Regulation Of Corticotropin-releasing Hormone Research Articles

Extracellular Nampt (eNampt/visfatin/PBEF) directly and indirectly stimulates ACTH and CCL2 protein secretion from isolated rat corticotropes.

Nicotinamide phosphoribosyltransferase (Nampt/visfatin/PBEF) acts both as an enzyme in the nicotinamide adenine dinucleotide (NAD) synthesis pathway as well as an extracellular hormone (eNampt). Among its effects, eNampt exerts potent pro-inflammatory effects. We have recently shown that, in rats, eNampt stimulates corticosterone secretion by acting through the pituitary rather than the hypothalamus. To investigate the mechanism of action of eNampt on the secretion of adrenocorticotropic hormone (ACTH) and chemokine (C-C motif) ligand 2 (CCL2), which are cytokines secreted by pituitary neuroendocrine tumors. The research was carried out on the AtT-20 murine cell line, primary rat pituitary cell culture, isolated pituitary corticotropes, and in vivo. The effects of the performed experiments were examined using the following methods: gene expression profiling using microarrays, quantitative polymerase chain reaction (qPCR) and enzyme-linked immunosorbent assay (ELISA). The results suggest that eNampt stimulates ACTH secretion from rat corticotropes both directly and indirectly. Indirect action most likely occurs through interleukin (IL)-6 secreted by folliculostellate cells of the pituitary gland. In isolated ACTH cells of the rat pituitary gland, eNampt stimulates the expression of genes involved in the immune response. Among them, the protein encoded by the CCL2 gene seems to also be involved in the regulation of corticotropin-releasing hormone (CRH)-dependent metabolism. Unlike rat corticotropes, murine AtT-20 corticotropic cells do not react to either eNampt or Fk866 (the inhibitor of Nampt enzymatic action). The eNampt stimulates the secretion of ACTH from rat corticotropes indirectly and directly, likely by stimulating IL-6 secretion from folliculostellate cells of the pituitary gland. This effect was not observed in the AtT-20 corticotropic cell cancer cell line.

Secretagogin Mediates the Regulatory Effect of Electroacupuncture on Hypothalamic-Pituitary-Adrenal Axis Dysfunction in Surgical Trauma.

Electroacupuncture (EA) improves hypothalamic-pituitary-adrenal (HPA) axis disorder by reducing corticotropin-releasing hormone (CRH) synthesis and release in the paraventricular nucleus (PVN). However, the potential mechanism underlying CRH regulation remains unclear. Secretagogin (SCGN) is closely related to stress and is involved in regulating the release of CRH. We hypothesized that SCGN in the PVN might trigger the HPA system and be involved in EA-mediated modulation of HPA dysfunction caused by surgical trauma. Serum CRH and adrenocorticotropic hormone (ACTH) and plasma corticosterone (CORT) levels at 6 h and 24 h after hepatectomy were determined by radioimmunoassay. CRH and SCGN protein levels in the PVN were detected by western blot and immunofluorescence, and CRH and SCGN mRNA levels in the PVN were determined by means of real-time polymerase chain reaction (RT-PCR) and in situ hybridization (ISH). Our studies showed that serum CRH, ACTH, and CORT levels and PVN CRH expression were significantly increased at 6 h and 24 h after hepatectomy in the hepatectomy group compared with the control group, and those in the EA+hepatectomy group were decreased compared with those in the hepatectomy group. The protein and mRNA levels of SCGN in the PVN were also increased after hepatectomy, and their expression in the EA+hepatectomy group was decreased compared with that in the hepatectomy group. When SCGN expression in the PVN was functionally knocked down by a constructed CsCI virus, we found that SCGN knockdown decreased the serum CRH, ACTH, and CORT levels in the SCGN shRNA+hepatectomy group compared with the hepatectomy group, and it also attenuated CRH expression in the PVN. In summary, our findings illustrated that EA normalized HPA axis dysfunction after surgical trauma by decreasing the transcription and synthesis of SCGN.

The role of glucocorticoids and corticotropin-releasing hormone regulation on anxiety symptoms and response to treatment.

