Genotypic variation in the promoter region of the CRH-248 gene interacts with early rearing experiences to disrupt the development of the HPA axis in infant rhesus macaques (Macaca mulatta).

Aberrant functioning of the hypothalamic-pituitary-adrenal (HPA) axis is a hallmark of conditions such as depression, anxiety disorders, and post-traumatic stress disorder. Early-life adversity and genetic variation can interaction to disrupt HPA axis regulation, potentially contributing to certain forms of psychopathology. This study employs a rhesus macaque model to investigate how early parental neglect interacts with a single nucleotide polymorphism within the promoter region of the corticotropin-releasing hormone (CRH-248) gene, impacting the development of the HPA axis. For the initial six months of life, 307 rhesus monkey infants (n = 146 females, n = 161 males) were either reared with their mothers (MR) in conditions emulating the natural environment (control group) or raised without maternal care in groups with constant or 3-hours daily access to same-aged peers (NR). Blood samples collected on days 30, 60, 90, and 120 of life under stressful conditions were assayed for plasma cortisol and adrenocorticotropic hormone (ACTH) concentrations. Findings revealed that NR subjects exhibited a significant blunting of both ACTH and cortisol concentrations. Notably, there was a gene-by-environment interaction observed for ACTH and cortisol levels, with NR subjects with the polymorphism displaying higher ACTH concentrations and lower cortisol concentrations. To the extent that these results generalize to humans, they suggest that early parental neglect may render individuals vulnerable to HPA axis dysfunction, a susceptibility that is modulated by CRH-248 genotype-a gene-by-environment interaction that leaves a lasting developmental signature.

Depression Exacerbates Hepatic Steatosis in C57BL/6J Mice by Activating the Hypothalamic-Pituitary-Adrenal Axis.

BACKGROUND/AIM: Depression is associated with metabolic disorders, including non-alcoholic fatty liver disease (NAFLD). However, the mechanisms underlying the interaction between them are still poorly known. MATERIALS AND METHODS: In this study, mice on a choline deficiency, L-amino acid-defined, high-fat diet (CDAHFD) developing steatosis were challenged with chronic restraint stress (CRS), a protocol widely used to induce depression. The development of depression and steatosis was evaluated using histopathology analysis, ELISA, q-PCR and Western Blot. RESULTS: The contribution of the activated HPA axis to hepatic steatosis progress was fully established, which was validated using a hepatocyte model. Histopathological and biochemical analysis indicated that steatosis was exacerbated by CRS challenge, and behavioral tests indicated that the mice developed depression. Among the screened endocrinal pathways, the hypothalamic-pituitary-adrenal (HPA) axis was significantly activated and the synergistic effect of CDAHFD and CRS in activating the HPA axis was observed. In the hypothalamus, expression of corticotropin-releasing hormone (CRH) was increased by 86% and the protein levels of hypothalamic CRH were upregulated by 25% to 33% by CRS treatment. Plasma CRH levels were elevated by 45-56% and plasma adrenocorticotropic hormone (ACTH) levels were elevated by 29-58% by CRS treatment. In the liver, target genes of the HPA axis were activated, accompanied by disruption of the lipid metabolism and progression of steatohepatitis. The lipid metabolism in the Hepa1-6 cell line treated with endogenous corticosterone (CORT) was in accordance with the aforementioned in vivo responses. CONCLUSION: Depression aggravated hepatic steatosis in CDAHFD-fed mice by activating the HPA axis. The risk of NAFLD development should be fully considered in depressive patients and improvement of psychotic disorders could be an etiological treatment strategy for them.

Mechanisms of Peitu Yimu acupuncture in repairing intestinal mucosal barrier by regulating CRF/CRFR1 pathway in diarrhea-predominant irritable bowel syndrome rats. / 培土抑木针法调节CRF/CRFR1é€šè·¯ä¿®å¤è ¹æ³»åž‹è‚ æ˜“æ¿€ç»¼åˆå¾å¤§é¼ è‚ é»è†œå±éšœçš„æœºåˆ¶ç ”ç©¶.

