Activation Of Corticotropin-releasing Factor Receptors Research Articles

Pharmacological Manipulation of Corticotropin-Releasing Factor Receptors in the Anterior and Posterior Subregions of the Insular Cortex Differently Affects Anxiety-Like Behaviors in the Elevated Plus Maze in Rats.

Neuroimaging data in humans and neurobiological studies in rodents have suggested an involvement of the insular cortex (IC) in anxiety manifestations. However, the local neurochemical mechanisms involved are still poorly understood. Corticotropin-releasing factor (CRF) neurotransmission has been described as a prominent neurochemical mechanism involved in the expression of anxiety-like behaviors, but the brain sites related are poorly understood. Additionally, several findings indicate that control of physiological and behavioral responses by the IC occurs in a site-specific manner along its rostrocaudal axis. Thus, this study is aimed at evaluating the effect of CRF receptor agonism and antagonism within the anterior and posterior subregions of the IC in controlling anxiety-related behaviors in the elevated plus maze (EPM). For this, independent groups (six groups) of animals received bilateral microinjections of vehicle, the selective CRF1 receptor antagonist CP376395, or CRF into either the anterior or posterior subregions of the IC. Ten minutes later, the behavior in the EPM was evaluated for five minutes. Treatment of the anterior IC with CP376395, but not with CRF, increased the time and number of entries into the open arms of the EPM. CRF, but not the CRF1 receptor antagonist, microinjected into the posterior IC also increased exploration of the EPM open arms. Taken together, these data indicate that CRFergic neurotransmission in the anterior IC is involved in the expression of anxiety-related behaviors in the EPM. This neurochemical mechanism does not seem to be activated within the posterior IC during exposure to the EPM, but the effects caused by CRF microinjection indicate that activation of CRF receptors in this IC subregion might evoke anxiolytic-like effects.

Abstract 16840: A Novel Long-Acting CRF2 Receptor Agonist COR-1167 Improves Cardiac and Renal Function on Top of Furosemide in a Sheep Model of Pacing Induced Heart Failure

Urocortin-2 (UCN-2), a member of the corticotrophin releasing factor family, has cardiovascular protective effects mediated via activation of CRF2 receptors. Exploratory studies in ovine heart failure (HF) and in HF patients demonstrated that acute intravenous (IV) infusion of UCN-2 (1-4hrs), on top of standard of care, temporarily improved cardiac function. However, the short t 1/2 of recombinant UCN-2 (~15 min) limits its therapeutic utility. A stable CRF2 peptide agonist was developed, named COR-1167, that is potent and selective for CRF2 versus CRF1 receptors and it was tested chronically in vivo. Adult female sheep were anesthetized, and instrumented with 7F His bundle electrodes, arterial, venous and Swan-Ganz catheters, pressure transducers, and flow probes. After recovery, HF was induced by rapid cardiac pacing that was maintained during the study. COR-1167 (5 μg/kg/day SC) in combination with furosemide (0.35 mg/kg/day IV) versus furosemide alone was administered for 5 days and cardiorenal parameters were constantly monitored in conscious sheep. Furosemide had no effect on cardiac function while it increased urine Na + , when compared to pre-treatment measurements. Compared to furosemide, COR-1167 plus furosemide significantly enhanced cardiac and renal function parameters (Table). Mean + SEM for Days 1-5 (CV, biomarker and renal parameters), F (n=5-7), F + COR-1167 (n=6-7), Repeated Measures ANOVA; *BW Day 0 minus Day 5 = BW change, Unpaired t test. COR-1167 cardiorenal benefits occurred within 2 hours of the first dose and were sustained, without changing SBP. It increased natriuresis and decreased net fluid balance (water intake minus urine volume) and BW (reflecting a reduction in congestion). In conclusion, chronic CRF2 receptor activation by COR-1167 significantly improved both cardiac and renal function on top of furosemide in ovine HF and potentially is a unique therapy for HF patients.

Corticotropin-releasing factor receptor activation in the nucleus accumbens shell induces pup neglect in lactating rats

Corticotropin-releasing factor receptor activation in the nucleus accumbens shell induces pup neglect in lactating rats

Estrus cycle-dependent working memory effects of prefrontal cortex corticotropin-releasing factor neurotransmission.