The stress response has been linked to the expression of anxiety and depression, but the mechanisms for these connections are under continued consideration. The activation and expression of glucocorticoids and CRH are variable and may hold important clues to individual experiences of mood disorders. This paper explores the interactions of glucocorticoids and CRH in the presentation of anxiety and depressive disorders in an effort to better describe their differing roles in each of these clinical presentations. In addition, it focuses on ways in which extra-hypothalamic glucocorticoids and CRH, often overlooked, may play important roles in the presentation of clinical disorders.

Regulation of corticotropin-releasing hormone.

Regulation of corticotropin-releasing hormone.

Homeodomain Protein Otp and Activity-Dependent Splicing Modulate Neuronal Adaptation to Stress

Regulation of corticotropin-releasing hormone (CRH) activity is critical for the animal's adaptation to stressful challenges, and its dysregulation is associated with psychiatric disorders in humans. However, the molecular mechanism underlying this transcriptional response to stress is not well understood. Using various stress paradigms in mouse and zebrafish, we show that the hypothalamic transcription factor Orthopedia modulates the expression of CRH as well as the splicing factor Ataxin 2-Binding Protein-1 (A2BP1/Rbfox-1). We further show that the G protein coupled receptor PAC1, which is a known A2BP1/Rbfox-1 splicing target and an important mediator of CRH activity, is alternatively spliced in response to a stressful challenge. The generation of PAC1-hop messenger RNA isoform by alternative splicing is required for termination of CRH transcription, normal activation of the hypothalamic-pituitary-adrenal axis and adaptive anxiety-like behavior. Our study identifies an evolutionarily conserved biochemical pathway that modulates the neuronal adaptation to stress throughtranscriptional activation and alternative splicing.

Steroid receptor coactivator-1 is necessary for regulation of corticotropin-releasing hormone by chronic stress and glucocorticoids

Adaptation to stress in vertebrates occurs via activation of hormonal and neuronal signaling cascades in which corticotropin-releasing hormone (CRH) plays a central role. Expression of brain CRH is subject to strong, brain-region specific regulation by glucocorticoid hormones and neurogenic intracellular signals. We hypothesized that Steroid Receptor Coactivator 1 (SRC-1), a transcriptional coregulator of the glucocorticoid receptor, is involved in the sensitivity of CRH regulation by stress-related factors. In the brains of SRC-1 knockout mice we found basal CRH mRNA levels to be lower in the central nucleus of the amygdala. Hypothalamic CRH up-regulation after chronic (but not acute) stress, as well as region-dependent up- and down-regulation induced by synthetic glucocorticoids, were significantly attenuated compared with wild type. The impaired induction of the crh gene by neurogenic signals was corroborated in AtT-20 cells, where siRNA and overexpression experiments showed that SRC-1 is necessary for full induction of a CRH promoter reporter gene by forskolin, suggestive of involvement of transcription factor CREB. In conclusion, SRC-1 is involved in positive and negative regulation of the crh gene, and an important factor for the adaptive capacity of stress.

Corticotropin-releasing hormone activates connexin 43 via activator protein-1 transcription factor in human myometrial smooth muscle cells

Corticotropin-releasing hormone (CRH) and connexin 43 (Cx43) play crucial roles in uterine contraction and the onset of labor. The aim of the present study was to investigate the regulatory effects of CRH on Cx43 expression in human myometrial smooth muscle cells (SMCs) and, potentially, its activation of the c-Fos/activator protein (AP)-1 signaling pathway. Human myometrial SMCs collected from nonpregnant women were treated with different concentrations of CRH. Transient transfection of AP-1 decoy oligodeoxynucleotide (ODN) was used to block AP-1 sites of Cx43. The transcriptional activity of AP-1 was detected by luciferase assay. Cx43 protein expression was visualized by immunofluorescence staining. mRNA and protein expression of c-Fos and Cx43 were demonstrated by real-time quantitative RT-PCR and Western blot, respectively. CRH facilitated Cx43 expression and enhanced AP-1 promoter activity in human uterine SMCs. After CRH treatment, Cx43 expression in the cytoplasm increased significantly. CRH significantly increased mRNA and protein expression of c-Fos and Cx43 in a dose-dependent manner (P < 0.01). A transient transfection of AP-1 decoy ODN did not affect CRH regulation of c-Fos (P > 0.05) but almost completely abolished CRH-induced enhancement of Cx43 expression (P < 0.01). In human primary myometrial SMCs, CRH enhances Cx43 mRNA and protein expression through upregulation of c-Fos expression. Blockade of AP-1 sites to the Cx43 promoter can neutralize the CRH-induced upregulation of Cx43.

