Catherine Rivier

Chemical and biological characterization of the inhibin family of protein hormones.

Publisher Summary This chapter discusses the chemical and biological characterization of the inhibin family of protein hormones, which is a family of peptides isolated from the follicular fluid or rete testis fluid on the basis of their ability to inhibit the secretion of the follicle-stimulating hormone (FSH) by cultured rat anterior pituitary cells. It also reviews the possible roles of inhibin and fibre-reinforced plastic (FRP)/activin in placenta, brain, and bone marrow. Inhibin-related dimers are broadly distributed anatomically and have powerful activities in several biological systems where inhibin and FRP/activin often exhibit opposite effects. While the physiologic roles of inhibin to regulate FSH secretion in the female rat and immature male rat are strongly supported, the significance of these hormones within the gonad, brain, placenta, and bone marrow have yet to be placed in in vivo context. Although the panoply of functions of inhibin and FRP/activin are certainly incompletely understood at this time, this family has already demonstrated a powerful mechanism for the generation of signal diversity whereby differential subunit association can result in the generation of dimers with opposing biological actions in multiple tissues.

Chemical and Biological Characterization of Corticotropin Releasing Factor

Publisher Summary Hypothalamus liberates a substance into the hypophysial portal blood that stimulates the adrenocorticotrophic hormone (ACTH) activity of the pituitary. This chapter discusses the chemical and biological characterization of this corticotropin releasing factor (CRF). Several known naturally occurring substances including vasopressin, oxytocin, norepinephrine, epinephrine, and angiotensin II are found to stimulate ACTH secretion. Partially purified preparations of CRF stimulates the secretion of a number of peptides derived from the proopiomelanocortin (POMC) precursor—including the opioid peptide, β-endorphin. The chapter explains that CRF is likely to be distributed outside of the hypothalamus and possess extra hypophysiotropic actions. In vitro systems are vulnerable to non specific secretagogs in extracts including myelin basic protein, histones, potassium ion, and the components of various buffers and solvents. Ovine CRF is homologous with several known peptides including sauvagine and urotensin I. CRF also shows some homology with calmodulin and with angiotensinogen. The tetrapeptide Phe-His-Leu-Leu is common to both angiotensinogen and CRF and is the site in angiotensinogen of renin and converting enzyme cleavage. The chapter concludes with the evidence that supports CRF or a closely related peptide in the neuroregulation of the pituitary corticotropic cells.

Corticotropin-releasing factor: Effects on the sympathetic nervous system and oxygen consumption

Corticotropin-releasing factor administered intracerebroventricularly produces prolonged elevation of plasma concentration of epinephrine, norepinephrine and glucose. These hormonal changes are associated with an increase in motor activity and oxygen consumption. No change in body temperature is observed. CRF produces changes in animal physiology that are similar to those observed in response to stress.

Corticotropin-Releasing Factor (CRF) and Endocrine Responses to Stress: CRF Receptors, Binding Protein, and Related Peptides

Abstract Corticotropin-releasing factor (CRF) is a 41 -amino acid neuropeptide, which is recognized as a critical mediator of complimentary, stress-related endocrine, autonomic, and behavioral responses in mammalian species. CRF belongs to a family of structurally related peptides including frogskin sauvagine and fish urotensin I. The effects of CRF and related peptides are mediated by two distinct receptors, which differ in their anatomical distribution, as well as in their pharmacological characteristics. In addition, CRF is bound with high affinity by a CRF binding protein (CRF-BP), which is a putative inhibitor of CRF action. CRF is probably not the sole endogenous ligand for CRF receptors or the CRF-BP, since a second mammalian member of the CRF family, urocortin, has recently been identified. This article describes recent findings with respect to CRF, its receptors, binding protein, and CRF-related peptides, which provide further insights into the role and mechanisms of CRF action in stress responses.