The prefrontal cortex (PFC) supports a diversity of cognitive processes. Impairment in PFC-dependent cognition is associated with multiple psychiatric disorders, including those known to display sex differences. Our ability to treat this impairment is limited, in part due to an incomplete understanding of the neural mechanisms that support PFC-dependent cognition. In previous studies in male rats, we demonstrated that corticotropin-releasing factor (CRF) receptors and neurons in caudal dorsomedial PFC (dmPFC) regulate PFC-dependent working memory. Subcortically, CRF can exert sex-specific actions, a subset of which are ovarian steroid dependent. To date, the cognitive actions of dmPFC CRF neurotransmission in females are unknown. To address this gap, the current studies examined the effects of chemogenetic and pharmacological manipulations of CRF receptors and neurons within the dmPFC of female rats tested in a spatial working memory task. Outside of proestrus, activation of both CRF receptors and neurons in the caudal, but not rostral, dmPFC impaired working memory. Meanwhile, blockade of CRF receptors in the caudal dmPFC or globally in the brain, improved working memory performance, similar to that seen in males. In contrast, these effects were not observed during proestrus. These observations demonstrate that while CRF neurotransmission in the PFC regulates working memory similarly in males and females, these actions are not observed in females when ovarian steroids are at peak levels.

Role of CRF1 and CRF2 receptors in the lateral hypothalamus in anxiogenic response evoked by acute and repeated restraint stress in rats

The lateral hypothalamus (LH) is an important hypothalamic area that has been implicated in the integration of behavioral and physiological responses to stress. The corticotropin‐releasing factor (CRF) system has been shown to be an important neurochemical mechanism in the central nervous system involved in the etiology of the behavioral changes evoked by exposure to aversive situations. The exposure to chronic stress affects the expression of CRF neurotransmission components in brain regions controlling stress responses. However, the role of CRF neurotransmission within the LH in the anxiogenic responses evoked by acute and repeated restraint stress (RS) has not been investigated. Here, was investigated the effect of bilateral microinjection into the LH of the selective CRF1 receptor antagonist CP376395 or the selective CRF2 receptor antagonist antisauvagine‐30 (ASV‐30) in the anxiogenic responses following the first and the 10th session of restraint stress (RS) in male rats. For this, male Wistar rats (250g) had cannula‐guide bilaterally implanted within the LH. The RS was realized by placing the animals in a plastic cylindrical tube for 60 minutes. Independent sets of animals received CP376395 (0.01 mol/100nL), ASV‐30 (0.01 mol/100nL) or vehicle into the LH 10 minutes before the onset of the first or 10th session of RS. Immediately after the end of the stress session, the animals were individually placed in the elevated plus maze (EPM) for 5 minutes to assess anxiety‐related behaviors. The behavioral analysis revealed that an acute RS session decreased the % of entries and time in the open arms of the EPM in vehicle‐treated animals, when compared to animals that were not submitted to restraint stress (i.e., naïve group) (P<0.05). Bilateral microinjection of either CP376395 or ASV‐30 into the LH inhibited the decreased open‐arms exploration caused by acute restraint stress (P<0.05). We did not identify behavioral changes in the EPM in animals repeatedly subjected to restraint stress, and pharmacological treatments also had no behavioral effect when administered before the 10th RS session (P>0.05). Taken together, the results indicates that the CRFergic neurotransmission in the LH, acting through activation of both CRF1 and CRF2 receptors, is involved in the generation of anxiety‐like behaviors induced by acute restraint stress, and the previous repeated exposure to this stressor affect this control.

Moderate prenatal alcohol exposure modifies sex-specific CRFR1 activity in the central amygdala and anxiety-like behavior in adolescent offspring.

