Corticotropin-releasing Hormone mRNA Expression Research Articles

Rapid acclimation of the cortisol stress response in adult turquoise killifish Nothobranchius furzeri.

The turquoise killifish Nothobranchius furzeri is an increasingly popular model species for comparative vertebrate research, and the basic physiology including responses to stressful stimuli are of primary interest. We exposed adult killifish to a single or repeated periods of acute confinement followed by analysis of tissue cortisol and plasma cortisol concentrations. Individuals were also sampled for messenger RNA (mRNA) expression of corticotropin-releasing hormone (CRH), mineralocorticoid receptor (MR), and glucocorticoid receptor (GR) in the brain to examine the effects of repeated stress events on constitutive expression of these important stress axis components. Following a single 30-minute confinement stress, male plasma cortisol significantly differed from baseline (p = 0.04). Both male and female whole-body cortisol were significantly increased (p = 0.004 and p = 0.04, respectively) at 15 and 30 minutes poststress. Despite obvious dimorphic behavior and morphology, cortisol concentrations did not differ between the sexes. Exposure to daily repeated confinement for one week altered the cortisol response in both sexes. Time 0, 15, and 60 minutes poststress cortisol concentrations were depressed in repeatedly stressed males (p ≤ 0.05), and times 0, 30 and 120 minutes poststress cortisol concentrations were depressed in repeatedly stressed females (p ≤ 0.05). Constitutive expression of CRH, MR, and GR mRNA in the brain following one week of repeated stress events did not differ among treatments or sexes. This study introduces the first description of hypothalamic-pituitary-interrenal axis activity in this important model species. Reduced cortisol production in repeatedly stressed adult killifish suggests acclimation to repeated stressors. Furthermore, acclimation was rapid, and plasma cortisol concentrations altered significantly in as little as one week.

Sex differences in hippocampal mineralocorticoid and glucocorticoid receptor mRNA expression in response to acute mate pair separation in zebra finches (Taeniopygia guttata).

Mate separation has been shown to mediate changes in physiological and behavioral processes via activation of the hypothalamo-pituitary-adrenal (HPA) axis in both mammalian and avian species. To elucidate the neural mechanisms associated with changes in the HPA axis in response to social stress, we investigated the effects of mate pair separation on circulating corticosterone concentrations as well as gene expression levels of mineralocorticoid receptor (MR), glucocorticoid receptor (GR), and corticotropin releasing hormone (CRH) in the hypothalamus and hippocampus of both male and female zebra finches, a species that forms strong pair bonds. Zebra finches (Taeniopygia guttata) were housed three to a cage (a mated pair plus a stimulus female), and were assigned to one of three new housing treatment groups: (1) male or female removed from their respective mate and placed in a cage with a new opposite sex conspecific and stimulus female (2) male or female that remained with their mate, but a new stimulus female was introduced, or (3) the subjects were handled but not separated from their mate or the stimulus female. After 48 hr in the new housing condition, we observed significant increases in plasma corticosterone concentrations in response to both mate pair and stimulus female separation. No significant differences in MR, GR, or CRH mRNA expression in the hypothalamus were observed in response to any treatment for both males and females. Females exhibited a significant up regulation in hippocampal MR, but not GR mRNA, whereas males exhibited a significant down regulation of both hippocampal MR and GR mRNA in response to mate pair separation. Thus, the hippocampus appears to play a key role in regulating sex specific responses to social stressors.

Action of neurotensin, corticotropin-releasing hormone, and RFamide-related peptide-3 in E2-induced negative feedback control: studies using a mouse arcuate nucleus hypothalamic cell model.

The recently established immortalized hypothalamic cell model mHypoA-55 possesses characteristics similar to those of Kiss-1 neurons in the arcuate nucleus (ARC) region of the hypothalamus. Here, we show that Kiss-1 gene expression in these cells was downregulated by 17β-estradiol (E2) under certain conditions. Both neurotensin (NT) and corticotropin-releasing hormone (CRH) were expressed in these cells and upregulated by E2. Stimulation of mHypoA-55 cells with NT and CRH significantly decreased Kiss-1 mRNA expression. A mammalian gonadotropin-inhibitory hormone homolog, RFamide-related peptide-3 (RFRP-3), was also found to be expressed in mHypoA-55 cells, and RFRP-3 expression in these cells was increased by exogenous melatonin stimulation. E2 stimulation also upregulated RFRP-3 expression in these cells. Stimulation of mHypoA-55 cells with RFRP-3 significantly increased the expression of NT and CRH. Furthermore, melatonin stimulation resulted in the increase of both NT and CRH mRNA expression in mHypoA-55 cells. On the other hand, in experiments using mHypoA-50 cells, which were originally derived from hypothalamic neurons in the anteroventral periventricular nucleus, Kiss-1 gene expression was upregulated by both NT and CRH, although E2 increased both NT and CRH expression, similarly to the mHypoA-55 cells. Our observations using the hypothalamic ARC cell model mHypoA-55 suggest that NT and CRH have inhibitory effects on Kiss-1 gene expression under the influence of E2 in association with RFRP-3 expression. Thus, these neuropeptides might be involved in E2-induced negative feedback mechanisms.

