Corticotropin-releasing hormone, its binding protein and receptors in human cervical tissue at preterm and term labor in comparison to non-pregnant state

Background  

Preterm birth is still the leading cause of neonatal morbidity and mortality. The level of corticotropin-releasing hormone (CRH) is known to be significantly elevated in the maternal plasma at preterm birth. Although, CRH, CRH-binding protein (CRH-BP), CRH-receptor 1 (CRH-R1) and CRH-R2 have been identified both at mRNA and protein level in human placenta, deciduas, fetal membranes, endometrium and myometrium, no corresponding information is yet available on cervix. Thus, the aim of this study was to compare the levels of the mRNA species coding for CRH, CRH-BP, CRH-R1 and CRH-R2 in human cervical tissue and myometrium at preterm and term labor and not in labor as well as in the non-pregnant state, and to localize the corresponding proteins employing immunohistochemical analysis.     

Corticotropin-releasing hormone activates protein kinase C in an isoenzyme-specific manner

Protein kinase C (PKC) has recently emerged as mediator of corticotropin-releasing hormone (CRH) effects. Aim of the present study was to study the effects of CRH on each PKC isoenzyme. As a model we have used the PC12 rat pheochromocytoma cell line, expressing the CRH type 1 receptor (CRHR1). Our data were as follows: (a) CRH-induced rapid phosphorylation of conventional PKCalpha and PKCbeta, accompanied by parallel increase of their concentration within nucleus. (b) CRH suppressed the phosphorylation of novel PKCdelta and PKCtheta;, which remained in the cytosol. (c) CRH-induced transient phosphorylation of atypical PKClambda and had no effect on PKCmu. (d) The effect of CRH on each PKC isoenzyme was blocked by a CRHR1 antagonist. (e) Blockade of conventional PKC phosphorylation inhibited CRH-induced calcium ion mobilization from intracellular stores as well as the CRH-induced apoptosis and Fas ligand production. In conclusion, our findings suggest that CRH via its CRHR1 receptor differentially regulates PKC-isoenzyme phosphorylation, an apparently physiologically relevant effect since blockade of conventional PKC phosphorylation abolished the biological effect of CRH.     

Corticotropin-releasing hormone in the regulation of adaptive behavior

Recent findings on the role of corticotropin-releasing hormone (CRH) in the regulation of stress and its consequences are summarized and analyzed in the review. Being involved in stress-activating system this neurohormone is referred to as a neurochemical factor triggering and integrating both endocrine and behavioral functions. The CRH distribution in hypothalamus and extrahypothalamic brain regions relevant to its involvement in the controlling of endocrine processes and behavior is viewed in details. Distinct behavioral outcomes of stress and the contribution of amygdalar, hippocampal, and striatal CRH-structures, implicated in general organism response to external influences, are widely discussed. From this viewpoint the mechanisms involved in the development of post-stress psychopathology, as well as drug addiction and alcoholism are treated.     

Corticotropin-releasing hormone in depression and post-traumatic stress disorder

Corticotropin-releasing hormone (CRH) has been implicated in the regulation of a wide range of behaviors including arousal, motor function, feeding, and reproduction. Because depressed patients are often hypercortisolemic and intracerebroventricular administration of CRH to experimental animals produces a syndrome reminiscent of depression, dysregulation of this compound has been suggested to be involved in the pathogenesis of depressive and anxiety disorders. Studies of cerebrospinal fluid CRH levels and clinical neuroendocrine tests in patients with anxiety and affective disorders have supported this hypothesis. This review discusses these neuroendocrine findings in melancholic and atypical depression as well as post-traumatic stress disorder (PTSD). Overall, the data suggest that melancholic depression is characterized by hyperactive central CRH systems with overactivity of the pituitary-adrenal (HPA) axis. On the other hand, atypical depression is characterized by hypoactive central CRH systems and accompanying underactivity of the hypothalamic-pituitary-adrenal axis. Furthermore, the neuroendocrinology of PTSD appears to be unique, in that patients have hyperactive central CRH systems with underactivity of the pituitary-adrenal axis.     

Corticotropin-releasing hormone and pituitary-adrenocortical responses in chronically stressed rats

Brain corticotropin-releasing hormone (CRH) concentration and pituitary adreno-cortical responses were examined in chronically stressed rats: body restraint stress (6 h/day) for 4 or 5 weeks. Stressed rats showed a reduction in weight gain. CRH concentration in the median eminence and the rest of the hypothalamus were not different between control and chronically immobilized rats. The anterior pituitary adenocorticotropic hormone (ACTH) concentration was elevated in chronically stressed rats, whereas plasma ACTH and corticosterone levels did not differ from the control values. The median eminence CRH concentration was reduced to the same extent at 5 min after onset of ether exposure (1 min) in chronically immobilized rats and controls. However, plasma ACTH and corticosterone showed greater responses to ether stress in chronically immobilized rats than in control rats. Plasma ACTH and corticosterone responses to exogenous CRH were not different between control and chronically immobilized rats, while the response to arginine vasopressin (AVP) was significantly greater in chronically immobilized rats. These results suggest that chronic stress caused an increase in the ACTH-secreting mechanism and that pituitary hypersensitivity to vasopressin might at least be partly responsible for this.     