OBJECTIVES: To investigate the effect of Peitu Yimu(strengthening spleen and soothing liver) acupuncture on intestinal mucosal barrier function and corticotropin-releasing factor (CRF)/CRF receptor 1 (CRFR1) pathway in rats with diarrhea-predominant irritable bowel syndrome (IBS-D), so as to explore its underlying mechanism in alleviating IBS-D. METHODS: Forty female SD rats were randomly divided into blank, model, electroacupuncture (EA), and agonist groups, with 10 rats in each group. Except for the blank group, rats in the other groups were given folium sennae infusion by gavage combined with chronic unpredictable mild stress to establish IBS-D model. Rats in the EA group received acupuncture at "Tianshu"(ST25) and EA at "Zusanli"(ST36) and "Taichong"(LR3) (2 Hz/15 Hz) on one side for 20 min, with the side chosen alternately every other day, for 14 days after modeling. Rats in the agonist group received acupuncture 30 min after intravenous injection of CRFR1 agonist urocortin, with the same manipulation method and time as the EA group. Before and after intervention, visceral pain threshold and stool Bristol scores were measured. Elevated plus maze test and open field test were used to detect anxiety and depression like behavior of rats. ELISA was used to detect the contents of CRF and CRFR1 in rats serum. Immunohistochemistry was used to detect the positive expressions of CRF, CRFR1, zonula occludens protein 1(ZO-1), occlusal protein(Occludin), and closure protein 1 (Claudin-1) in colon tissue. RESULTS: Compared with the blank group, the visceral pain threshold, open arm time percentage (OT%), total distance of movement in the open field test, and positive expression of ZO-1, Occludin, and Claudin-1 in colon were decreased (P<0.01, P<0.05), while Bristol stool scores, serum CRF and CRFR1 contents, and positive expressions of CRF and CRFR1 in colon were increased (P<0.01) in the model group. After intervention and compared with the model group, the visceral pain threshold, OT%, total distance of movement in the open field test, and positive expressions of ZO-1, Occludin, and Claudin-1 in colon were increased (P<0.05, P<0.01), while Bristol stool scores, serum CRF and CRFR1 contents, and positive expressions of CRF and CRFR1 in colon were decreased (P<0.01) in the EA group；the Bristol stool scores, serum CRF content, and CRF positive expression in colon were significantly decreased in the agonist group (P<0.01). CONCLUSIONS: Peitu Yimu acupuncture can significantly improve visceral hypersensitivity and anxiety-depression state in IBS-D rats. Its mechanism may be related to the inhibition of CRF/CRFR1 pathway and restoration of intestinal tight junction protein expressions.

Response of CRH system in brain and gill of marine medaka to seawater acidification.

Corticotropin-releasing hormone (CRH) is mainly secreted by the hypothalamus to regulate stress when environmental factors change. Gills contact with water directly and may also secrete CRH to maintain local homeostasis. Ocean acidification changes water chemical parameters and is becoming an important environmental stressor for marine fish. The response of brain and gill CRH systems to ocean acidification remains unclear. In this study, marine medaka were exposed to CO2-acidified seawater (440 ppm, 1000 ppm, and 1800 ppm CO2) for 2 h, 4 h, 24 h, and 7 d, respectively. At 2 h and 4 h, the expression of crh mRNA in gills increased with increasing CO2 concentration. Crh protein is expressed mainly in the lamellae cells. crhbp and crhr1 expression also increased significantly. However, at 2 h and 4 h, acidification caused little changes in these genes and Crh protein expression in the brain. At 7 d, Crh-positive cells were detected in the hypothalamus; moreover, Crh protein expression in the whole brain increased. It is suggested that CRH autocrine secretion in gills is responsible for local acid-base regulation rather than systemic mobilization after short-term acidification stress, which may help the rapid regulation of body damage caused by environmental stress.

Two Adverse Early Life Events Induce Differential Changes in Brain CRH and Serotonin Systems in Rats along with Hyperphagia and Depression.