Dominance relationships in Syrian hamsters modulate neuroendocrine and behavioral responses to social stress

Stress is a well-known risk factor for psychopathology and rodent models of social defeat have strong face, etiological, construct and predictive validity for these conditions. Syrian hamsters are highly aggressive and territorial, but after an acute social defeat experience they become submissive and no longer defend their home territory, even from a smaller, non-aggressive intruder. This defeat-induced change in social behavior is called conditioned defeat (CD). We have shown that dominant hamsters show increased neural activity in the ventromedial prefrontal cortex (vmPFC) following social defeat stress and exhibit a reduced CD response at social interaction testing compared to subordinates. Although the vmPFC can inhibit the neuroendocrine stress response, it is unknown whether dominants and subordinates differ in stress-induced activity of the extended hypothalamic-pituitary-adrenal (HPA) axis. Here, we show that, following acute social defeat, dominants exhibit decreased submissive and defensive behavior compared to subordinates but do not differ from subordinates or social status controls (SSCs) in defeat-induced cortisol concentrations. Furthermore, both dominants and SSCs show greater corticotropin-releasing hormone (CRH) mRNA expression in the basolateral/central amygdala compared to subordinates, while there was no effect of social status on CRH mRNA expression in the paraventricular nucleus of the hypothalamus or bed nucleus of the stria terminalis. Overall, status-dependent differences in the CD response do not appear linked to changes in stress-induced cortisol concentrations or CRH gene expression, which is consistent with the view that stress resilience is not a lack of a physiological stress response but the addition of stress coping mechanisms.Lay summaryDominant hamsters show resistance to the behavioral effects of acute social defeat compared to subordinates, but it is unclear whether social status modulates the neuroendocrine stress response in Syrian hamsters. This study indicates that dominant social status does not alter stress-induced activity of the extended hypothalamic-pituitary-adrenal (HPA) axis, which suggests that the ability of dominants to cope with social defeat stress is not associated with changes in their neuroendocrine stress response.

Single oral administration of flavan 3-ols induces stress responses monitored with stress hormone elevations in the plasma and paraventricular nucleus

We previously confirmed that postprandial alterations in the circulation and metabolism after a single oral dose of flavan 3-ols (mixture of catechin and catechin oligomers) were involved in an increase in sympathetic nervous activity. However, it is well known that, in response to various stresses, activation of the hypothalamic-pituitary-adrenal (HPA) axis occurs together with sympathetic nerve activity, which is associated with activation of the sympathetic-adrenal-medullary (SAM) axis. In this study, we examined whether the HPA axis was activated after a single dose of flavan 3-ols. We administered an oral dose of 10 or 50 mg/kg flavan 3-ols to male ICR mice, removed the brains, and fixed them in paraformaldehyde-phosphate buffer. Other animals that were treated similarly were decapitated, and blood was collected. In the paraventricular nucleus (PVN), c-fos mRNA expression increased significantly at 15 min after administration of either 10 or 50 mg/kg flavan 3-ols. Corticotropin-releasing hormone (CRH) mRNA expression levels significantly increased at 240 min after administration of 10 mg/kg flavan 3-ols, and at 60 min after administration of 50 mg/kg flavan 3-ols. Plasma corticosterone levels were also significantly increased at 240 min after ingestion of 50 mg/kg flavan 3-ols. In this experiment, we confirmed that the ingestion of flavan 3-ols acted as a stressor in mammals with activation both the SAM and HPA axes.