Corticotropin-releasing factor receptors and actions in rat Leydig cells

Rat Leydig cells possess functional high affinity receptors for corticotropin-releasing factor (CRF). CRF inhibited human chorionic gonadotropin (hCG)-induced androgen production in cultured fetal and adult Leydig cells in a dose-dependent manner, but it had no effect on basal testosterone secretion. Comparable inhibitory effects of CRF were observed in the presence or absence of 3-isobutyl-1-methylxanthine. CRF treatment caused a marked reduction of steroid precursors of the androgen pathway (from pregnenolone to testosterone) during gonadotropin stimulation, but it did not influence their basal levels. The inhibitory action of CRF on hCG-induced steroidogenesis was fully reversed by 8-bromo-cAMP but was not affected by pertussis toxin. The action of CRF was rapid; and it was blocked by coincubation with anti-CRF antibody. CRF caused no changes in hCG binding to Leydig cells, and in contrast to other target tissues, CRF did not stimulate cAMP production, indicating that CRF receptors are not coupled to Gs in Leydig cells. These studies have demonstrated that CRF-induced inhibition of the acute steroidogenic action of hCG is exerted at sites related to receptor/cyclase coupling or cAMP formation. The inhibitory effects of CRF in the Leydig cell do not occur through the Gi unit of adenylate cyclase, but could involve pertussis toxin-insensitive G protein(s). These observations demonstrate that CRF has a novel and potent antireproductive effect at the testicular level. Since CRF is synthesized in the testis and is present in Leydig cells, it is likely that locally produced CRF could exert negative autocrine modulation on the stimulatory action of luteinizing hormone on Leydig cell function.     

Thyroid hormone action at the cellular level

Thyroid hormones influence numerous physiological and biochemical functions. The expression of the hormonal effects involves several events. The interaction of T3 with nuclear receptors, and the stimulation of mRNA production appears to be a major step. Extranuclear binding of thyroid hormones could account for early responses. Plasma membrane receptors may play a role in the cellular uptake of T3 and the stimulation of amino acids and sugar transport. A direct control of oxidative phosphorylation through binding of T3 to mitochondrial binding sites has been proposed. The role of cytosolic binding proteins remains unclear. The understanding of the mode of action of thyroid hormones requires a better knowledge of the molecular events occurring at the nuclear level, and the relation between the nuclear and extranuclear binding sites in the hormonal expression.     

Corticotropin-releasing hormone induces keratinocyte differentiation in the adult human epidermis

Previously we documented that human epidermis exclusively expresses corticotropin releasing hormone receptor 1 (CRH-R1). To define the role of CRH in the epidermis, we investigated its effects on differentiation of normal human adult epidermal keratinocytes. Thus, CRH inhibited proliferation in a dose dependent fashion and significantly decreased Ki-67 antigen expression. This effect was independent of either the presence or the absence of growth factors in the medium. Flow cytometry analysis demonstrated that CRH inhibited the transition from G0/1 to S phase of the cell cycle, which was accompanied by an increased expression of cdk inhibitor p16 (Ink4a) protein. The antiproliferative effect was attenuated by protein kinase C inhibitor (GF109203X) but not by H89 (protein kinase A inhibitor), PD98059, or SB203580 (MAP kinase inhibitors). The cell cycle withdrawal was associated with the induction of keratinocyte differentiation. Thus, CRH stimulated the expression of cytokeratin 1 and involucrin, and inhibited cytokeratin 14 on both mRNA and protein levels. It also increased cell granularity and cell size. Furthermore, CRH induced signal transduction cascade that included stimulation of inositol 1,4,5-triphosphate, which was time and dose dependent. CRH also increased activator protein-1 DNA binding activity with JunD identified as the most important element. Thus, activation of CRH-R1 induces a non-random and sequential signal transduction cascade governing both keratinocyte differentiation and the inhibition of cell proliferation through G0/1 arrest. We propose that this program, triggered by CRH interaction with CRH-R1, includes induction of a transduction pathway involving the sequential activation of phospholipase C, protein kinase C, activator protein-1 (including Jun D), and p16.     