[38] Assay of corticotropin releasing factor

Publisher Summary The corticotropin releasing factor (CRF) can be assayed by a variety of biological and immunological techniques. The chapter describes two methods: an in vitro assay employing primary anterior pituitary cell cultures and a radioimmunoassay (RIA) procedure. The chapter presents the acid extraction method for tissue fragments and a Bond Elute method for aqueous samples for extraction of CRF. The cell culture assay is the exclusive assay used for the isolation and subsequent characterization of ovine CRF and has, thereby, been useful for the detection and quantification of CRF in crude or partially purified extracts. The method has been applied for a wide range of studies investigating the mode of action of CRF and its interactions with other physiological modulators such as glucocorticoids, catecholamines, and neurohypophysial peptides. This method is useful for the assay of both agonist and antagonist analogs of CRF and of highly purified CRF-like peptides of various mammalian and nonmammalian species.

Cocaine stimulates adrenocorticotropin (ACTH) secretion through a corticotropin-releasing factor (CRF)-mediated mechanism

Cocaine was injected intravenously to non-anesthetized, freely moving adult male rats and caused dose-dependent elevations in plasma adrenocorticotropin (ACTH) levels. The observation that this stimulatory effect was completely abolished by pretreatment with a corticotropin-releasing factor (CRF) antiserum, coupled with the lack of effect of cocaine on ACTH secretion by cultured pituitary cells, suggests that cocaine acts within the brain to release endogenous CRF.

Alcohol self‐administration acutely stimulates the hypothalamic‐pituitary‐adrenal axis, but alcohol dependence leads to a dampened neuroendocrine state

Clinical studies link disruption of the neuroendocrine stress system with alcoholism, but remaining unknown is whether functional differences in the hypothalamic‐pituitary‐adrenal (HPA) axis precede alcohol abuse and dependence or result from chronic exposure to this drug. Using an operant self‐administration animal model of alcohol dependence and serial blood sampling, we show that long‐term exposure to alcohol causes significant impairment of HPA function in adult male Wistar rats. Acute alcohol (voluntary self‐administration or experimenter‐administered) stimulated the release of corticosterone and its upstream regulator, adrenocorticotropic hormone, but chronic exposure sufficient to produce dependence led to a dampened neuroendocrine state. HPA responses to alcohol were most robust in ‘low‐responding’ non‐dependent animals (averaging < 0.2 mg/kg/session), intermediate in non‐dependent animals (averaging ∼0.4 mg/kg/session), and most blunted in dependent animals (averaging ∼1.0 mg/kg/session) following several weeks of daily 30‐min self‐administration sessions, suggesting that neuroendocrine tolerance can be initiated prior to dependence and relates to the amount of alcohol consumed. Decreased expression of corticotropin‐releasing factor (CRF) mRNA expression in the paraventricular nucleus of the hypothalamus and reduced sensitivity of the pituitary to CRF may contribute to, but do not completely explain, neuroendocrine tolerance. The present results, combined with previous studies, suggest that multiple adaptations to stress regulatory systems may be brought about by excessive drinking, including a compromised hormonal response and a sensitized brain stress response that together contribute to dependence.

Corticosteroid-Dependent Plasticity Mediates Compulsive Alcohol Drinking in Rats

Alcoholism is characterized by a compulsion to seek and ingest alcohol, loss of control over intake, and the emergence of a negative emotional state during abstinence. We hypothesized that sustained activation of neuroendocrine stress systems (e.g., corticosteroid release via the hypothalamic-pituitary-adrenal axis) by alcohol intoxication and withdrawal and consequent alterations in glucocorticoid receptor (GR) and mineralocorticoid receptor (MR) activation drive compulsive alcohol drinking. Our results showed that rats exposed to alcohol vapor to the point of dependence displayed increased alcohol intake, compulsive drinking measured by progressive-ratio responding, and persistent alcohol consumption despite punishment, assessed by adding quinine to the alcohol solution, compared with control rats that were not exposed to alcohol vapor. No group differences were observed in the self-administration of saccharin-sweetened water. Acute alcohol withdrawal was accompanied by downregulated GR mRNA in various stress/reward-related brain regions [i.e., prefrontal cortex, nucleus accumbens (NAc), and bed nucleus of the stria terminalis (BNST)], whereas protracted alcohol abstinence was accompanied by upregulated GR mRNA in the NAc core, ventral BNST, and central nucleus of the amygdala. No significant alterations in MR mRNA levels were found. Chronic GR antagonism with mifepristone (RU38486) prevented the escalation of alcohol intake and compulsive responding induced by chronic, intermittent alcohol vapor exposure. Chronic treatment with mifepristone also blocked escalated alcohol drinking and compulsive responding during protracted abstinence. Thus, the GR system appears to be involved in the development of alcohol dependence and may represent a potential pharmacological target for the treatment of alcoholism.