The Protein Status of Rats Affects the Rewarding Value of Meals Due to their Protein Content

Protein status is controlled by the brain, which modulates feeding behavior to prevent protein deficiency. This study tested in rats whether protein status modulates feeding behavior through brain reward pathways. Experiments were conducted in male Wistar rats (mean ± SD weight; 230±16 g). In experiment 1, rats adapted for 2 wk to a low-protein (LP; 6% of energy) or a normal-protein (NP; 14% of energy) diet were offered a choice between 3 cups containing high-protein (HP; 50% of energy), NP, or LP feed; their intake was measured for 24 h. In 2 other experiments, the rats were adapted for 2 wk to NP and either HP or LP diets and received, after overnight feed deprivation, a calibrated HP, NP, or LP meal daily. After the meal, on the last day, rats were killed and body composition and blood protein, triglycerides, gut neuropeptides, and hormones were determined. In the brain, neuropeptide mRNAs in the hypothalamus and c-Fos protein and opioid and dopaminergic receptor mRNAs in the nucleus accumbens (NAcc) were measured. Rats fed an LP compared with an NP diet had 7% lower body weight, significantly higher protein intake in a choice experiment (mean ± SD: 30.5%±0.05% compared with 20.5%±0.05% of energy), higher feed-deprived blood ghrelin, lower postmeal blood leptin, and higher neuropeptide Y (Npy) and corticotropin-releasing hormone (Crh) mRNA expression in the hypothalamus. In contrast to NP, rats fed an LP diet showed postmeal c-Fos protein expression in the NAcc, which was significantly different between meals, with LP < NP < HP. In contrast, in rats adapted to an HP diet compared with an NP diet, energy intake was lower; and in the NAcc, meal-induced c-Fos protein expression was 20% lower, and mRNA expression was 17% higher for dopamine receptor 2 (Drd2) receptors and 38% lower for κ opioid receptor (Oprk1) receptors. A protein-restricted diet induced a reward system-driven appetite for protein, whereas a protein-rich diet reduced the meal-induced activation of reward pathways and lowered energy intake in male rats.

Age Dependent Hypothalamic and Pituitary Responses to Novel Environment Stress or Lipopolysaccharide in Rats.

Previously, we have shown that the transcription factor nuclear factor interleukin (NF-IL)6 can be used as an activation marker for inflammatory lipopolysaccharide (LPS)-induced and psychological novel environment stress (NES) in the rat brain. Here, we aimed to investigate age dependent changes of hypothalamic and pituitary responses to NES (cage switch) or LPS (100 μg/kg) in 2 and 24 months old rats. Animals were sacrificed at specific time points, blood and brains withdrawn and analyzed using immunohistochemistry, RT-PCR and bioassays. In the old rats, telemetric recording revealed that NES-induced hyperthermia was enhanced and prolonged compared to the young group. Plasma IL-6 levels remained unchanged and hypothalamic IL-6 mRNA expression was increased in the old rats. Interestingly, this response was accompanied by a significant upregulation of corticotropin-releasing hormone mRNA expression only in young rats after NES and overall higher plasma corticosterone levels in all aged animals. Immunohistochemical analysis revealed a significant upregulation of NF-IL6-positive cells in the pituitary after NES or LPS-injection. In another important brain structure implicated in immune-to-brain communication, namely, in the median eminence (ME), NF-IL6-immunoreactivity was increased in aged animals, while the young group showed just minor activation after LPS-stimulation. Interestingly, we found a higher amount of NF-IL6-CD68-positive cells in the posterior pituitary of old rats compared to the young counterparts. Moreover, aging affected the regulation of cytokine interaction in the anterior pituitary lobe. LPS-treatment significantly enhanced the secretion of the cytokines IL-6 and TNFα into supernatants of primary cell cultures of the anterior pituitary. Furthermore, in the young rats, incubation with IL-6 and IL-10 antibodies before LPS-stimulation led to a robust decrease of IL-6 production and an increase of TNFα production by the pituitary cells. In the old rats, this specific cytokine interaction could not be detected. Overall, the present results revealed strong differences in the activation patterns and pathways between old and young rats after both stressors. The prolonged hyperthermic and inflammatory response seen in aged animals seems to be linked to dysregulated pituitary cytokine interactions and brain cell activation (NF-IL6) in the hypothalamus-pituitary-adrenal axis.

