Biomedical subjects

W Vale

Publications and source records attributed to W Vale.

At least 19 recordsLinked to original sources

PubMed2005

CRF2 receptor activation prevents colorectal distension induced visceral pain and spinal ERK1/2 phosphorylation in rats.

BACKGROUND AND AIMS: Activation of corticotropin releasing factor 1 (CRF1) receptors is involved in stress related responses and visceral pain, while activation of CRF2 receptors dampens the endocrine and some behavioural stress responses. We hypothesised that CRF2 receptor activation may influence visceral pain induced by colorectal distension (CRD) in conscious rats, and assessed the possible sites and mechanisms of action. METHODS: Male Sprague-Dawley rats were exposed to CRDs (60 mm Hg, 10 minutes twice, with a 10 minute rest interval). Visceromotor responses (VMR) were measured by electromyography or visual observation. Spinal (L6-S1) extracellular signal regulated kinase 1/2 (ERK 1/2) activation following in vivo CRD and CRF2 receptor gene expression in the T13-S1 dorsal root ganglia (DRG) and spinal cord were determined. Inferior splanchnic afferent (ISA) activity to CRD (0.4 ml, 20 seconds) was assessed by electrophysiological recording in an in vitro ISA nerve-inferior mesenteric artery (intra-arterial)-colorectal preparation. RESULTS: In controls, VMR to the second CRD was mean 31 (SEM 4)% higher than that of the first (p<0.05). The selective CRF2 agonist, human urocortin 2 (hUcn 2, at 10 and 20 microg/kg), injected intravenous after the first distension, prevented sensitisation and reduced the second response by 8 (1)% and 30 (5)% (p<0.05) compared with the first response, respectively. RT-PCR detected CRF2 receptor gene expression in the DRG and spinal cord. CRD (60 mm Hg for 10 minutes) induced phosphorylation of ERK 1/2 in neurones of lumbosacral laminae I and IIo and the response was dampened by intravenous hUcn 2. CRD, in vitro, induced robust ISA spike activity that was dose dependently blunted by hUcn 2 (1-3 microg, intra-arterially). The CRF2 receptor antagonist, astressin2-B (200 microg/kg subcutaneously or 20 microg intra-arterially) blocked the hUcn 2 inhibitory effects in vivo and in vitro. CONCLUSIONS: Peripheral injection of hUcn 2 blunts CRD induced visceral pain, colonic afferent, and spinal L6-S1 ERK 1/2 activity through CRF2 receptor activation in rats.

Animals↗

PubMed2004

Cardiovascular effects of long-term central and peripheral administration of urocortin, corticotropin-releasing factor, and adrenocorticotropin in sheep.

The neuroendocrine hormones ACTH and corticotropin- releasing factor (CRF), which are involved in the stress response, have acute effects on arterial pressure. New evidence indicates that urocortin (UCN), the putative agonist for the CRF type 2 receptor, has selective cardiovascular actions. The responses to long-term infusions of these hormones, both peripherally and centrally, in conscious animals have not been studied. Knowledge of the long-term effects is important because they may differ considerably from their acute actions, and stress is frequently a chronic stimulus. The present experiments investigated the cardiovascular effects of CRF, UCN, and ACTH in conscious sheep. Infusions were made either into the lateral cerebral ventricles (i.c.v.) or i.v. over 4 d at 5 microg/h. UCN infused i.c.v. or i.v. caused a prolonged increase in heart rate (HR) (P < 0.01) and a small increase in mean arterial pressure (MAP) (P < 0.05). CRF infused i.c.v. or i.v. progressively increased MAP (P < 0.05) but had no effect on HR. Central administration of ACTH had no effect, whereas systemic infusion increased MAP and HR (P < 0.001). In conclusion, long-term administration of these three peptides associated with the stress response had prolonged, selective cardiovascular actions. The striking finding was the large and sustained increase in HR with i.c.v. and i.v. infusions of UCN. These responses are probably mediated by CRF type 2 receptors because they were not reproduced by infusions of CRF.

Adrenocorticotropic Hormone↗

PubMed2003

Residues in the C-terminal region of activin A determine specificity for follistatin and type II receptor binding.