BACKGROUND: Different types of stress inflicted in early stages of life elevate the risk, among adult animals and humans, to develop disturbed emotional-associated behaviors, such as hyperphagia or depression. Early-life stressed (ELS) adults present hyperactivity of the hypothalamus-pituitary-adrenal (HPA) axis, which is a risk factor associated with mood disorders. However, the prevalence of hyperphagia (17%) and depression (50%) is variable among adults that experienced ELS, suggesting that the nature, intensity, and chronicity of the stress determines the specific behavioral alteration that those individuals develop. METHODS: We analyzed corticosterone serum levels, Crh, GR, Crhr1 genes expression in the hypothalamic paraventricular nucleus, amygdala, and hippocampus due to their regulatory role on HPA axis in adult rats that experienced maternal separation (MS) or limited nesting material (LNM) stress; as well as the serotonergic system activity in the same regions given its association with the corticotropin-releasing hormone (CRH) pathway functioning and with the hyperphagia and depression development. RESULTS: Alterations in dams' maternal care provoked an unresponsive or hyper-responsive HPA axis function to an acute stress in MS and LNM adults, respectively. The differential changes in amygdala and hippocampal CRH system seemed compensating alterations to the hypothalamic desensitized glucocorticoids receptor (GR) in MS or hypersensitive in LNM. However, both adult animals developed hyperphagia and depression-like behavior when subjected to the forced-swimming test, which helps to understand that both hypo and hypercortisolemic patients present those disorders. CONCLUSION: Different ELS types induce neuroendocrine, brain CRH and 5-hydroxytriptamine (5-HT) systems' alterations that may interact converging to develop similar maladaptive behaviors.

Blood mRNA expression levels of glucocorticoid receptors and FKBP5 are associated with depressive disorder and altered HPA axis.

BACKGROUND: While depression has been associated with alterations in the hypothalamic-pituitary adrenal (HPA) axis function, there is still controversy regarding the nature and extent of the dysfunction, such as in the debate about hypercortisolism vs. hypocortisolism. It may therefore be necessary to understand whether and how HPA axis function in depression is linked to mRNA expression of key genes regulating this system. METHODS: We studied 163 depressed outpatients, most of whom were chronically ill, and 181 healthy controls. Blood mRNA expression levels of NR3C1 (including GRα, GRß, and GR-P isoforms), FKBP4, and FKBP5 were measured at baseline. HPA axis feedback sensitivity was measured by the dexamethasone (Dex)/corticotropin-releasing hormone (CRH) test. The association between mRNA expression levels and HPA axis feedback sensitivity was examined. RESULTS: Compared to controls, patients showed significantly higher expression of GRα and lower expression of FKBP5, and higher post-Dex cortisol levels, even after controlling for age and sex. FKBP5 expression was significantly positively correlated with cortisol levels in patients, while GRα expression was significantly negatively correlated with cortisol levels in controls. LIMITATIONS: Most patients were taking psychotropic medications. The large number of correlation tests may have caused type I errors. CONCLUSIONS: The tripartite relationship between depression, mRNA expression of GR and FKBP5, and HPA axis function suggests that the altered gene expression affects HPA axis dysregulation and, as a result, impacts the development and/or illness course of depressive disorder. The combination of increased GRα expression and decreased FKBP5 expression may serve as a biomarker for chronic depression.

The expression of corticotropin-releasing hormone family peptides in premalignant and malignant vulvar lesions.

OBJECTIVES: To examine the relation of corticotropin-releasing hormone (CRH) family peptides with inflammatory processes and oncogenesis, emphasizing in vulvar inflammatory, premalignant and malignant lesions, as well as to investigate the possibility of lesion cells immunoescaping, utilizing FAS/FAS-L complex. METHODS: Immunohistochemical expression of CRH, urocortin (UCN), FasL and their receptors CRHR1, CRHR2 and Fas was studied in vulvar tissue sections obtained from patients with histologically confirmed diagnosis of lichen, vulvar intraepithelial neoplasia (VIN) and vulvar squamous cell carcinoma (VSCC). The patient cohort was selected from a tertiary teaching Hospital in Greece, between 2005 and 2015. For each of the disease categories, immunohistochemical staining was evaluated and the results were statistically compared. RESULTS: A progressive increase of the cytoplasmic immunohistochemical expression of CRH and UCN, from precancerous lesions to VSCC was observed. A similar increase was detected for Fas and FasL expression. Nuclear localization of UCN was demonstrated in both premalignant and VSCC lesions, with staining being significantly intensified in carcinomas, particularly in the less differentiated tumor areas or in the areas at invasive tumor front. CONCLUSIONS: Stress response system and CRH family peptides seem to have a role in inflammation maintenance and progression of vulvar premalignant lesions to malignancy. It seems that stress peptides may locally modulate the stroma through Fas/FasL upregulation, possibly contributing to vulvar cancer development.