Estrogen Receptor (ER)β Isoforms Rather Than ERα Regulate Corticotropin-Releasing Hormone Promoter Activity through an Alternate Pathway

The hypothalamic-pituitary-adrenal axis regulates mammalian stress responses by secreting glucocorticoids. The magnitude of the response is in part determined by gender, for in response to a given stressor, circulating glucocorticoids reach higher levels in female rats than in males. This gender difference could result from estrogen regulation of the corticotropin-releasing hormone (CRH) promoter via either of its receptors: estrogen receptor (ER)alpha or ERbeta. Immunocytochemistry revealed that a subset (12%) of medial parvocellular CRH neurons in the rat hypothalamus contain ERbeta but not ERalpha. To determine whether ERs could regulate CRH promoter activity, we cotransfected cells with a CRH promoter construct and either ERalpha or individual ERbeta isoforms. ERalpha weakly stimulated CRH promoter transcriptional activity in a ligand-independent manner. Conversely, all ERbeta isoforms tested stimulated CRH promoter activity with different ligand profiles. ERbeta1 and ERbeta2delta3 displayed constitutive activity (ERbeta1 more than ERbeta2delta3). Ligand-dependent activity of beta isoforms 1 and 2 was altered by an Exon3 splice variant (delta3) or by the additional 18 amino acids in the ligand-binding domain of ERbeta2 isoforms. Lastly, we suggest that ER regulation of CRH takes place through an alternate pathway, one that requires protein-protein interactions with other transcription factors or their associated complexes. However, a pure ER-activator protein-1 alternate pathway does not appear to be involved.

Corticotropin-releasing hormone deficiency increases allergen-induced airway inflammation in a mouse model of asthma

Corticotropin-releasing hormone (CRH) is a major regulator of adrenocorticotropic hormone and the production of glucocorticoids by the adrenal gland. Abnormal regulation of CRH and endogenous glucocorticoids has been implicated in the pathogenesis of asthma. We postulated that CRH deficiency could increase asthma severity by disrupting hypothalamus-pituitary-adrenal axis function and the induction of glucocorticoids through inflammatory and physiologic stress. However, CRH is expressed by several types of immune cells and might be induced at sites of inflammation, where it has local immunostimulatory actions. Thus CRH deficiency could decrease asthma severity. To test these possibilities, we subjected CRH-knockout mice to an ovalbumin-induced airway inflammation protocol that mimics many features of asthma. CRH-knockout mice had an increase in airway inflammation of approximately 80% to 300% and an increase in goblet cell hyperplasia of approximately 70% compared with wild-type mice. In contrast, IgE induction was unaffected by CRH deficiency. The increased inflammation in knockout mice was associated with increased tissue resistance, elastance, and hysteresivity. Levels of IL-4, IL-5, IL-13, RANTES, IFN-gamma, and eotaxin were all increased in knockout mice. Serum corticosterone levels were decreased in knockout mice and might account for some of the differences between knockout and wild-type mice. We conclude that CRH deficiency disrupts endogenous glucocorticoid production and enhances allergen-induced airway inflammation and lung mechanical dysfunction in mice. Thus inherited or acquired CRH deficiency could increase asthma severity in human subjects.

GABAergic mechanisms constraining the activity of the hypothalamo-pituitary-adrenocortical axis.

There are two major inhibitory mechanisms that constrain the activity of the hypothalamo-pituitary-adrenocortical axis: the hormonal negative feedback and the neural inhibition including that posed by the GABAergic neurons. This chapter summarizes our recent morphologic and functional findings on the role of gamma-aminobutyric acid (GABA) in the transcriptional regulation of hypophyseotropic neuropeptide genes in the parvocellular neurosecretory cells of the hypothalamic paraventricular nucleus (PVH). We used organotypic hypothalamic slice cultures and in vivo microinjection protocols in combination with in situ histologic and ultrastructural procedures to address the role of local interneurons in the regulation of hypothalamic effector neurons. Under basal conditions, an intrinsic GABAergic mechanism in the PVH microenvironment was revealed that by itself, without limbic contribution, impinged a tonic inhibitory influence on the parvocellular corticotropin-releasing hormone (CRH) neurons in vitro. In vivo, remote inputs were superimposed on the local circuit, allowing differential transcriptional regulation of CRH and arginine vasopressin (AVP) genes in the hypophyseotropic neurons. During stress, GABAergic cells that are known to project to the PVH become activated and are involved in restraining the cellular, transcriptional, and hormonal responses to stress.