Sea Buckthorn Pomace Supplementation in the Diet of Growing Pigs—Effects on Fatty Acid Metabolism, HPA Activity and Immune Status

There is evidence that sea buckthorn, as a source of n-3 polyunsaturated fatty acids (n-3 PUFA), possesses health-enhancing properties and may modulate neuroendocrine and immune functions. In the present study, we investigated the effect of sea buckthorn pomace (SBP) supplementation in the diet of growing German Landrace pigs on fatty acids in the blood and hypothalamus, peripheral immune parameters and mRNA expression of corticotropin-releasing hormone (CRH), mineralocorticoid receptor (MR) and glucocorticoid receptor (GR) in the hypothalamus and spleen. Pigs were fed diets supplemented with 12% of dried SBP or 0% SBP (control group) over an intervention period of eight weeks. The fatty acid profiles in blood plasma were significantly affected by SBP supplementation only for C18:2n-6 and n-6/n-3 PUFA ratio compared with the control group. SBP supplementation did not significantly affect the fatty acid concentrations in the hypothalamus. Furthermore, there were no significant differences in mRNA expression of CRH, MR and GR in the hypothalamus or of GR mRNA expression in the spleen. Concerning the immune status, the plasma IgG levels tended to be higher in SBP pigs, whereas the leukocyte distribution, mitogen-stimulated lymphocyte proliferation, and serum IgM levels remained unchanged. In conclusion, the SBP supplementation of the diet only caused moderate effects on fatty acid metabolism, but no significant effects on hypothalamic–pituitary–adrenal (HPA) activity and immunity in growing pigs. It seems that a beneficial effect of dietary n-3 PUFA on health and welfare is more likely to be expected during stressful situations.

Resveratrol ameliorates chronic unpredictable mild stress-induced depression-like behavior: involvement of the HPA axis, inflammatory markers, BDNF, and Wnt/β-catenin pathway in rats.

Classic antidepressant drugs are modestly effective across the population and most are associated with intolerable side effects. Recently, numerous lines of evidence suggest that resveratrol (RES), a natural polyphenol, possesses beneficial therapeutic activity for depression. The aim of the present study was to explore whether RES exhibits an antidepressant-like effect in a depression model and to explore the possible mechanism. A depression model was established via chronic unpredictable mild stress (CUMS), after which the model rats in the RES and fluoxetine groups received a daily injection of RES or fluoxetine, respectively. The sucrose preference test, open field test, and forced swimming test were used to explore the antidepressant-like effects of RES. The activity of the hypothalamic–pituitary–adrenal (HPA) axis was evaluated by detecting the plasma corticosterone concentration and hypothalamic mRNA expression of corticotrophin-releasing hormone. The plasma interleukin-6 (IL-6), C-reactive protein (CRP), and tumor necrosis factor-α (TNF-α) concentrations were measured by enzyme-linked immunosorbent assay. Hippocampal protein expression of brain-derived neurotrophic factor (BDNF) and the Wnt/β-catenin pathway were analyzed by western blot. The results showed that RES relieved depression-like behavior of CUMS rats, as indicated by the increased sucrose preference and the decreased immobile time. Rats that received RES treatment exhibited reduced plasma corticosterone levels and corticotrophin-releasing hormone mRNA expression in the hypothalamus, suggesting that the hyperactivity of the HPA axis in CUMS rats was reversed by RES. Moreover, after RES treatment, the rats exhibited increased plasma IL-6, CRP, and TNF-α concentrations. Furthermore, RES treatment upregulated the hippocampal protein levels of BDNF and the relative ratio of p-β-catenin/β-catenin while downregulating the relative ratio of p-GSK-3β/GSK-3β. Our findings suggest that RES improved depressive behavior in CUMS rats by downregulating HPA axis hyperactivity, increasing BDNF expression and plasma IL-6, CRP, and TNF-α concentrations, and regulating the hippocampal Wnt/β-catenin pathway.

Comparison of efficacy of adrenocorticotrophic hormone and methylprednisolone on rat models of infantile spasm