Anxiety disorders are highly prevalent among individuals with a history of prenatal alcohol exposure (PAE), and adolescent rodents demonstrate anxiety-like behavior following moderate PAE on Gestational Day (G) 12. A likely systemic target of PAE is the stress peptide corticotropin-releasing factor (CRF), as activation of CRF receptor 1 (CRFR1) in the medial nucleus of the central amygdala (CeM) is known to increase anxiety-like behavior in adults. To determine if CRF-CRFR1 interactions underly PAE-induced anxiety, functional changes in CRF system activity were investigated in adolescent male and female Sprague Dawley rats following G12 PAE. Compared to air-exposed controls, PAE increased basal spontaneous (s) inhibitory postsynaptic current (IPSC) frequency in the CeM of males, but not females. Furthermore, PAE blunted CRFR1-regulated miniature (m) IPSCs in a sex- and concentration-specific manner, and only PAE males demonstrated tonic CRFR1 activity in the CeM. It was further determined that G12 PAE decreased CRFR1 mRNA in the CeM of males while increasing regional expression in females. Finally, infusion of a CRFR1 agonist into the CeM of adolescents produced a blunted expression of CRFR1-induced anxiety-like behavior exclusively in PAE males, mirroring the blunted physiology demonstrated by PAE males. Cumulatively, these data suggest that CRFR1 function within the CeM is age- and sex-specific, and PAE not only increases the expression of anxiety-like behavior, but may reduce the efficacy of treatment for PAE-induced anxiety through CRFR1-associated mechanisms. Therefore, future research will be necessary to develop targeted treatment of anxiety disorders in individuals with a history of PAE.

Central amygdala corticotropin-releasing factor neurons promote hyponeophagia but do not control alcohol drinking in mice.

Corticotropin-releasing factor (CRF) signaling in the central nucleus of the amygdala (CeA) plays a critical role in rodent models of excessive alcohol drinking. However, the source of CRF acting in the CeA during alcohol withdrawal remains to be identified. In the present study, we hypothesized that CeA CRF interneurons may represent a behaviorally relevant source of CRF to the CeA increasing motivation for alcohol via negative reinforcement. We first observed that Crh mRNA expression in the anterior part of the mouse CeA correlates positively with alcohol intake in C57BL/6J males with a history of chronic binge drinking followed by abstinence and increases upon exposure to chronic intermittent ethanol (CIE) vapor inhalation. We then found that chemogenetic activation of CeA CRF neurons in Crh-IRES-Cre mouse brain slices increases gamma-aminobutyric acid (GABA) release in the medial CeA, in part via CRF1 receptor activation. While chemogenetic stimulation exacerbated novelty-induced feeding suppression (NSF) in alcohol-naïve mice, thereby mimicking the effect of withdrawal from CIE, it had no effect on voluntary alcohol consumption, following either acute or chronic manipulation. Furthermore, chemogenetic inhibition of CeA CRF neurons did not affect alcohol consumption or NSF in chronic alcohol drinkers exposed to air or CIE. Altogether, these findings indicate that CeA CRF neurons produce local release of GABA and CRF and promote hyponeophagia in naïve mice, but do not drive alcohol intake escalation or negative affect in CIE-withdrawn mice. The latter result contrasts with previous findings in rats and demonstrates species specificity of CRF circuit engagement in alcohol dependence.

Roles of corticotropin-releasing factor signaling in the lateral habenula in anxiety-like and alcohol drinking behaviors in male rats

Corticotropin-releasing factor (CRF) signaling in the mesocorticolimbic system is known to modulate anxiety-like behavior and alcohol consumption, behaviors that also have been associated with the hyper-glutamatergic state of the lateral habenula (LHb) neurons in rats. However, the role of CRF signaling in the LHb on the glutamate transmission, anxiety-like behaviors and alcohol consumption is unknown. Here, we used male rats that had been consuming alcohol for three months to address this gap in the literature. First, using electrophysiological techniques, we evaluated CRF's effects on the glutamate transmission in LHb neurons in brain slices. CRF facilitated glutamate transmission. The facilitation was greater in neurons of alcohol-withdrawing rats than in those of naïve rats. The facilitation was mimicked by the activation of CRF receptor 1 (CRF1R) but attenuated by the activation of CRF receptor 2 (CRF2R). This facilitation was mediated by upregulating CRF1R-protein kinase A signaling. Conversely, protein kinase C blockade attenuated CRF's facilitation in neurons of naïve rats but promoted it in neurons of alcohol-withdrawing rats. Next, using site-direct pharmacology, we evaluated the role of CRF signaling in the LHb on anxiety-like behaviors and alcohol consumption. Intra-LHb inhibition of CRF1R or activation of CRF2R ameliorated the anxiety-like behaviors in alcohol-withdrawing rats and reduced their alcohol intake when drinking was resumed. These observations provide the first direct behavioral pharmacological and cellular evidence that CRF signaling in the LHb modulates glutamate transmission, anxiety-like behaviors and alcohol consumption, and that adaptation occurs in CRF signaling in the LHb after chronic alcohol consumption.