RETRACTED: In vitro regulation of corticotropin-releasing hormone

Studies involving regulation of corticotropin-releasing hormone (CRH) in vitro have been used to validate findings obtained in vivo and more importantly have been used as model systems to better understand signalling mechanisms responsible for the expression of the CRH gene and peptide. Many in vitro studies examining CRH have utilized hypothalamic tissue while a few have focused on the amygdala. Clonal cell lines have also been utilized as models of central nervous system CRH neurons. Stimuli that have been implicated in regulating hypothalamic CRH regulation in vitro include protein kinase A (PKA) and protein kinase C (PKC) activators, glucocorticoids, biogenic amines, cytokines and the gaseous neurotransmitters. Amygdalar CRH levels in vitro are affected by some of the same stimuli that regulate hypothalamic CRH; however there is evidence supporting differential regulation of CRH in these two brain regions by some of the same stimuli. Only a few studies in aggregate have investigated signal transduction mechanisms and these studies have focused on PKA- and glucocorticoid-mediated changes in CRH expression. Thus, much more investigative work in better understanding CRH regulation in vitro is needed.

Interleukin-1 beta depolarizes paraventricular nucleus parvocellular neurones.

Interleukin-1 beta (IL-1 beta) is involved in hypothalamic regulation of corticotropin releasing hormone (CRH) secretion and consequent downstream modulation of the neuroimmune response. In this study, whole-cell patch clamp recordings of rat parvocellular neurones in a slice preparation of the paraventricular nucleus (PVN) of the hypothalamus were performed to examine the cellular effects of IL-1 beta. In response to 1 nm IL-1 beta, 65% of parvocellular neurones tested exhibited a clear depolarization, which was abolished in the presence of tetrodotoxin (TTX). This depolarization was partially dependent on nitric oxide formation, as demonstrated by attenuation of the response in the presence of N-omega-nitro-L-arginine methylester, a nitric oxide synthase inhibitor. The effects of IL-1 beta on responsive parvocellular neurones were associated with a decrease in the frequency of inhibitory post synaptic potentials (IPSPs). Bicuculline administration blocked the effects of IL-1 beta, suggesting that this cytokine modulates GABA-ergic output, resulting in a decrease in inhibitory input (IPSPs) and consequent depolarization. These data support the conclusion that IL-1 beta influences the excitability of parvocellular neurones in the PVN, as a secondary consequence of nitric oxide generation and modulation of GABAergic inhibitory input to these cells. They elucidate cellular correlates underlying the well-established neuroimmune roles of IL-1 beta in the paraventricular nucleus of the hypothalamus.

Expression of Corticotropin-Releasing Hormone Type 1 Receptor in Paraventricular Nucleus after Acute Stress

We have previously proposed the existence of ultrashort loop-positive feedback regulation of corticotropin-releasing hormone (CRH) in the hypothalamus. To gain a better understanding of this effect, we performed double-label in situ hybridization to identify the neurons in the paraventricular nucleus (PVN) that express CRH type 1 receptor (CRH-R1) following stress. We also conducted immunohistochemistry to determine whether CRH-R1 mRNA was translated to CRH-R1 protein in the PVN. Thirty-minute restraint stress given to male Wistar rats increased c-fos mRNA expression primarily in the CRH-producing neurons of the parvocellular PVN. Small numbers of vasopressin and oxytoxin-producing cells were also labeled by c-fos probes. Approximately 70% of CRH-R1 positive neurons exhibited CRH mRNA 2 h after the beginning of stress, while only a small percentage of the vasopressin and oxytocin-producing cells coexpressed CRH-R1 mRNA. CRH-R1 immunoreactivity, which was detected in the perikarya and fibers of PVN neurons, appeared to increase in response to stress, though this was not statistically significant. Pretreatment with a selective CRH-R1 antagonist, CP-154,526, significantly attenuated stress-induced corticotropin (ACTH) secretion as well as c-fos mRNA expression in the PVN. These results demonstrate that acute stress increases neuronal activation and CRH-R1 mRNA expression primarily in CRH-producing neurons of the parvocellular PVN, that CRH-R1 message is translated to CRH-R1 protein, and that PVN neurons are activated at least in part through CRH-R1 under acute stress. The data further support the possibility of feedback regulation of CRH itself in CRH-producing neurons.