Exploring transcriptional and post-transcriptional epigenetic regulation of crf and 11ßhsd2 in rainbow trout brain during chronic social stress.

Using dominance hierarchies in juvenile rainbow trout (Oncorhynchus mykiss) as a model of chronic social stress in fish, we explored whether epigenetic transcriptional and post-transcriptional mechanisms are involved in the gene expression of corticotropin-releasing factor (crf) and 11ß-hydroxysteroid dehydrogenase (11ßhsd2), key factors involved in the regulation of the endocrine stress axis response. In juvenile rainbow trout pairs, subordinate individuals display sustained elevation of circulating cortisol concentrations. Cortisol production is controlled by the hypothalamic-pituitary-interrenal (HPI) axis in fish and initiated by CRF release from the preoptic area (POA). Given that crf is modulated during chronic social stress, and that such stress has been implicated in the epigenetic regulation of crf in other taxa, we probed a role for epigenetic regulation of crf transcript abundance in chronically stressed rainbow trout. We also investigated the regulation of the cortisol-metabolising enzyme 11ßhsd2 in the POA, which is upregulated in subordinates. The potential involvement of DNA methylation and microRNAs (miRNAs) in the regulation of crf transcript abundance was investigated during social stress in the POA of fish, as was the potential involvement of miRNAs in 11ßhsd2 regulation. Although transcript abundances of crf were elevated in subordinate fish after 4 days, DNA methylation profiles within putative promoter sequences upstream of the crf gene were not significantly affected by chronic stress. An inverse relationship between crf and its predicted posttranscriptional regulator miR-103a-3p in the POA suggests that miRNAs may be involved in mediating the effects of chronic social stress on key components of the endocrine stress axis.

Downregulation of CRHBP is associated with trophoblast invasion inhibition in recurrent pregnancy loss.

In brief: Corticotropin-releasing hormone binding protein (CRHBP) is fundamental to the stress response and plays an important role in parturition during pregnancy. This study shows that abnormal CRHBP expression could be an early warning sign of recurrent pregnancy loss and that CRHBP knockdown could suppress HTR8/SVneo cell invasion by the PKC signaling pathway via interacting with CRH receptor 2. Abstract: Trophoblast invasion is critical for placentation and pregnancy success. Trophoblast dysfunction results in many pregnancy complications, including recurrent pregnancy loss (RPL). Corticotropin-releasing hormone binding protein (CRHBP) is fundamental to the stress response and plays an important role in parturition during pregnancy via binding with CRH. To further characterize its function in early pregnancy, we explored the expression of CRHBP in villi during early pregnancy. Compared with normal pregnant women, we demonstrated that the expression of CRHBP decreased in the trophoblasts and villi in RPL patients and that knockdown of CRHBP expression could suppress HTR8/SVneo cell invasion significantly. Our further exploration indicated that the capacity of CRHBP for regulating trophoblast invasion was associated with the PKC signaling pathway via interacting with CRH receptor 2. These findings might provide a new fundamental mechanism for successful pregnancy and a new diagnostic and therapeutic target for RPL.

Single-cell transcriptomic analysis reveals rich pituitary-Immune interactions under systemic inflammation.

The pituitary represents an essential hub in the hypothalamus-pituitary-adrenal (HPA) axis. Pituitary hormone-producing cells (HPCs) release several hormones to regulate fundamental bodily functions under normal and stressful conditions. It is well established that the pituitary endocrine gland modulates the immune system by releasing adrenocorticotropic hormone (ACTH) in response to neuronal activation in the hypothalamus. However, it remains unclear how systemic inflammation regulates the transcriptomic profiles of pituitary HPCs. Here, we performed single-cell RNA-sequencing (scRNA-seq) of the mouse pituitary and revealed that upon inflammation, all major pituitary HPCs respond robustly in a cell type-specific manner, with corticotropes displaying the strongest reaction. Systemic inflammation also led to the production and release of noncanonical bioactive molecules, including Nptx2 by corticotropes, to modulate immune homeostasis. Meanwhile, HPCs up-regulated the gene expression of chemokines that facilitated the communication between the HPCs and immune cells. Together, our study reveals extensive interactions between the pituitary and immune system, suggesting multifaceted roles of the pituitary in mediating the effects of inflammation on many aspects of body physiology.