Objective To compare the efficacy of adrenocorticotrophic hormone(ACTH) and methylprednisolone on the rat models of infantile spasms(IS). Methods The SD rats on postnatal 10 day (P10) were divided into blank group (n=18), control group (n=18) and model group (n=110) according to the random number table method.The rats of model group were prepared by adopting prenatal stress exposure and N- methyl -D aspartate (NMDA) injection.In the model group, after inducing epileptic seizures, the rats were divided into different groups (18 rats in each group) according to the random number table method as following: model group Ⅰ (subcutaneous injection of ACTH, 50 IU/kg, at P10: 1400, 2100; P11, P12: 700, 1400, 2100; P13: 700), model group Ⅱ(subcutaneous injection of 9 g/L saline), model group Ⅲ (intraperitoneal injection of methylprednisolone, 60 mg/kg, at P11, P12, P13: 900, once per day), model group Ⅳ (intraperitoneal injection of 9 g/L saline) and model group Ⅴ (positive control group, with no drug or saline injection). Three days later, epilepsy was induced again, and the rats of model group were intraperitoneally injected with NMDA (12 mg/kg) at P13(1000). The rats of control group were injected intraperitoneally with same volume of 9 g/L saline, but the rats of blank group were not treated.Behaviors of rats with epilepsy seizures were observed and epilepsy scores were given.The expression of corticotropin-releasing hormone (CRH) in the hypothalamus of each group was detected by using immunohistochemistry and fluorescence quantitative polymerase chain reaction.The learning and memorizing capacity of the rats were measured by Y-maze experiment. Results There was no death in the model group after the onset of seizure.In the model group Ⅰ, 13 cases were attacked(72.22%), and 14 cases were attacked in the model group Ⅲ(78.78%). The level of attack was decreased.The buckling state was not observed in model group and Ⅲ, but the latency period of epilepsy was prolonged and the epilepsy scores were significantly decreased.There were no significant differences of onset latency [(2 369.38±628.70) s vs.(1 922.93±462.36) s] and epilepsy score[(2.15±1.14) scores vs.(2.07±0.83) scores] between the 2 groups (all P>0.005). The rats of model group Ⅱ, Ⅳ, Ⅴ were all attacked completely and presented buckling state.There was no onset or death in blank group and control group.The number of CRH positive cells and CRH mRNA relative expression of each model group were higher than those in the blank group and control group.The number of CRH positive cells and CRH mRNA expression of model group Ⅰ and Ⅲ were lower than those in model group Ⅱ, Ⅳ and Ⅴ, and the differences were significant (all P 0.002 4). No significant difference was found between blank group and control group, or among model group Ⅱ, Ⅳ and Ⅴ (all P>0.002 4). There were no significant differences in the learning capacity among all groups (F=2.196, P>0.002 4). The correct response rate after 24 hours of the model group was lower than the blank group and control group, and ACTH and methylprednisolone pretreatment did not influence the memorizing capacity (P>0.002 4). Conclusion The effect of pretreatment of ACTH is similar to that of methylprednisolone in the rat model of IS. Key words: Infantile spasm; Adrenocorticotrophic hormone; Methylprednisolone; Rats model; Corticotropin-releasing hormone

Chronic salt-loading reduces basal excitatory input to CRH neurons in the paraventricular nucleus and accelerates recovery from restraint stress in male mice

Neurons synthesizing corticotrophin-releasing hormone (CRH) in the paraventricular nucleus of the hypothalamus (PVN) are activated during acute stress and act via the hypothalamic-pituitary-adrenal (HPA) axis to increase systemic levels of corticosterone (CORT). Recent data indicates that CRH neurons in the PVN are inhibited by acute salt-loading, and that this inhibition blunts the response to restraint stress as measured by increases in plasma CORT. The current study evaluates the effects of chronic rather than acute salt-loading on stress-induced activation of the HPA axis. Relative to euhydrated controls, chronic salt-loading over a 5-day period elevated plasma sodium and fluid intake without eliciting hypovolemia or substantial alterations in food intake or body weight. Chronic salt-loading also decreased expression of CRH mRNA in the anterior but not posterior portion of the PVN. Similarly, whole cell patch clamp recordings revealed that salt-loading effectively decreases spontaneous excitatory input to CRH neurons in the PVN without altering spontaneous inhibitory input. Generally consistent with these observations, chronic salt attenuated HPA axis activation as indicated by a significant reduction of plasma CORT during recovery from restraint stress.

Altered functionality of the corticotrophin-releasing hormone receptor-2 in the hypothalamic paraventricular nucleus of hyperphagic maternally separated rats

Early-life stress induces endocrine and metabolic alterations that increase food intake and overweight in adulthood. The stress response activates the corticotropin-releasing hormone (CRH) and urocortins' (Ucns) system in the hypothalamic paraventricular nucleus (PVN). These peptides induce anorexic effects through CRH-R2 receptor activation; however, chronic stressed animals develop hyperphagia despite of high PVN CRH expression.We analyzed this paradoxical behavior in adult rats subjected to maternal separation (MS) for 180min/daily during post-natal days 2–14, evaluating their body weight gain, food intake, serum corticosterone and vasopressin concentrations, PVN mRNA expression of CRH-R1, CRH-R2, CRH, Ucn2, Ucn3, vasopressin and CRH-R2 protein levels.MS adults increased their feeding, weight gain as well as circulating corticosterone and vasopressin levels, evincing chronic hyperactivity of the stress system. MS induced higher PVN CRH, Ucn2 and CRH-R2 mRNA expression and protein levels of CRH-R2 showed a tendency to decrease in the cellular membrane fraction. An intra-PVN injection of the CRH-R2 antagonist antisauvagine-30 in control adults increased receptor's mRNA expression, mimicking the observed PVN receptor's up-regulation of early-life MS adults.An injection of Ucn-2 directly into the PVN reduced food intake and increased PVN pCREB/CREB ratio in control animals; in contrast, Ucn-2 was unable to reduce food intake and enhance phosphorylated-CREB levels in PVN of MS rats.In conclusion, the chronic hyperactivity of the stress axis and PVN CRH-R2 resistance to Ucn2 effects, supported impaired receptor functionality in MS animals, probably due to its chronic stimulation by CRH or Ucn2, induced by early-life stress.