Regulation of corticotropin-releasing hormone in vitro

Studies examining regulation of corticotropin-releasing hormone (CRH) in vitro have been used to validate findings obtained in vivo and more importantly have been used as model systems to better understand signalling mechanisms responsible for the expression of the CRH gene and peptide. Most in vitro studies examining CRH have utilized hypothalamic tissue while a few have focused on the amygdala. Furthermore, clonal cell lines have also been utilized as models of central nervous system CRH neurons. Stimuli that have been implicated in regulating hypothalamic CRH in vitro include protein kinase A (PKA) and protein kinase C (PKC) activators, glucocorticoids, biogenic amines, cytokines and the gaseous neurotransmitters. CRH levels in the amygdala in vitro are affected by some of the same stimuli that regulate hypothalamic CRH; however there is evidence supporting differential regulation of CRH in these two brain regions by some of the same stimuli. Only a few studies in aggregate have investigated the signal transduction mechanisms responsible for CRH expression. These mechanistic studies have focused on PKA- and glucocorticoid-mediated changes in CRH expression. Clearly much more investigative work in better understanding CRH regulation in vitro is needed.

Decreased CRH mRNA expression in the fetal guinea pig hypothalamus following maternal nutrient restriction

The regulation of corticotropin-releasing hormone (CRH) mRNA expression following maternal nutrient restriction was examined in the fetal hypothalamus. Pregnant guinea pigs were food restricted for 48 h or fed normally during late gestation. After nutrient restriction, CRH mRNA levels in the hypothalamic paraventricular nucleus of the fetus were determined using in situ hybridization and were found to be significantly decreased ( P<0.0001) compared to controls. In conclusion, we have successfully sequenced the coding sequence of the guinea pig CRH gene, and have shown that a short period (48 h) of maternal nutrient restriction inhibits CRH mRNA expression in the fetal hypothalamus.

Neuronal activity and stress differentially regulate hippocampal and hypothalamic corticotropin-releasing hormone expression in the immature rat

Corticotropin-releasing hormone, a major neuromodulator of the neuroendocrine stress response, is expressed in the immature hippocampus, where it enhances glutamate receptor-mediated excitation of principal cells. Since the peptide influences hippocampal synaptic efficacy, its secretion from peptidergic interneuronal terminals may augment hippocampal-mediated functions such as learning and memory. However, whereas information regarding the regulation of corticotropin-releasing hormone’s abundance in CNS regions involved with the neuroendocrine responses to stress has been forthcoming, the mechanisms regulating the peptide’s levels in the hippocampus have not yet been determined. Here we tested the hypothesis that, in the immature rat hippocampus, neuronal stimulation, rather than neuroendocrine challenge, influences the peptide’s expression. Messenger RNA levels of corticotropin-releasing hormone in hippocampal CA1, CA3 and the dentate gyrus, as well as in the hypothalamic paraventricular nucleus, were determined after cold, a physiological challenge that activates the hypothalamic pituitary adrenal system in immature rats, and after activation of hippocampal neurons by hyperthermia. These studies demonstrated that, while cold challenge enhanced corticotropin-releasing hormone messenger RNA levels in the hypothalamus, hippocampal expression of this neuropeptide was unchanged. Secondly, hyperthermia stimulated expression of hippocampal immediate-early genes, as well as of corticotropin-releasing hormone. Finally, the mechanism of hippocampal corticotropin-releasing hormone induction required neuronal stimulation and was abolished by barbiturate administration. Taken together, these results indicate that neuronal stimulation may regulate hippocampal corticotropin-releasing hormone expression in the immature rat, whereas the peptide’s expression in the hypothalamus is influenced by neuroendocrine challenges.