Role of Basolateral Amygdalar Somatostatin 2 Receptors in a Rat Model of Chronic Anxiety

Repeated exposure to stress has been implicated in inducing chronic anxiety states. Stress related increases in anxiety responses are likely mediated by activation of corticotropin-releasing factor receptors (CRFR) in the amygdala, particularly the basolateral amygdala (BLA). Within the BLA, acute injections of the CRFR agonist urocortin 1 (Ucn1) leads to acute anxiety, whereas repeated daily injections of subthreshold-doses of Ucn1 produces a long-lasting, persistent anxiety-like phenotype, a phenomenon referred to as Ucn1-priming. Relative gene expressions from the BLA of vehicle and Ucn1-primed rats were analyzed with quantitative RT-PCR using a predesigned panel of 82 neuroscience-related genes. Compared to vehicle-primed rats, only expression of the somatostatin receptor 2 gene (Sstr2) was significantly reduced in the BLA of Ucn1-primed rats. The contribution of Sstr2 on an anxiety phenotype was tested by injecting a Sstr2 antagonist into the BLA in un-primed rats. The Sstr2 antagonist increased anxiety-like behavior. Notably, pretreatment with Sstr2 agonist injected into the BLA blocked anxiety-inducing effects of acute Ucn1 BLA-injections and delayed anxiety expression during Ucn1-priming. However, concomitant Sstr2 agonist pretreatment during Ucn-1 priming did not prevent either the development of a chronic anxiety state or a reduction of BLA Sstr2 expression induced by priming. The data demonstrate that the persistent anxiety-like phenotype observed with Ucn1-priming in the BLA is associated with a selective reduction of Sstr2 gene expression. Although Sstr2 activation in the BLA blocks acute anxiogenic effects of stress and down-regulation of BLA Sstr2, it does not suppress the long-term consequences of prolonged exposure to stress-related challenges.

Functional and Anatomical Characterization of Corticotropin-Releasing Factor Receptor Subtypes of the Rat Spinal Cord Involved in Somatic Pain Relief

Corticotropin-releasing factor (CRF) orchestrates our body’s response to stressful stimuli. Pain is often stressful and counterbalanced by activation of CRF receptors along the nociceptive pathway, although the involvement of the CRF receptor subtypes 1 and/or 2 (CRF-R1 and CRF-R2, respectively) in CRF-induced analgesia remains controversial. Thus, the aim of the present study was to examine CRF-R1 and CRF-R2 expression within the spinal cord of rats with Freund’s complete adjuvant-induced unilateral inflammation of the hind paw using reverse transcriptase polymerase chain reaction, Western blot, radioligand binding, and immunofluorescence confocal analysis. Moreover, the antinociceptive effects of intrathecal (i.t.) CRF were measured by paw pressure algesiometer and their possible antagonism by selective antagonists for CRF-R1 and/or CRF-R2 as well as for opioid receptors. Our results demonstrated a preference for the expression of CRF-R2 over CRF-R1 mRNA, protein, binding sites and immunoreactivity in the dorsal horn of the rat spinal cord. Consistently, CRF as well as CRF-R2 agonists elicited potent dose-dependent antinociceptive effects which were antagonized by the i.t. CRF-R2 selective antagonist K41498, but not by the CRF-R1 selective antagonist NBI35965. In addition, i.t. applied opioid antagonist naloxone dose-dependently abolished the i.t. CRF- as well as CRF-R2 agonist-elicited inhibition of somatic pain. Importantly, double immunofluorescence confocal microscopy of the spinal dorsal horn showed CRF-R2 on enkephalin (ENK)-containing inhibitory interneurons in close opposition of incoming mu-opioid receptor-immunoreactive nociceptive neurons. CRF-R2 was, however, not seen on pre- or on postsynaptic sensory neurons of the spinal cord. Taken together, these findings suggest that i.t. CRF or CRF-R2 agonists inhibit somatic inflammatory pain predominantly through CRF-R2 receptors located on spinal enkephalinergic inhibitory interneurons which finally results in endogenous opioid-mediated pain inhibition.

Otilonium Bromide treatment prevents nitrergic functional and morphological changes caused by chronic stress in the distal colon of a rat IBS model.