Musclin prevents depression-like behavior in male mice by activating urocortin 2 signaling in the hypothalamus.

Introduction: Physical activity is recommended as an alternative treatment for depression. Myokines, which are secreted from skeletal muscles during physical activity, play an important role in the skeletal muscle-brain axis. Musclin, a newly discovered myokine, exerts physical endurance, however, the effects of musclin on emotional behaviors, such as depression, have not been evaluated. This study aimed to access the anti-depressive effect of musclin and clarify the connection between depression-like behavior and hypothalamic neuropeptides in mice. Methods: We measured the immobility time in the forced swim (FS) test, the time spent in open arm in the elevated-plus maze (EPM) test, the mRNA levels of hypothalamic neuropeptides, and enumerated the c-Fos-positive cells in the paraventricular nucleus (PVN), arcuate nucleus (ARC), and nucleus tractus solitarii (NTS) in mice with the intraperitoneal (i.p.) administration of musclin. Next, we evaluated the effects of a selective corticotropin-releasing factor (CRF) type 1 receptor antagonist, selective CRF type 2 receptor antagonist, melanocortin receptor (MCR) agonist, and selective melanocortin 4 receptor (MC4R) agonist on changes in behaviors induced by musclin. Finally we evaluated the antidepressant effect of musclin using mice exposed to repeated water immersion (WI) stress. Results: We found that the i.p. and i.c.v. administration of musclin decreased the immobility time and relative time in the open arms (open %) in mice and increased urocortin 2 (Ucn 2) levels but decreased proopiomelanocortin levels in the hypothalamus. The numbers of c-Fos-positive cells were increased in the PVN and NTS but decreased in the ARC of mice with i.p. administration of musclin. The c-Fos-positive cells in the PVN were also found to be Ucn 2-positive. The antidepressant and anxiogenic effects of musclin were blocked by central administration of a CRF type 2 receptor antagonist and a melanocortin 4 receptor agonist, respectively. Peripheral administration of musclin also prevented depression-like behavior and the decrease in levels of hypothalamic Ucn 2 induced by repeated WI stress. Discussion: These data identify the antidepressant effects of musclin through the activation of central Ucn 2 signaling and suggest that musclin and Ucn 2 can be new therapeutic targets and endogenous peptides mediating the muscle-brain axis.

Growth differentiation factor-15 stimulates the synthesis of corticotropin-releasing factor in hypothalamic 4B cells.

Growth differentiation factor-15 (GDF15) is a stress-activated cytokine that regulates cell growth and inflammatory and stress responses. We previously reported the role and regulation of GDF15 in pituitary corticotrophs. Dexamethasone increases Gdf15 gene expression levels and production. GDF15 suppresses adrenocorticotropic hormone synthesis in pituitary corticotrophs and subsequently mediates the negative feedback effect of glucocorticoids. Here, we analyzed corticotropin-releasing factor (Crf) promoter activity in hypothalamic 4B cells transfected with promoter-driven luciferase reporter constructs. The effects of time and GDF15 concentration on Crf mRNA levels were analyzed using quantitative real-time polymerase chain reaction. Glial cell-derived neurotrophic factor family receptor α-like (GFRAL) protein is expressed in 4B cells. GDF15 increased Crf promoter activity and Crf mRNA levels in 4B cells. The protein kinase A and C pathways also contributed to the GDF15-induced increase in Crf gene expression. GDF15 stimulates GFRAL, subsequently increasing the phosphorylation of AKT, an extracellular signal-related kinase, and the cAMP response element-binding protein. Therefore, GDF15-dependent pathways may be involved in regulating Crf expression under stressful conditions in hypothalamic cells.