Activin is a secreted growth factor that signals by binding two related classes of single transmembrane receptors at the cell surface. The interaction of activin with its receptors is highly regulated by other cell surface receptors, antagonistic ligands, and high affinity extracellular binding proteins such as follistatin. Two activin A mutants, the deletion mutant des[85-109]-activin A and the point mutant K102E-activin A (K102E), were investigated with respect to their ability to bind cell surface receptors and the binding protein follistatin. The deletion mutant exhibits low affinity for both receptors and follistatin whereas the point mutant fails to bind cell surface receptors but binds follistatin-288 with high affinity. K102E is able to compete with wild type activin to bind to follistatin and can thus increase the concentration of activin available for receptor binding and signaling. These findings underline the importance of the C-terminal region of activin for binding interactions and show that different residues in this region are involved in cell surface receptor and follistatin interactions.

Activins↗

PubMed2002

Potent and long-acting corticotropin releasing factor (CRF) receptor 2 selective peptide competitive antagonists.

We present evidence that members of the corticotropin releasing factor (CRF) family assume distinct structures when interacting with the CRF(1) and CRF(2) receptors. Predictive methods, physicochemical measurements, and structure-activity relationship studies have suggested that CRF, its family members, and competitive antagonists such as astressin [cyclo(30-33)[DPhe(12),Nle(21),Glu(30),Lys(33),Nle(38)]hCRF((12-41))] assume an alpha-helical conformation when interacting with their receptors. We had shown that alpha-helical CRF((9-41)) and sauvagine showed some selectivity for CRF receptors other than that responsible for ACTH secretion(1) and later for CRF2.(2) More recently, we suggested the possibility of a helix-turn-helix motif around a turn encompassing residues 30-33(3) that would confer high affinity for both CRF(1) and CRF(2)(2,4) in agonists and antagonists of all members of the CRF family.(3) On the other hand, the substitutions that conferred ca. 100-fold CRF(2) selectivity to the antagonist antisauvagine-30 [[DPhe(11),His(12)]sauvagine((11-40))] did not confer such property to the corresponding N-terminally extended agonists. We find here that a Glu(32)-Lys(35) side chain to side chain covalent lactam constraint in hCRF and the corresponding Glu(31)-Lys(34) side chain to side chain covalent lactam constraint in sauvagine yield potent ligands that are selective for CRF(2). Additionally, we introduced deletions and substitutions known to increase duration of action to yield antagonists such as cyclo(31-34)[DPhe(11),His(12),C(alpha)MeLeu(13,39),Nle(17),Glu(31),Lys(34)]Ac-sauvagine((8-40)) (astressin(2)-B) with CRF(2) selectivities greater than 100-fold. CRF receptor autoradiography was performed in rat tissue known to express CRF(2) and CRF(1) in order to confirm that astressin(2)-B could indeed bind to established CRF(2) but not CRF(1) receptor-expressing tissues. Extended duration of action of astressin(2)-B vs that of antisauvagine-30 is demonstrated in the CRF(2)-mediated animal model whereby the inhibition of gastric emptying of a solid meal in mice by urocortin administered intraperitoneally at time zero is antagonized by the administration of astressin(2)-B but not by antisauvagine-30 at times -3 and -6 h while both peptides are effective when given 10 min before urocortin.

Animals↗

PubMed2001

Hypothalamic paraventricular nucleus injections of urocortin alter food intake and respiratory quotient.

Corticotropin releasing hormone (CRH) acts on the central nervous system to alter energy balance and influence both food intake and sympathetically-mediated thermogenesis. CRH is also reported to inhibit food intake in several models of hyperphagia including neuropeptide Y (NPY)-induced eating. The recently identified CRH-related peptide, urocortin (UCN), also binds with high affinity to CRH receptor subtypes and decreases food intake in food-deprived and non-deprived rats. The present experiment characterized further the feeding and metabolic effects of UCN by examining its impact after direct injections into the paraventricular nucleus (PVN) of the hypothalamus. In feeding tests (n=8), UCN (50-200 pmol) was injected into the PVN at the onset of the dark cycle and food intake was measured 1, 2 and 4 h postinjection. In separate rats (n=8), the metabolic effects of UCN were monitored using an open circuit calorimeter which measured oxygen consumption (V(O2)) and carbon dioxide production (V(CO2)). Respiratory quotient (RQ) was calculated as V(CO2)/V(O2). UCN suppressed feeding at all times studied and reliably decreased RQ within 30 min of infusion. Additional work examined the effect of UCN (50-100 pmol) pretreatment on the feeding and metabolic effects of NPY. NPY, injected at the start of the dark period, reliably increased 2 h food intake. This effect was blocked by PVN UCN administration. Similarly, UCN blocked the increase in RQ elicited by NPY alone. These results suggest that UCN-sensitive mechanisms within the PVN may modulate food intake and energy substrate utilization, possibly through an interaction with hypothalamic NPY.