The flame retardant 2,2′,4,4′-Tetrabromodiphenyl ether enhances the expression of corticotropin-releasing hormone in the placental cell model JEG-3

Polybrominated diphenyl ethers (PBDEs) are chemicals used as flame retardants in household products. After disposing of these items, PBDEs leach from the products by surface water. BDE-47 is a PBDE congener commonly isolated from contaminated food and is the most studied isomer. The placenta is the major source of hormones during pregnancy, and an elevated level of corticotrophin-releasing hormone (CRH) is associated with premature delivery. In the present study, we examined changes in the placental CRH expression under BDE-47 exposure in the JEG-3 cell model system. These placental cells are derived from human choriocarcinoma. Our result showed that this pollutant induced the CRH mRNA expression at 0.5 nM or above in the cells. A similar trend was observed when CRH peptide was determined by Western analysis in the cell lysates. As previous studies have shown the importance of signal transduction pathways in the gene regulation, the status of some protein kinases in the present study was investigated. The phosphorylated PKCα, JNK, and P38 were increased by the toxicant treatment, and administering the specific inhibitors could counteract the induced CRH expression. It appeared that the signaling transduction pathway of PKC was a significant contributor in the transcriptional regulation. Further study by using Electrophoretic Mobility Shift Assay suggested that AP-2 was the ultimate DNA-binding element for the initiation of gene transcription. Because an untimely increased CRH may compromise fetal development and induce preterm birth, the present study suggested that endocrine changes in pregnancy should be taken into consideration in the next assessment.

Functional evidence that the nucleus of the hippocampal commissure shows an earlier activation from a stressor than the paraventricular nucleus: Implication of an additional structural component of the avian hypothalamo-pituitary-adrenal axis

Despite extensive data addressing the regulation of the hypothalamo-pituitary-adrenal (HPA) axis in vertebrates, the neuroendocrine regulation of stress in birds remains incomplete. The paraventricular nucleus (PVN) contains the key neuropeptides, corticotropin releasing hormone (CRH) and arginine vasotocin (AVT), containing neurons. However, another population of CRH neurons was recently identified in a septal nucleus called the nucleus of the hippocampal commissure (NHpC). Therefore, the current study investigated changes in gene expression of CRH and AVT in the PVN and CRH in the NHpC, as well as changes in plasma corticosterone concentrations following a stressor, food deprivation. In the NHpC, a rapid increase in CRH mRNA levels was observed as early as 2h, while relative CRH mRNA expression in the PVN increased thereafter from 4 to 12h of food deprivation. On the other hand, relative mRNA levels of AVT in the PVN were not observed until 8h and significantly increased at 12 and 24h following food deprivation. Furthermore, at the level of the anterior pituitary, relative expression of proopiomelanocortin transcripts followed gene expression patterns of CRH and AVT in the brain. In the absence of food, the pattern of plasma CORT showed an initial rise at 2h and a fourfold increase was measured at 4h that was sustained through 24h. Taken together, results from this study suggest that (1) CRH neurons in the NHpC appear to be the first responsive neurons to stress stimuli compared to those in the PVN, (2) CRH is predominantly functional in the early phase of stress while AVT is involved in the later phase of the stress period and (3) in birds, CRH neurons in the NHpC appear to be part of the classical HPA axis.

Assessing placental corticotrophin-releasing hormone disruption by hexestrol in a cell model.

Studies have shown that corticotrophin-releasing hormone (CRH) and relaxin are associated with early delivery. Our lab previously has shown the mycotoxin zeranol increases placental CRH expression. The mycotoxin is used in the farming industry to promote cattle growth, and some synthetic hormones are also used for the same purposes. In order to complete the picture of these growth promoting agents, we attempted to examine the synthetic hormones on the placental gene expression in the current study. Among the tested compounds, hexestrol induced the CRH mRNA and protein expression at 100nM in JEG-3 cells. As signal transduction pathways have been described in the transcriptional control previously, the activations of several protein kinases were determined. P38, PKCβ and JNK were activated upon hexestrol treatment. Since the P38-inhibitor SB203580 prevented hexestrol from inducing CRH in a subsequent experiment, P38 was likely involved in the transcriptional regulation. Electrophoretic mobility shift assay revealed an increase in the CRE binding activity in CRH promoter. This study showed that hexestrol exposure might be a concern for pregnant women.

