cyanamide (45 mg/kg, intraperitoneally) also increased ethanol-related plasma corticosterone levels. These effects of AT and cyanamide on ethanol-induced corticosterone values were observed under treatment conditions that decreased significantly brain catalase activity. Indeed, a significant correlation between effects of catalase manipulations on both variables was found. Finally, we found that the administration of naltrexone enhanced the levels of plasma corticosterone after the administration of saline or ethanol. CONCLUSIONS: This study shows that the inhibition of brain catalase increases ethanol-induced plasma corticosterone levels. Results are discussed together with previous findings suggesting a putative linkage between brain ethanol metabolism and the endogenous opioid system to explain some of the neuroendocrine effects of ethanol.     

Insulin resistance contributes to aberrant stress responses in the Tg2576 mouse model of Alzheimer's disease

We previously reported aberrant stress responses and impaired glucose tolerance in transgenic Tg2576 mice, a model of Alzheimer's disease (AD). Here we report that by 8 months of age, Tg2576 mice had lower basal serum insulin concentrations and exhibited a delayed insulin-induced reduction in blood glucose levels relative to wild-type mice. However, the basal levels of blood glucose and percent glycosylated hemoglobin (%HbA1c) were similar between the two groups of mice. While the basal levels of serum corticosterone were similar between Tg2576 and wild-type mice, an overnight fasting caused a greater rise in serum corticosterone levels and an excessive reduction in serum insulin concentrations in the transgenics. At 9 months of age, we began administering Tg2576 mice rosiglitazone, an agonist of peroxisome proliferator-activated receptor-gamma that increases peripheral insulin sensitivity, and after 6 weeks of administration the Tg2576 mice had the same response to insulin and increase in serum corticosterone levels after an overnight fast as did wild-type mice. By 13 months of age, untreated Tg2576 mice had become hyperinsulinemic, in contrast to Tg2576 mice administered rosiglitazone for 4 months where the serum insulin concentrations were maintained at levels observed in wild-type mice. These results provide evidence for a relationship between insulin resistance, impaired regulation of insulin and glucose levels, and aberrant stress responses in Tg2576 mice.     

Stress-induced REM sleep increase is antagonized by naltrexone in rats

Rationale The expression of sleep is influenced by situations that take place during the preceding waking period, giving rise to different patterns of sleep architecture. Immobilization stress (IMB) induces an increase of both rapid eye movement (REM) and slow wave sleep (SWS). It has been suggested that these changes are mediated in part by noradrenaline and by the corticotrophin releasing factor.Objective To determine the participation of mu receptors in the stress-induced increase of REM sleep using naltrexone (NTX).Methods Twelve adult male rats were implanted for sleep recordings. Subjects were recorded under control conditions as well as after: a) IMB stress (1 h); b) injection of NTX (1.5 mg/kg); c) NTX plus IMB. To assess corticosterone levels, additional groups (n=5) were decapitated at 0, 1, 3 and 6 h after vehicle injection and after immobilization. Four groups were decapitated at 0, 1, 3, and 6 h after NTX plus IMB. Corticosterone plasma levels were determined by HPLC.Results IMB induces an increase in REM and SWS, and a decrease in wakefulness. Administration of NTX before IMB suppresses the effects of stress on sleep. NTX administration is innocuous in non-stressed animals. However, NTX administration does not prevent the rise of corticosterone normally observed after IMB stress.Conclusion These data suggest that NTX prevents the effects of IMB stress on sleep by acting outside of the hypothalamic-pituitary-adrenal axis that partially mediates the stress response.     

Effects of simulated environmental conditions on glucocorticoid metabolite measurements in white-tailed deer feces

Environmental conditions may influence fecal glucocorticoid metabolite measurements if feces cannot be collected immediately after deposition. To evaluate the influence of environmental conditions on fecal glucocorticoid metabolite concentrations, we exposed fresh fecal samples to 1 of 5 simulated conditions: (1) room temperature (22 degrees C), (2) high heat (38 degrees C), (3) alternating high heat and room temperature cycle, (4) alternating freezing (-20 degrees C) and room temperature cycle, and (5) simulated rainfall (0.85 cm every other day at 22 degrees C) for 7 days. We collected fresh white-tailed deer (Odocoileus virginianus) feces at various times pre- and post-adrenocorticotropin injection to provide samples with initially low (n=5), medium (n=5), and high (n=5) glucocorticoid concentrations. Feces were mixed thoroughly and then allocated into five 10-g samples. Also, a 5-g sub-sample was taken from each fecal mass prior to treatment and stored at -20 degrees C until assayed. We subsampled from all treatments once every 24-h for 7 days. Fecal samples were assayed using [125I]corticosterone radioimmunoassay kits. Fecal glucocorticoid metabolites in all three groups in the simulated rainfall treatment and the low group in the alternating freezing and room temperature treatment increased significantly over the 7-day period. We believe increased microbial metabolism of fecal glucocorticoids may partly explain these results. Other biochemical processes (e.g., cleavage of conjugate side groups from hormone metabolites by non-microbial action or release of glucocorticoids from lipid micelles) may also have increased fecal glucocorticoid measurements. Our findings suggest that fecal samples exposed to rainfall for one week may artificially inflate fecal glucocorticoid measurements. Thus, researchers should recognize the potential bias when collecting fecal samples exposed to rainfall. Non-fresh samples may prove useful when care is taken to address the elevation in immunoreactive glucocorticoid concentrations.