Animals↗

PubMed2001

Antagonism of activin by inhibin and inhibin receptors: a functional role for betaglycan-glycan.

Activin and inhibin research has provided important insight into reproductive physiology as well as many areas involving regulation of cell growth, differentiation and function. Progress in understanding the roles of these hormones in various cell and tissue types has been complimented by novel discoveries at the molecular level that have shed light on ligand/receptor interactions, signaling mechanisms and regulation. While the receptors and signaling pathway for activin are now well characterized, the molecular basis for inhibin action has remained relatively unclear. Here we summarize recent advances in understanding inhibin's mode of action focusing on our recent identification of betaglycan-glycan as an inhibin co-receptor capable of mediating inhibin action.

Activin Receptors↗

PubMed2001

Mice that lack corticotropin-releasing factor (CRF) receptors type 1 show a blunted ACTH response to acute alcohol despite up-regulated constitutive hypothalamic CRF gene expression.

BACKGROUND: The purpose of this work was to determine the influence of acute alcohol treatment, injected intraperitoneally, on the hypothalamic-pituitary-adrenal axis of mice that lack type 1 receptor for corticotropin-releasing factor (CRFR1). METHODS: CRFR1-deficient (CRFR1-/-), heterozygous (CRFR1+/-), and wild-type (CRFR1+/+) mice were generated and maintained under standard conditions. Homozygous, heterozygous, and wild-type offspring were identified by polymerase chain reaction analysis of tail DNA. Experiments were performed on 9- to 16-week-old male and female mice. All blood samples were obtained by rapid decapitation of conscious mice conducted between 10 AM-12 PM. Blood sample collection was completed within 20 to 30 sec of disturbing the animals, and all samples were terminal. Preliminary experiments were conducted to determine the time-course of the ACTH and hypothalamic responses to alcohol in all three groups of mice, and a single time point (30 min and 2 hr, respectively), corresponding to peak responses, was chosen to measure the corresponding parameters in all subsequent studies. RESULTS: In vehicle-injected animals, basal ACTH and corticosterone levels were statistically comparable in heterozygotes and mice with a null allele for the CRFR1 gene, although values of this latter hormone were slightly lower in the mutants. Alcohol (4.0 g/kg) elicited the expected significant (p < 0.01) increase in plasma ACTH and corticosterone levels in heterozygous mice. These responses were virtually abolished or markedly decreased, respectively, in CRFR1-deficient animals. As previously reported, constitutive CRF mRNA levels were elevated in the paraventricular nucleus (PVN) of the hypothalamus in mice that lacked CRFR1, compared to wild-type control mice. Interestingly, this was not the case for transcripts of the immediate early gene NGFI-B. When measured 2 hr after alcohol, PVN NGFI-B gene expression was significantly (p < 0.01) increased in both control and mutant mice, as were CRF mRNA levels in mutant mice, but the hypothalamic responses of the mutants were larger (p < 0.01) than those of the control mice. This difference may be due, at least in part, to the lack of steroid feedback in the mutants. CONCLUSION: These results indicate that although the intraperitoneal injection of alcohol remains capable of eliciting PVN CRF neuronal activation in mice that lack CRFR1, the ACTH and corticosterone responses are significantly blunted, a phenomenon believed to be due to the lack of CRFR1 in the pituitary of these animals.

Adrenocorticotropic Hormone↗

PubMed2000

Distribution of mRNAs encoding CRF receptors in brain and pituitary of rat and mouse.