Irritable bowel syndrome (IBS) is a highly prevalent gastrointestinal disorder characterized by periods of remission and exacerbation. Among the risk factors to develop IBS, psychosocial stress is widely acknowledged. The water avoidance stress repeatedly applied (rWAS) is considered effective to study IBS etio‐pathogenesis. Otilonium bromide (OB), a drug with multiple mechanisms of action, is largely used to treat IBS patients. Orally administered, it concentrates in the large bowel and significantly ameliorates the IBS symptomatology. Presently, we tested whether rWAS rats developed neuro‐muscular abnormalities in the distal colon and whether OB treatment prevented them. The investigation was focussed on the nitrergic neurotransmission by combining functional and morphological methodologies. The results confirm rWAS as reliable animal model to investigate the cellular mechanisms responsible for IBS: exposure to one‐hour psychosocial stress for 10 days depressed muscle contractility and increased iNOS expression in myenteric neurons. OB treatment counteracted these effects. We hypothesize that these effects are due to the corticotropin‐releasing factor (CRF) release, the main mediator of the psychosocial stress, followed by a CRF1receptor activation. OB, that was shown to prevent CRF1r activation, reasonably interrupted the cascade events that bring to the mechanical and immunohistochemical changes affecting rWAS rat colon.

Prefrontal corticotropin-releasing factor neurons impair sustained attention via distal transmitter release.

The prefrontal cortex (PFC) supports cognitive processes critical for goal-directed behavior. Although the PFC contains a high density of corticotropin-releasing factor (CRF) neurons, their role in cognition has been largely unexplored. We recently demonstrated that CRF neurons in the caudal dorsomedial PFC (dmPFC) of rats act to impair working memory via activation of local CRF receptors. However, there is heterogeneity in the neural mechanisms that support the diversity of PFC-dependent cognitive processes. Currently, the degree to which PFC CRF neurons impact other forms of PFC-dependent cognition is unknown. To address this issue, the current studies examined the effects of chemogenetic manipulations of PFC CRF neurons on sustained attention in male rats. Similar to working memory, activation of caudal dmPFC CRF neurons impaired, while inhibition of these neurons or global CRF receptor antagonism improved, sustained attention. However, unlike working memory, the sustained attention-impairing effect of PFC CRF neurons was not dependent on local CRF receptors. Moreover, CRF infusion into the caudal dmPFC or other medial PFC subregions had no effect on task performance. Together, these observations demonstrate that while caudal dmPFC CRF neurons impair both working memory and sustained attention, these actions involve distinct neural circuits (local CRF release for working memory and extra-PFC release for sustained attention). Nonetheless, the procognitive actions of systemically administered CRF antagonists across both tasks are similar to those seen with attention deficit hyperactivity disorder-related treatments. Thus, CRF antagonists may have potential for use in the treatment of PFC cognitive dysfunction.

Role of CRF 2 Receptors in Colonic Ion Secretion in Response to Stress‐related Peptides

Background and Aims Stress increases intestinal secretion and exacerbates symptoms of irritable bowel syndrome (IBS). Peripherally-derived corticotropin-releasing factor (CRF) is known to mediate stress-induced intestinal secretion, presumably by activation of CRF1 receptors in the gut. The present study aimed to ascertain the role of CRF2 activation in intestinal secretion by three other members of CRF peptide family, urocortin (UCN) 1-3, in wild-type (WT) and CRF2 knockout (Crhr2-/-) mice. Methods Mucosal/submucosal preparations from proximal colon of WT and Crhr2-/- mice of adult male mice were mounted in Ussing chambers for measurement of short-circuit current (Isc) as an indicator of ion secretion. Results The colonic mucosa/submucosa preparations were exposed to different concentrations of UCN1, 2, or 3 (30 nM, 100 nM, 300 nM, and 1 mM) on the basolateral side. All three UCNs evoked increases in colonic ISC (ΔISC) in a dose-dependent manner. The EC50s were 392.64nM for UCN1, 103.28nM for UCN2, and 231.21nM for UCN3. Application of CRF or stressin1 (a selective CRF1 receptor agonist) (30nM – 1mM) also caused increases in ISC with an EC50 of 597.04nM or 588.84nM, respectively. The potency order was therefore UCN2 > UCN3 > UCN1 > stressin1 » CRF. At each concentration tested, UCN3 produced significantly greater ΔISC responses than that caused by stressin1, CRF, UCN1, or UCN2. Involvement of the CRF receptor subtypes in UCN1-3 evoked ΔISC responses was investigated with the selective CRF1 receptor antagonist NBI27914 and the selective CRF2 receptor antagonist antisauvagine-30. Both NBI27914 and antisauvagine-30 significantly suppressed UCN1 (300nM)-induced ΔISC responses (p < 0.05). The ΔISC responses evoked by UCN2 (100nM) and UCN3 (100nM) were significantly suppressed by antisauvagine-30 (p < 0.05 and p < 0.0001, respectively), but were not influenced by NBI27914 (p > 0.05). In the Crhr2-/- mice, UCN2 (1mM) failed to evoke any ΔISC compared with WT controls (p < 0.01). UCN3 (1mM)-evoked ΔISC responses were significantly decreased by 57.21% in the Crhr2-/- mice compared with WT mice (p < 0.05). In contrast, stressin1 (1mM)-evoked ΔISC response did not differ between WT and Crhr2-/- mice (p > 0.05). Conclusions UCN1-3 all caused an increase of colonic ion secretion, which mimicked the effects of stress and CRF. UCN1-induced increase of colonic ion secretion was mediated by both CRF1 and CRF2, whereas UCN2 and UCN3-induced increase of colonic ion secretion was mediated only by CRF2. Since UCN2 and UCN3 have greater potency and efficacy on colonic ion secretion compared with UCN1, CRF, and stressin1, it is logical to conclude that CRF2 receptors played a predominant role in the pro-secretory effects of UCNs.