Postnatal light alters hypothalamic-pituitary-adrenal axis function and induces a depressive-like phenotype in adult mice.

The postnatal light environment that a mouse experiences during the critical first three postnatal weeks has long-term effects on both its circadian rhythm output and clock gene expression. Furthermore, data from our lab suggest that postnatal light may also impact the hypothalamic-pituitary-adrenal (HPA) axis, which is a key regulator of stress. To test the effect of postnatal light exposure on adult stress responses and circadian rhythmicity, we raised mice under either 24-h light-dark cycles (LD), constant light (LL) or constant dark (DD) during the first three postnatal weeks. After weaning we then exposed all animals to LD cycles (basal conditions), followed by LL (stressed conditions) environments. We examined brain neuropeptide and glucocorticoid receptor (GR) expression, plasma corticosterone concentration rhythm and body temperature rhythm, together with depression- and anxiety-related behaviour. Results showed that LL- and DD-raised mice exhibited decreased GR expression in the hippocampus, increased plasma corticosterone concentration at the onset of the dark phase and a depressive phenotype when exposed to LD cycles later in life. Furthermore, LL-raised mice showed increased corticotrophin-releasing hormone mRNA expression in the paraventricular nucleus of the hypothalamus. When exposed to LL as adults, LL-raised mice showed a significant circadian rhythm of plasma corticosterone concentration, together with a shorter period and stronger circadian rhythm of body temperature compared to DD-raised mice. Taken together, these data suggest that altered postnatal light environments have long-term effects on the HPA axis and the circadian system, which can lead to altered stress responses and a depressive phenotype in adulthood.

Resveratrol Ameliorates the Anxiety- and Depression-Like Behavior of Subclinical Hypothyroidism Rat: Possible Involvement of the HPT Axis, HPA Axis, and Wnt/β-Catenin Pathway.

Metabolic disease subclinical hypothyroidism (SCH) is closely associated with depression-like behavior both in human and animal studies, and our previous studies have identified the antidepressant effect of resveratrol (RES) in stressed rat model. The aim of this study was to investigate whether RES would manifest an antidepressant effect in SCH rat model and explore the possible mechanism. A SCH rat model was induced by hemi-thyroid electrocauterization, after which the model rats in the RES and LT4 groups received a daily intragastric injection of RES at the dose of 15 mg/kg or LT4 at the dose of 60 μg/kg for 16 days. The rats’ plasma concentrations of thyroid hormones were measured. Behavioral performance and hypothalamic–pituitary–adrenal (HPA) activity were evaluated. The protein expression levels of the Wnt/β-catenin in the hippocampus were detected by western blot. The results showed that RES treatment downregulated the elevated plasma thyroid-stimulating hormone concentration and the hypothalamic mRNA expression of thyrotropin-releasing hormone in the SCH rats. RES-treated rats showed increased rearing frequency and distance in the open-field test, increased sucrose preference in the sucrose preference test, and decreased immobility in the forced swimming test compared with SCH rats. The ratio of the adrenal gland weight to body weight, the plasma corticosterone levels, and the hypothalamic corticotrophin-releasing hormone mRNA expression were reduced in the RES-treated rats. Moreover, RES treatment upregulated the relative ratio of phosphorylated-GSK3β (p-GSK3β)/GSK3β and protein levels of p-GSK3β, cyclin D1, and c-myc, while downregulating the relative ratio of phosphorylated-β-catenin (p-β-catenin)/β-catenin and expression of GSK3β in the hippocampus. These findings suggest that RES exerts anxiolytic- and antidepressant-like effect in SCH rats by downregulating hyperactivity of the HPA axis and regulating both the HPT axis and the Wnt/β-catenin pathway.

Fish oil enhances intestinal barrier function and inhibits corticotropin-releasing hormone/corticotropin-releasing hormone receptor 1 signalling pathway in weaned pigs after lipopolysaccharide challenge.