Two G protein-coupled receptors have been identified that bind corticotropin-releasing factor (CRF) and urocortin (UCN) with high affinity. Hybridization histochemical methods were used to shed light on controversies concerning their localization in rat brain, and to provide normative distributional data in mouse, the standard model for genetic manipulation in mammals. The distribution of CRF-R1 mRNA in mouse was found to be fundamentally similar to that in rat, with expression predominating in the cerebral cortex, sensory relay nuclei, and in the cerebellum and its major afferents. Pronounced species differences in distribution were few, although more subtle variations in the relative strength of R1 expression were seen in several forebrain regions. CRF-R2 mRNA displayed comparable expression in rat and mouse brain, distinct from, and more restricted than that of CRF-R1. Major neuronal sites of CRF-R2 expression included aspects of the olfactory bulb, lateral septal nucleus, bed nucleus of the stria terminalis, ventromedial hypothalamic nucleus, medial and posterior cortical nuclei of the amygdala, ventral hippocampus, mesencephalic raphe nuclei, and novel localizations in the nucleus of the solitary tract and area postrema. Several sites of expression in the limbic forebrain were found to overlap partially with ones of androgen receptor expression. In pituitary, rat and mouse displayed CRF-R1 mRNA signal continuously over the intermediate lobe and over a subset of cells in the anterior lobe, whereas CRF-R2 transcripts were expressed mainly in the posterior lobe. The distinctive expression pattern of CRF-R2 mRNA identifies additional putative central sites of action for CRF and/or UCN. Constitutive expression of CRF-R2 mRNA in the nucleus of the solitary tract, and stress-inducible expression of CRF-R1 transcripts in the paraventricular nucleus may provide a basis for understanding documented effects of CRF-related peptides at a loci shown previously to lack a capacity for CRF-R expression or CRF binding. Other such "mismatches" remain to be reconciled.

Animals↗

PubMed2000

Betaglycan binds inhibin and can mediate functional antagonism of activin signalling.

Activins and inhibins, structurally related members of the TGF-beta superfamily of growth and differentiation factors, are mutually antagonistic regulators of reproductive and other functions. Activins bind specific type II receptor serine kinases (ActRII or IIB) to promote the recruitment and phosphorylation of the type I receptor serine kinase, ALK4 (refs 7-9), which then regulates gene expression by activating Smad proteins. Inhibins also bind type II activin receptors but do not recruit ALK4, providing a competitive model for the antagonism of activin by inhibin. Inhibins fail to antagonize activin in some tissues and cells, however, suggesting that additional components are required for inhibin action. Here we show that the type III TGF-beta receptor, betaglycan, can function as an inhibin co-receptor with ActRII. Betaglycan binds inhibin with high affinity and enhances binding in cells co-expressing ActRII and betaglycan. Inhibin also forms crosslinked complexes with both recombinant and endogenously expressed betaglycan and ActRII. Finally, betaglycan confers inhibin sensitivity to cell lines that otherwise respond poorly to this hormone. The ability of betaglycan to facilitate inhibin antagonism of activin provides a variation on the emerging roles of proteoglycans as co-receptors modulating ligand-receptor sensitivity, selectivity and function.

Activin Receptors↗

PubMed2000

The inhibitory effect of hormones associated with stress on Na appetite of sheep.

Stress is a large stimulus of Na appetite in rabbits, rats, and mice. This study investigated the influence of some peptides implicated in stress, i.e., adrenocorticotropin (ACTH), corticotropin-releasing factor (CRF), and the recently discovered member of the CRF family, urocortin, on the ingestive behavior of sheep. Intracerebroventricular infusion of these peptides over 4 days decreased the need-free Na intake of Na-repleted sheep. Intracerebroventricular infusion of urocortin, however, did not alter Na intake of Na-depleted sheep. Systemic infusion of ACTH increased, whereas systemic infusion of either urocortin or CRF decreased, Na intake of Na-repleted sheep. The increase in Na intake caused by the peripheral infusion of ACTH was blocked by concurrent i.v. infusion of urocortin, substantiating the inhibitory role of this peptide on Na appetite. Central administration of all peptides and i.v. administration of urocortin or urocortin and ACTH combined decreased food intake. Water intake was not directly influenced by the peptides. Rather, decreased water intake, when observed, was secondary to decreased food intake, as determined by pair-feeding experiments. Whereas systemic infusion of ACTH mimics the increase in Na intake observed in several different stressful situations, CRF and urocortin actually inhibit Na intake, indicating a direct central action overriding any effect of these peptides on ACTH release. Indeed, the inhibition of Na intake by urocortin occurred despite its stimulation of ACTH release and the subsequent increase in peripheral level of cortisol. Thus it would appear that hormones associated with stress have both excitatory and inhibitory influences on Na intake. Presumably, other physiological processes entrained by stress also will be important in determining the quantitative outcome on Na appetite.