Detection and purification of inhibin using antisera generated against synthetic peptide fragments

Publisher Summary Inhibin is a hormone whose best established physiological role is selective suppression of the release of follicle-stimulating hormone (FSH) from the pituitary. Subsequent to the identification of inhibin, two laboratories isolated a protein from porcine follicular fluid that selectively released FSH from pituitary cell cultures. Characterization of this protein––named FSH releasing protein (FRP)/activin––showed that it was a homo- or heterodimer of inhibin β subunits. This chapter also describes the production of antibodies to the α, βA, and βB subunits of inhibin; development of a radioimmunoassay (RIA) specific for inhibin, using an antibody directed to the α subunit; use of these antibodies for Western blot analysis; and a method for concentrating inhibin and FRP from biological fluids. It also discusses a method that was developed for the rapid isolation of inhibin from ram rete testis fluid (RTF) using immunoaffinity chromatography with an antibody directed against the α subunit.

Stress and Interleukin‐1 β‐Induced Activation of c‐fos, NGFI‐B ann CRF Gene Expression in the Hypothalamic PVN: Comparison Between Sprague‐Dawley, Fisher‐344 and Lewis Rats

Various signals are known to activate the hypothalamic‐pituitary‐adrenal (HPA) axis, an event largely dependent on the release of corticotropin‐releasing factor (CRF) which originates mainly from the parvocellular paraventricular nucleus (PVN) of the hypothalamus. These signals include neurogenic stimuli such as exposure to mild electroshocks, and systemic stimuli like administration of cytokines. The HPA axis activity of Lewis rats has been reported to be hyporesponsive to such stimuli, but the exact mechanisms involved in this phenomenon are poorly understood. The present study investigated the effect of footshock exposure and central injection of interleukin (IL)‐1β, on CRF neuronal activity and gene expression in the PVN of adult male Sprague‐Dawley (SD), Fisher‐344 (F344) and Lewis (LEW) rats. The animals were deeply anesthetized and rapidly perfused transcardially with a solution of 4% paraformaldehyde 3 h after the beginning of the footshock session (1.5 mA, 2 s duration, 4/min over 1 h), or the i.c.v. injection of IL‐1β (100 ng in 10 μl). mRNA encoding the immediate ‘early’ genes (lEGs) c‐fos and NGFI‐B, as well as CRF, were assayed by in situ hybridization histochemistry, while the localization of Fos protein within CRF‐immunoreactive (ir) neurons in the PVN was determined using a dual immunostaining protocol. Both stress and IL‐1β induced robust Fos‐ir expression within the parvocellular division of the PVN in all 3 strains. The number of cells immunoreactive for both Fos and CRF proteins in the PVN was similar in SD, F344 and LEW rats following either challenge. While control animals did not display detectable levels of c‐fos or NGFI‐B mRNA in the PVN, both treatments induced significant expression of these transcripts in this hypothalamic nucleus and no significant differences were observed among SD, F344 and LEW rats. Relative levels of CRF mRNA in the PVN were also significantly and comparably increased following either stress or central IL‐1β treatment. In contrast, plasma ACTH and corticosterone levels were significantly higher in F344 and SD rats than in LEW animals during the stress session.