Repeated toluene and cyclohexane inhalation produces differential effects on HPA and HPT axes in adolescent male rats.

Misused volatile solvents typically contain toluene (TOL) as the main psychoactive ingredient. Cyclohexane (CHX) can also be present and is considered a safer alternative. Solvent misuse often occurs at early stages of life, leading to permanent neurobehavioral impairment and growth retardation. However, a comprehensive examination of the effects of TOL and CHX on stress regulation and energy balance is lacking. Here, we compared the effect of a binge-pattern exposure to TOL or CHX (4,000 or 8,000 ppm) on body weight, food intake, the hypothalamus-pituitary-adrenal (HPA) and hypothalamus-pituitary-thyroid (HPT) axes in male adolescent Wistar rats. At 8,000 ppm, TOL decreased body weight gain without affecting food intake. In addition, TOL and CHX altered the HPA and HPT axes' function in a solvent- and concentration-dependent manner. The highest TOL concentration produced HPA axis hyperactivation in animals not subjected to stress, which was evidenced by increased corticotropin-releasing-factor (CRF) release from the median eminence (ME), elevated adrenocorticotropin hormone (ACTH) and corticosterone serum levels, and decreased CRF mRNA levels in the hypothalamic paraventricular nucleus (PVN). TOL (8,000 ppm) also increased triiodothyronine (T3) serum levels, decreased pro-thyrotropin-releasing-hormone (pro-TRH) mRNA transcription in the PVN, pro-TRH content in the ME, and serum thyroid stimulating hormone (TSH) levels. CHX did not affect the HPA axis. We propose that the increased HPT axis activity induced by TOL can be related to the impaired body weight gain associated with inhalant misuse. These findings may contribute to a better understanding of the effects of the misused solvents TOL and CHX.

CRH-R2 signalling modulates feeding and circadian gene expression in hypothalamic mHypoA-2/30 neurons.

The hypothalamic type 2 corticotropin releasing hormone receptor (CRH-R2) plays critical roles in homeostatic regulation, particularly in fine tuning stress recovery. During acute stress, the CRH-R2 ligands CRH and urocortins promote adaptive responses and feeding inhibition. However, in rodent models of chronic stress, over-exposure of hypothalamic CRH-R2 to its cognate agonists is associated with urocortin 2 (Ucn2) resistance; attenuated cAMP-response element binding protein (CREB) phosphorylation and increased food intake. The molecular mechanisms involved in these altered CRH-R2 signalling responses are not well described. In the present study, we used the adult mouse hypothalamus-derived cell line mHypoA-2/30 to investigate CRH-R2 signalling characteristics focusing on gene expression of molecules involved in feeding and circadian regulation given the role of clock genes in metabolic control. We identified functional CRH-R2 receptors expressed in mHypoA-2/30 cells that differentially regulate CREB and AMP-activated protein kinase (AMPK) phosphorylation and downstream expression of the appetite-regulatory genes proopiomelanocortin (Pomc) and neuropeptide Y (Npy) in accordance with an anorexigenic effect. We studied for the first time the effects of Ucn2 on clock genes in native and in a circadian bioluminescence reporter expressing mHypoA-2/30 cells, detecting enhancing effects of Ucn2 on mRNA levels and rhythm amplitude of the circadian regulator Aryl hydrocarbon receptor nuclear translocator-like protein 1 (Bmal1), which could facilitate anorexic responses in the activity circadian phase. These data uncover novel aspects of CRH-R2 hypothalamic signalling that might be important in regulation of circadian feeding during stress responses.

Effect of hypoxia duration and pattern on channel Catfish (Ictalurus punctatus) neuropeptide gene expression and hematology.