Stress induces injury in intestinal barrier function in piglets. Long-chain n-3 PUFA have been shown to exhibit potential immunomodulatory and barrier protective effects in animal models and clinical trials. In addition, corticotropin-releasing hormone (CRH)/CRH receptor (CRHR) signalling pathways play an important role in stress-induced alterations of intestinal barrier function. We hypothesised that fish oil could affect intestinal barrier function and CRH/CRHR signalling pathways. In total, thirty-two weaned pigs were allocated to one of four treatments. The experiment consisted of a 2×2 factorial design, and the main factors included immunological challenge (saline or lipopolysaccharide (LPS)) and diet (5 % maize oil or 5 % fish oil). On d 19 of the trial, piglets were treated with saline or LPS. At 4 h after injection, all pigs were killed, and the mesenteric lymph nodes (MLN), liver, spleen and intestinal samples were collected. Fish oil decreased bacterial translocation incidence and the number of translocated micro-organisms in the MLN. Fish oil increased intestinal claudin-1 protein relative concentration and villus height, as well as improved the intestinal morphology. In addition, fish oil supplementation increased intestinal intraepithelial lymphocyte number and prevented elevations in intestinal mast cell and neutrophil numbers induced by LPS challenge. Moreover, fish oil tended to decrease the mRNA expression of intestinal CRHR1, CRH and glucocorticoid receptors. These results suggest that fish oil supplementation improves intestinal barrier function and inhibits CRH/CRHR1 signalling pathway and mast cell tissue density.

Novel stress increases hypothalamic-pituitary-adrenal activity in mice with a raised bite

Background and objectiveIn humans, occlusal disharmony may cause various physical complaints, including head and neck ache, stiffness in the shoulder and neck, and arthrosis of the temporomandibular joints. Occlusal disharmony induced by raising the bite in rodents, increases plasma corticosterone levels, which leads to morphologic changes in the hippocampus and altered hippocampus-related behavior. The paraventricular nucleus (PVN) of the hypothalamus regulates the hypothalamic-pituitary-adrenal system. Chronically stressed animals exposed to a novel stress exhibit higher adrenocorticotropic hormone levels than naive control animals. We hypothesized that there would be different response of the corticotrophin releasing hormone (CRH) and arginine vasopressin (AVP) to a novel acute stress with occlusal disharmony. DesignIn order to investigate how exposure of mice with occlusal disharmony to a novel acute stress (restraint stress) affects the PVN, we induced occlusal disharmony by raising the vertical dimension of the bite (bite-raised condition) and examined the expression of corticotrophin releasing hormone (CRH) mRNA and arginine vasopressin (AVP) mRNA in mouse PVN. ResultsCRH mRNA expression was increased in the PVN of the bite-raised group 90min after the bite-raising procedure, but the expression was recovered to the control level at 14days. AVP mRNA expression in the PVN was normal at 90min, and increased significantly 14days after the bite-raising procedure. Exposure to restraint stress in the bite-raised mice induced a significant increase in CRH mRNA expression in the PVN. ConclusionsThe bite-raising procedure induced a rapid CRH mRNA response and a slower AVP mRNA response in the parvocellular PVN of the hypothalamus. Exposure to a novel stress following the bite-raising procedure further reinforced the CRH stress response. Thus, occlusal disharmony, such as that induced by raising the bite, may be a risk factor for hypersensitivity to a novel stress.

Expression of stress hormones AVP and CRH in the hypothalamus of Mus musculus following water and food deprivation

Neurohypophyseal hormone, arginine vasopressin (AVP), in addition to acting as antidiuretic hormone is also considered to be stress hormone like hypothalamic corticotropin-releasing hormone (CRH). Present study was designed to investigate the relative response of these stress hormones during water and food deprivation. In this study, male laboratory mice of Swiss strain were divided in 5 groups, control – provided water and food ad libitum, two experimental groups water deprived for 2 and 4days respectively (WD2 and WD4) and another two groups food deprived for 2 and 4days respectively (FD2 and FD4). Results indicate an increased expression of AVP mRNA as well as peptide in the hypothalamus of WD2 mice and the expression was further upregulated after 4days of water deprivation but the expression of CRH remained unchanged compare to their respective controls. On the other hand no change was observed in the expression of hypothalamic AVP mRNA while AVP peptide increased significantly in FD2 and FD4 mice compare to control. Further, the expression of CRH mRNA although increased in hypothalamus of both FD2 and FD4 mice, the immunofluorescent staining shows decreased expression of CRH in PVN of food deprived mice. Based on these findings it is concluded that since during osmotic stress only AVP expression is upregulated but during metabolic stress i.e. food deprivation transcription and translation of both the stress hormones are differentially regulated. Further, it is suggested that role of AVP and CRH may be stress specific.