Adrenocorticotropic Hormone↗

PubMed2000

Fos induction in selective hypothalamic neuroendocrine and medullary nuclei by intravenous injection of urocortin and corticotropin-releasing factor in rats.

CRF and urocortin, administrated systemically, exert peripheral biological actions which may be mediated by brain pathways. We identified brain neuronal activation induced by intravenous (i.v.) injection of CRF and urocortin in conscious rats by monitoring Fos expression 60 min later. Both peptides (850 pmol/kg, i.v.) increased the number of Fos immunoreactive cells in the paraventricular nucleus of the hypothalamus, supraoptic nucleus, central amygdala, nucleus tractus solitarius and area postrema compared with vehicle injection. Urocortin induced a 4-fold increase in the number of Fos-positive cells in the supraoptic nucleus and a 3.4-fold increase in the lateral magnocellular part of the paraventricular nucleus compared with CRF. Urocortin also elicited Fos expression in the accessory hypothalamic neurosecretory nuclei, ependyma lining the ventricles and choroid plexus which was not observed after CRF. The intensity and pattern of the Fos response were dose-related (85, 255 and 850 pmol/kg, i.v.) and urocortin was more potent than CRF. Neither CRF nor urocortin induced Fos expression in the lateral septal nucleus, Edinger-Westphal nucleus, dorsal raphe nucleus, locus coeruleus, or hypoglossal nucleus. These results show that urocortin, and less potently CRF, injected into the circulation at picomolar doses activate selective brain nuclei involved in the modulation of autonomic/endocrine function; in addition, urocortin induces a distinct activation of hypothalamic neuroendocrine neurons.

Amygdala↗

PubMed2000

Identification of a binding site on the type II activin receptor for activin and inhibin.

Type II activin receptors (ActRII and ActRIIB) are single-transmembrane domain serine/threonine kinase receptors that bind activin to initiate the signaling and cellular responses triggered by this hormone. Inhibin also binds type II activin receptors and antagonizes many activin effects. Here we describe alanine scanning mutagenesis of the ActRII extracellular domain. We identify a cluster of three hydrophobic residues (Phe(42), Trp(60), and Phe(83)) that, when individually mutated to alanine in the context of the full-length receptor, cause the disruption of activin and inhibin binding to ActRII. Each of the alanine-substituted ActRII mutants retaining activin binding maintains the ability to form cross-linked complexes with activin and supports activin cross-linking to the type I activin receptor ALK4. Unlike wild-type ActRII, the three mutants unable to bind activin do not cause an increase in activin signaling when transiently expressed in a corticotroph cell line. Together, our results implicate these residues in forming a critical binding surface on ActRII required for functional interactions with both activin and inhibin. This first identification of a transforming growth factor-beta family member binding site may provide a general basis for characterizing binding sites for other members of the superfamily.

Activin Receptors↗

PubMed2000

Behavioral effects of central administration of the novel CRF antagonist astressin in rats.

Astressin, a novel corticotropin releasing factor (CRF) antagonist, has been found to be particularly potent at inhibiting the hypothalamo-pituitary-adrenal axis. The aim of the present study was to determine the effects in rats of astressin in attenuating the anxiogenic-like response produced by social stress and intracerebroventricular (ICV) CRF administration on the elevated plus-maze, and ICV CRF-induced locomotor activation in the rat. Astressin significantly reversed the anxiogenic-like response induced by both social stress and ICV rat/humanCRF (r/hCRF) on the elevated plus-maze, but failed to block the effects of r/hCRF-induced locomotor activity in a familiar environment. When these results were compared to previous studies performed with the same paradigms using other CRF antagonists, astressin showed effects similar to those of D-PheCRF(12-41) on plus-maze performance. However, contrary to alpha-helicalCRF(9-41) and D-PheCRF(12-41), astressin had no effect on CRF-induced locomotor activity. These results suggest that astressin may have a unique anti-CRF profile compared to previously tested antagonists.

Aggression↗

PubMed1999

Reduced anxiety-like and cognitive performance in mice lacking the corticotropin-releasing factor receptor 1.