Commercial aquaculture production of channel catfish (Ictalurus punctatus) occurs in shallow ponds with daily cycling of dissolved oxygen concentration ranging from supersaturation to severe hypoxia. Once daily minimum dissolved oxygen concentration falls below 3.0 mg O2/L, channel catfish have a reduced appetite, leading to reduced growth rates. In other fishes, upregulation of the neuropeptides corticotropin-releasing factor (CRF) and urotensin I (UI) have been implicated as initiating the mechanism responsible for decreasing appetite once an environmental stressor is detected. Channel catfish maintained at 27 °C in aquaria were subjected to varying durations and patterns of hypoxia (1.75 ± 0.07 mg O2/L) to evaluate underlying physiological responses to hypoxia and determine if hypothalamic CRF and UI are responsible for hypoxia-induced anorexia in channel catfish. During a short exposure to hypoxia (12 h), venous PO2 was significantly lower within 6 h and was coupled with an increase of hematocrit and decrease of blood osmolality, yet all responses reversed within 12 h after returning to normoxia. When this pattern of hypoxia and normoxia was repeated cyclically for 5 days, these physiological responses repeated daily. Extended periods of hypoxia (5 days) resulted in similar hematological responses, which did not recover to baseline values during the hypoxia exposure. This study did not find a significant change in hypothalamic transcription of CRF and UI during hypoxia challenges but did identify multiple physiological adaptive responses that work together to reduce the severity of experimentally induced hypoxia in channel catfish.

Novel corticotropin-releasing hormone receptor genes (CRHR1 and CRHR2) linkage to and association with polycystic ovary syndrome.

BACKGROUND: Women with polycystic ovarian syndrome (PCOS) have increased hypothalamic-pituitary-adrenal (HPA) axis activation, pro-inflammatory mediators, and psychological distress in response to stressors. In women with PCOS, the corticotropin-releasing hormone (CRH) induces an exaggerated HPA response, possibly mediated by one of the CRH receptors (CRHR1 or CRHR2). Both CRHR1 and CRHR2 are implicated in insulin secretion, and variants in CRHR1 and CRHR2 genes may predispose to the mental-metabolic risk for PCOS. METHODS: We phenotyped 212 Italian families with type 2 diabetes (T2D) for PCOS following the Rotterdam diagnostic criteria. We analyzed within CRHR1 and CRHR2 genes, respectively, 36 and 18 microarray-variants for parametric linkage to and/or linkage disequilibrium (LD) with PCOS under the recessive with complete penetrance (R1) and dominant with complete penetrance (D1) models. Subsequentially, we ran a secondary analysis under the models dominant with incomplete penetrance (D2) and recessive with incomplete penetrance (R2). RESULTS: We detected 22 variants in CRHR1 and 1 variant in CRHR2 significantly (p < 0.05) linked to or in LD with PCOS across different inheritance models. CONCLUSIONS: This is the first study to report CRHR1 and CRHR2 as novel risk genes in PCOS. In silico analysis predicted that the detected CRHR1 and CRHR2 risk variants promote negative chromatin activation of their related genes in the ovaries, potentially affecting the female cycle and ovulation. However, CRHR1- and CRHR2-risk variants might also lead to hypercortisolism and confer mental-metabolic pleiotropic effects. Functional studies are needed to confirm the pathogenicity of genes and related variants.

Mapping of corticotropin-releasing factor, receptors, and binding protein mRNA in the chicken telencephalon throughout development.

Understanding the neural mechanisms that regulate the stress response is critical to know how animals adapt to a changing world and is one of the key factors to be considered for improving animal welfare. Corticotropin-releasing factor (CRF) is crucial for regulating physiological and endocrine responses, triggering the activation of the sympathetic nervous system and the hypothalamo-pituitary-adrenal axis (HPA) during stress. In mammals, several telencephalic areas, such as the amygdala and the hippocampus, regulate the autonomic system and the HPA responses. These centers include subpopulations of CRF containing neurons that, by way of CRF receptors, play modulatory roles in the emotional and cognitive aspects of stress. CRF binding protein also plays a role, buffering extracellular CRF and regulating its availability. CRF role in activation of the HPA is evolutionary conserved in vertebrates, highlighting the relevance of this system to help animals cope with adversity. However, knowledge on CRF systems in the avian telencephalon is very limited, and no information exists on detailed expression of CRF receptors and binding protein. Knowing that the stress response changes with age, with important variations during the first week posthatching, the aim of this study was to analyze mRNA expression of CRF, CRF receptors 1 and 2, and CRF binding protein in chicken telencephalon throughout embryonic and early posthatching development, using in situ hybridization. Our results demonstrate an early expression of CRF and its receptors in pallial areas regulating sensory processing, sensorimotor integration and cognition, and a late expression in subpallial areas regulating the stress response. However, CRF buffering system develops earlier in the subpallium than in the pallium. These results help to understand the mechanisms underlying the negative effects of noise and light during prehatching stages in chicken, and suggest that stress regulation becomes more sophisticated with age.