Differential Regulation of Corticotropin-Releasing Hormone and Vasopressin Transcription by Glucocorticoids

Differential Regulation of Corticotropin-Releasing Hormone and Vasopressin Transcription by Glucocorticoids

Immunoendocrinology of preterm labor: The link between corticotropin-releasing hormone and inflammation

Preterm labor is the final common pathway after several potential insults to the uterus or fetus. The preterm labor syndrome may be precipitated by several different pathophysiologic events, including intrauterine infection, uterine ischemia, uterine overdistention, hormonal disturbances, and other problems. Intrauterine infections (both clinically evident and subclinical) are associated with increased amniotic fluid concentrations of proinflammatory cytokines, and gestational tissues and the fetus are potential sources of these cytokines. In addition to culture-proven intrauterine infection, there may be an “intrauterine inflammatory response syndrome” that could account for cases of preterm labor in which no infectious organism can be identified. Because the immunologic and endocrinologic systems regulate each other extensively, there is potential for corticotropin-releasing hormone to regulate inflammatory responses and vice versa. The cytokine interleukin 1 stimulates production of corticotropin-releasing hormone, and corticotropin-releasing hormone in turn regulates cytokine production by immune effector cells. Because maternal stress is associated with preterm birth, abnormalities in the regulation of corticotropin-releasing hormone and the production of inflammatory cytokines may be a mechanism that could form the pathophysiologic basis for this association. (Am J Obstet Gynecol 1999;180:S251-6.)

Isolation and Characterization of the Rat Corticotropin-Releasing Hormone (CRH)-Binding Protein Gene: Transcriptional Regulation by Cyclic Adenosine Monophosphate and CRH1

The CRH-binding protein (CRH-BP) antagonizes the ACTH-releasing activity of the neuropeptide CRH in vitro. However, the function of CRH-BP in vivo and the molecular mechanisms that regulate CRH-BP expression are not well understood. In this study, the rat CRH-BP gene was characterized, and CRH-BP promoter sequences were identified. The rat CRH-BP gene spans almost 12 kilobases and contains 7 exons. Ribonuclease protection experiments indicate that transcription of the CRH-BP gene initiates at multiple sites in rat cerebral cortex. Transfection experiments with CRH-BP-reporter constructs, containing 88-3500 bp 5' flanking and 66 bp 5' untranslated DNA from the rat CRH-BP gene, demonstrate basal promoter activity in multiple cell lines. CRH-BP-reporter constructs also demonstrate positive regulation of promoter activity by cAMP in a variety of cell lines and by CRH in cells expressing the CRH receptor. The DNA sequences between -341 and -88 bp, including the cAMP response element-like sequence at -127 bp, are required for maximal cAMP and CRH regulation of CRH-BP promoter activity. These studies suggest that CRH-BP transcription in vivo may be positively regulated by cAMP and CRH.

Brainstem hemisection decreases corticotropin-releasing hormone mRNA in the paraventricular nucleus but not in the central amygdaloid nucleus.

Corticotropin-releasing hormone (CRH) neurons in the paraventricular nucleus (PVN) of the hypothalamus and in the central nucleus of the amygdala (ACE) participate in neurohumoral and behavioral responses to stress. To understand better the central regulation of CRH, the present study assessed the effects of ipsilateral surgical hemisection of the brainstem on expression of CRH mRNA in the PVN and the ACE. In situ hybridization was used to demonstrate PVN CRH mRNA expression in hemisected, sham-operated or intact rats before and after 3 h of immobilization (IMMO). In addition, hypothalamic-pituitary-adrenocortical (HPA) axis activity at baseline and during IMMO was assessed by measurements of plasma concentrations of ACTH and corticosterone. IMMO markedly increased CRH mRNA expression in the PVN in all experimental groups. Rats with brainstem hemisections had lower PVN CRH mRNA expression ipsilateral to the lesion and markedly blunted responses after IMMO, compared to values in sham-operated rats. In contrast, neither hemisection nor IMMO affected CRH mRNA expression in the ACE. Lesioned and SHAM-operated groups did not differ in baseline or IMMO-induced increases in plasma ACTH or corticosterone levels. The present results indicate that baseline levels and IMMO-induced increments in CRH mRNA expression in the PVN, but not in the ACE, depend on ipsilaterally ascending medullary tracts and that IMMO-induced HPA activation does not depend on these pathways.