Corticotropin-releasing factor (CRF) has been hypothesized to be involved in the pathophysiology of anxiety, depression, cognitive and feeding disorders. Two distinct CRF receptor subtypes, CRFR1 and CRFR2, are thought to mediate CRF actions in the CNS. However, the role for each receptor subtype in animal models of neuropsychiatric disorders remains to be determined. Using CRFR1 deficient mice, the present study investigated the functional significance of this CRF receptor subtype in anxiety-like and memory processes. CRFR1 knockout mice displayed an increased exploratory behavior in both the Elevated Plus-maze (EPM) and the Black and White (B-W) test box models of anxiety, indicating an anxiolytic-like effect of the CRFR1 gene deletion. In contrast, during the retrieval trial of a two-trial spatial memory task wild type mice made more visits to and spent more time in the novel arm as opposed to the two familiar ones of a Y-maze apparatus. No increase in the level of exploration of the novel arm by the CRFR1 deficient mice was observed. This indicates that CRFR1 knockout mice are impaired in spatial recognition memory. These results demonstrate that genetic deletion of the CRFR1 receptor can lead to impairments in anxiety-like and cognitive behaviors, supporting a critical role for this receptor in anxiety and cognitive biological processes.

Animals↗

PubMed1999

Corticotropin releasing factor (CRF) agonists with reduced amide bonds and Ser7 substitutions.

Strategies to generate competitive antagonists of bioactive peptides include several possible structural modifications such as the introduction of D-residues and of reduced amide bonds, the substitution of amino acid side chains, dimerization of fragments, and deletion of part of the sequence, among others. Whereas we have identified the two most likely residues responsible for receptor activation in corticotropin releasing factor (CRF) (Ser7 and Leu8)1 and generated potent antagonists by deleting residues 1-8,2,3 the question remained as to whether we could generate CRF antagonists with enhanced affinity after reduction of amide bonds at the N-terminus of CRF or through subtle alteration of those residues' side chains. Reduced amide bond replacements (psi[CH2NH]) between residues 6-9 in oCRF(5-41) (11, 12, 15) analogues consistently yielded potencies of <1% that of oCRF. Except for the 10psi11 and 12psi13 analogues 19 and 20, reduced amide bond replacements were generally well-tolerated in the longer hCRF(4-41) analogues, with the 7psi8-, 8psi9-, and 9psi10-modified peptides (13, 14, 18) yielding potencies that were 2-4 times that of hCRF. Although somewhat promising as agonists, they were, however, 3-7 times less potent than the parent [D-Pro4Nle21,38]-hCRF(4-41) (2). Since O-alkylation of Tyr3 in vasopressin yields an antagonist, and since Ser7 is one of the eight fully conserved residues in the CRF family (inclusive of sauvagine, urocortins, and urotensins) and likely to be critical for receptor binding, we synthesized cyclo(30-33)[Ser(OMe)7,D-Phe12,Nle21,Glu30,Lys33 ,Nle38]Ac-hCRF(7-41) (22), which was found to exhibit full efficacy and 40% of the potency of cyclo(30-33)[D-Phe12,Nle21,Glu30,Lys33, Nle38]Ac-hCRF(7-41) (5). Other substitutions at position 7 included aminoglycine (23, 24) and alkylated and/or acylated [alpha or alpha'-methyl (25-28), alpha'-formyl (29, 30), alpha'-formyl, alpha'-methyl (31), alpha'-acetyl (32), alpha'-acetyl, alpha'-methyl (33)], D- or L-aminoglycines. All analogues were active although less potent than the parent compound 2, and all elicited maximal ACTH response as compared to hCRF. The most potent analogue in this series (33) had the bulkiest side chain, Agl(Me, Ac), and was 60% and 80% as potent as the Ser7 analogue 5 and the Ala7 analogue 6, respectively. In conclusion, we found that neither reduction of the individual amide linkages between residues 6-11 and 12-13 nor introduction of a carbamide moiety in lieu of the side chain of Ser7 led to CRF antagonists.

Adrenocorticotropic Hormone↗

PubMed1999

Corticotropin-releasing factor antagonist attenuates the "anxiogenic-like" effect in the defensive burying paradigm but not in the elevated plus-maze following chronic cocaine in rats.