Corticotropin-releasing hormone in the human hypothalamus. Free-floating immunostaining method

We have clearly demonstrated corticotropin-releasing hormone (CRH) immunoreactive cell bodies and nerve fibers in the human hypothalamus by immunocytochemistry using free-floating sections instead of paraffin-embedded sections. Human hypothalami were obtained at autopsy, fixed and cryostat-sectioned at 40 microns. Free-floating sections were immunostained with antibody to CRH using the Vector ABC system. Most of CRH immunoreactive nerve fibers from the paraventricular nucleus pass under the fornix, while some CRH immunoreactive nerve fibers pass beyond the fornix and some through the fornix. Then the CRH immunoreactive nerve fibers run downward, medially to the supraoptic nucleus and toward the pituitary stalk. This method of immunocytochemistry is a very sensitive and suitable means for immunocytochemical studies of neuropeptides in the human brain.     

Corticotropin-releasing hormone receptor (CRHR)1 and CRHR2 are both trafficking and signaling receptors for urocortin

Transport of urocortin, a potent satiety peptide, occurs at the blood-brain barrier of the mouse. Endocytosis of urocortin by the cerebral microvessel endothelial cells composing the blood-brain barrier is a rate-limiting step of this transport, but the cellular mechanisms involved have not been fully elucidated. The presence of both CRH receptors R1 and R2 in isolated cerebral microvessels shown in this study suggested that both subtypes might mediate urocortin transport. The roles of these two receptors in the endocytosis and signal transduction of urocortin were tested by overexpression studies in human embryonic kidney 293 cells. Both receptors led to a significant increase of binding and endocytosis of radiolabeled urocortin. CRHR1-mediated urocortin endocytosis was blocked by astressin (antagonist for both CRHRs), whereas CRHR2-mediated urocortin endocytosis was also blocked by antisauvagine 30 (selective CRHR2beta antagonist). Chlorpromazine, filipin, and nystatin had no effect on urocortin endocytosis, indicating the lack of significant involvement of clathrin or caveolae membrane microdomains. Both CRHR1 and CRHR2 were able to mediate the ligand-induced increase of cAMP production, suggesting that the overexpressed receptors were biologically active. Elevation of intracellular cAMP by forskolin or dibutyryl-cAMP, however, did not show acute modulation of the binding and endocytosis of urocortin. Despite the substantial intracellular degradation of endocytosed urocortin in cells overexpressing either CRHR1 or CRHR2, intact urocortin could be exocytosed during the 1-h study interval. We conclude that both CRHR1 and CRHR2 play a facilitatory role in the non-clathrin-, non-caveolae-mediated endocytosis and intracellular signal transduction of this potent peptide.     

Amiodarone antagonizes the effects of T3 at the receptor level: an additional mechanism for its in vivo hypothyroid-like effects

Amiodarone is a diiodinated benzofuran derivative that has some structural similarities to the thyroid hormones and contains two iodine atoms per molecule. It has exhibited hypothyroid-like effects that are thought to be the result of an inhibition of thyroid hormone synthesis due to iodine load, a decrease in the T4 to T3 conversion, and (or) a competitive binding for T3 receptors. The aim of this study was to determine if this third mechanism contributes to the hypothyroid-like effects of amiodarone in vivo. To do so, some characteristic features known to be influenced by hypothyroidism were determined in surgically thyroidectomized rats (n = 48), which received replacement doses of T3 (0.5 and 1.0 microgram.100 g-1.day-1) with or without amiodarone (60 mg.kg-1.day-1). Thyroidectomy produced a hypothyroid state upon which amiodarone had no detectable effects except a negative body weight gain. T3 (0.5 microgram) nearly normalized the thyroid status of the animals, but the concomitant administration of amiodarone induced hypothyroid-like effects suggesting that these effects are dependent on T3. Higher doses of T3 (1.0 microgram) produced hyperthyroid-like effects and attenuated the effects of amiodarone. Unexpectedly, amiodarone decreased T3 plasma concentrations. To determine if the effects of amiodarone were the results of a decrease in T3 plasma and myocardial concentrations or a competition with T3 for its receptors, exogenous T3 pharmacokinetics were studied in thyroidectomized rats receiving T3 (0.5 microgram) with or without amiodarone. The results suggested that amiodarone increased T3 cardiac concentrations.(ABSTRACT TRUNCATED AT 250 WORDS)     

Melanin-concentrating hormone and its receptors: state of the art

Melanin-concentrating hormone (MCH) is a cyclic neuropeptide of nineteen amino acids in mammals. Its involvement in the feeding behaviour has been well established during the last few years. A first receptor subtype, now termed MCHIR, was discovered in 1999, following the desorphanisation of the SLCI orphan receptor, using either reverse pharmacology or systematic screening of agonist candidates. A second MCH receptor, MCH2R, has been discovered recently, by several groups working on data mining of genomic banks. The molecular pharmacology of these two receptors is only described on the basis of the action of peptides derived from MCH. The present review tentatively summarizes the knowledge on these two receptors and presents the first attempts to discover new classes of antagonists that might have major roles in the control of obesity and feeding behaviour.