Vasopressin Expressed in Hypothalamic CRF Neurons Causes Impaired Water Diuresis in Secondary Adrenal Insufficiency.

Patients with secondary adrenal insufficiency can present with impaired free water excretion and hyponatremia, which is due to the enhanced secretion of vasopressin (AVP) despite increased total body water. AVP is produced in magnocellular neurons in the paraventricular nucleus of the hypothalamus (PVH) and supraoptic nucleus and in parvocellular corticotropin-releasing factor (CRF) neurons in the PVH. This study aimed to elucidate whether magnocellular AVP neurons or parvocellular CRF neurons coexpressing AVP are responsible for the pathogenesis of hyponatremia in secondary adrenal insufficiency. The number of CRF neurons expressing copeptin, an AVP gene product, was significantly higher in adrenalectomized AVP-floxed mice (AVPfl/fl) than in sham-operated controls. Adrenalectomized AVPfl/fl mice supplemented with aldosterone showed impaired water diuresis under ad libitum access to water or after acute water loading. They became hyponatremic after acute water loading, and it was revealed under such conditions that aquaporin-2 (AQP2) protein levels were increased in the kidney. Furthermore, translocation of AQP2 to the apical membrane was markedly enhanced in renal collecting duct epithelial cells. Remarkably, all these abnormalities observed in the mouse model for secondary adrenal insufficiency were ameliorated in CRF-AVP-/- mice that lacked AVP in CRF neurons. Our study demonstrates that CRF neurons in the PVH are responsible for the pathogenesis of impaired water excretion in secondary adrenal insufficiency.

Sex-dependent association of DNA methylation in the coding region of the corticotropin-releasing hormone gene and schizophrenia spectrum disorder.

BACKGROUND: Schizophrenia spectrum disorder (SSD) is a common mental disorder causing severe and chronic disability. Epigenetic changes in genes related to the hypothalamic-pituitary-adrenal (HPA) axis are believed to play an important role in SSD pathogenesis. The methylation status of the corticotropin-releasing hormone (CRH) gene, which is central to the HPA axis, has not been investigated in patients with SSD. AIM: We investigated the methylation status of the coding region of the CRH gene (hereafter, CRH methylation) using peripheral blood samples from patients with SSD. SUBJECTS AND METHODS: We used sodium bisulphite and MethylTarget to determine CRH methylation after collecting peripheral blood samples from 70 patients with SSD who had positive symptoms and 68 healthy controls. RESULTS: CRH methylation was significantly increased in patients with SSD, especially in male patients. CONCLUSIONS: Differences in CRH methylation were detectable in the peripheral blood of patients with SSD. Epigenetic abnormalities in the CRH gene were closely related to positive symptoms of SSD, suggesting that epigenetic processes may mediate the pathophysiology of SSD.

Chronic UCN2 treatment desensitizes CRHR2 and improves insulin sensitivity.

Urocortin 2 (UCN2) acts as a ligand for the G protein-coupled receptor corticotropin-releasing hormone receptor 2 (CRHR2). UCN2 has been reported to improve or worsen insulin sensitivity and glucose tolerance in vivo. Here we show that acute dosing of UCN2 induces systemic insulin resistance in male mice and skeletal muscle. Inversely, chronic elevation of UCN2 by injection with adenovirus encoding UCN2 resolves metabolic complications, improving glucose tolerance. CRHR2 recruits Gs in response to low concentrations of UCN2, as well as Gi and ß-Arrestin at high concentrations of UCN2. Pre-treating cells and skeletal muscle ex vivo with UCN2 leads to internalization of CRHR2, dampened ligand-dependent increases in cAMP, and blunted reductions in insulin signaling. These results provide mechanistic insights into how UCN2 regulates insulin sensitivity and glucose metabolism in skeletal muscle and in vivo. Importantly, a working model was derived from these results that unifies the contradictory metabolic effects of UCN2.