RATIONALE: Chronic cocaine abuse is associated with the development of anxiogenic states in humans. Corticotropin-releasing factor (CRF) is an endogenous neurotropic factor well known to modulate stress responses. It has been postulated that CRF is involved in the neurobiological mechanisms underlying the anxiety and/or stress responses associated with removal of cocaine after chronic administration. OBJECTIVE: The present study investigated the role of endogenous CRF in mediating the "anxiety-like" effect 48 h after the cessation of saline or chronic cocaine treatment in rats, using the defensive burying paradigm and the elevated plus-maze. METHODS: Rats received daily injections of cocaine (20 mg/kg IP, for 14 consecutive days) or vehicle. Forty-eight hours after the last injection, animals were tested in the plus-maze and then in the defensive burying paradigm. In a second experiment, intracerebroventricular (ICV) cannulae were implanted at the lateral ventricle. Animals were allowed a 1-week period for recovery before starting the chronic drug treatment. The defensive burying testing took place 48 h after cessation of the treatment. The CRF antagonist [DPhe12, Nle21,38, CalphaMe Leu37] r/h CRF(12-41), (also known as D-phe CRF(12-41)) (0.04, 0.2 and 1.0 microg/5 microl) was injected 5 min before the 15-min testing. RESULTS: An "anxiogenic-like" effect following chronic cocaine treatment was demonstrated with the defensive burying paradigm, but not with the elevated plus-maze. This "anxiety-like" response was attenuated by ICV pretreatment with the CRF antagonist D-Phe CRF(12-41), with the highest dose of the CRF antagonist reversing the observed "anxiogenic-like" response. CONCLUSIONS: These data suggest that brain CRF may be substantially involved in the development of "anxiety-like" responses related to cocaine withdrawal and could be important for future drug dependence treatments.

Animals↗

PubMed1999

The disulfide bond arrangement in the extracellular domain of the activin type II receptor.

The initial step in the signaling cascade of the growth factor activin involves its binding to the extracellular domain of the activin type II receptor. This receptor domain contains 10 cysteine residues which are engaged in intramolecular disulfide bonds. To elucidate the structural framework of this domain we have characterized its disulfide-bonding pattern using an extracellular fragment of the receptor which binds activin A with high affinity. By combining proteolysis with mass spectroscopy and chemical sequence analysis, the disulfide connectivity was determined to be as follows: C1-C3, C2-C4, C5-C8, C6-C7, and C9-C10. A similar disulfide arrangement occurs in a family of snake toxins for which the three-dimensional structure is known.

Activin Receptors↗

PubMed1999

Urocortin stimulates placental adrenocorticotropin and prostaglandin release and myometrial contractility in vitro.

Urocortin is a new member of the CRF family. Multiple biological effects for urocortin have been shown in rats and in some in vitro models, showing a modulatory role in hormonal and behavioral functions. Human placenta expresses urocortin, but no information is available on the possible local biological actions. The aim of the present study was to evaluate the effect of urocortin on placental ACTH and prostaglandin (PG) secretion, as well as on myometrial contractility. Various in vitro models were used. For investigating the effect of urocortin on ACTH release, primary cultures of human trophoblast cells were used. Culture media, collected before and after 3 h exposure to different doses of urocortin and ACTH, were measured by RIA. Trophoblast tissue explants were incubated for 24 h in the presence of increasing doses of urocortin, and prostaglandin E2 (PGE2) levels were measured by RIA. Strips of myometrial tissue were incubated in an organ bath and connected to an isometric smooth-muscle transducer in the presence of urocortin, with or without prostaglandin F2alpha (PGF2a). In all these experiments, the effect of astressin (a CRF receptor antagonist) on urocortin-induced actions and the effect of equimolar doses of CRF were evaluated. A dose-related increase of trophoblast ACTH or PGE2 was induced by urocortin, whereas astressin inhibited urocortin-stimulated ACTH or PGE2 release. Equimolar doses of CRF showed a similar effect on both ACTH and PGE2. Urocortin increased PGF2alpha-induced myometrial contractility, and this effect was completely abolished by the addition of astressin. The present study showed that human urocortin stimulates placental secretion of ACTH and PGE2, and modulates myometrial contractility, suggesting a role for this peptide in placental and intrauterine CRF pathways.

Adrenocorticotropic Hormone↗