Effects of stress-related peptides on chloride secretion in the mouse proximal colon.

Stress increases intestinal secretion and exacerbates symptoms of irritable bowel syndrome (IBS). Peripherally derived corticotropin-releasing factor (CRF) is known to mediate stress-induced intestinal secretion, presumably by activation of CRF1 receptors in the gut. The present study aimed to ascertain the role of CRF2 activation in intestinal secretion by three other members of CRF peptide family, urocortin (UCN) 1-3, in wild type (WT) and CRF2 knockout (Crhr2-/- ) mice. Mucosal/submucosal preparations from proximal colon of WT and Crhr2-/- mice of both sexes were mounted in Ussing chambers for measurement of short-circuit current (Isc ) as an indicator of ion secretion. Male mice demonstrated a significantly higher baseline Isc than female in both WT and Crhr2-/- genotypes. CRF and UCN1-3 (1μM) caused greater increases in colonic Isc (ΔIsc ) in male than female. Colonic Isc response to the selective CRF1 agonist, stressin1, was similar in both sexes. In male mice, the selective CRF2 agonists (UCN2 and UCN3) caused significantly greater ΔIsc than CRF and stressin1. UCN2- and UCN3-evoked ΔISC was significantly reduced in preparations pretreated with the selective CRF2 antagonist antisauvagine-30 and in Crhr2-/- mice. The prosecretory effects of urocortins were due to increases in Cl- secretion and involved enteric neurons and mast cells. The findings revealed sex differences in baseline colonic secretion and responses to stress-related peptides. CRF2 receptors play a more prominent role in colonic secretion in male mice. The greater baseline secretion and responses to UCNs may contribute to the higher prevalence of diarrhea-predominant IBS in males.

Molecular Basis for Hormone Recognition and Activation of Corticotropin-Releasing Factor Receptors.

Corticotropin-releasing factor (CRF) and the three related peptides urocortins 1-3 (UCN1-UCN3) are endocrine hormones that control the stress responses by activating CRF1R and CRF2R, two members of class B G-protein-coupled receptors (GPCRs). Here, we present two cryoelectron microscopy (cryo-EM) structures of UCN1-bound CRF1R and CRF2R with the stimulatory G protein. In both structures, UCN1 adopts a single straight helix with its N terminus dipped into the receptor transmembrane bundle. Although the peptide-binding residues in CRF1R and CRF2R are different from other members of class B GPCRs, the residues involved in receptor activation and G protein coupling are conserved. In addition, both structures reveal bound cholesterol molecules to the receptor transmembrane helices. Our structures define the basis of ligand-binding specificity in the CRF receptor-hormone system, establish a common mechanism of class B GPCR activation and G protein coupling, and provide a paradigm for studying membrane protein-lipid interactions for class B GPCRs.

Activation of CRF2 receptor increases gastric vagal afferent mechanosensitivity.

Gastric vagal afferent (GVA) sensing of food-related mechanical stimuli is a crucial mechanism in the control of feeding behavior and gastric function. Stress is an important factor contributing to eating disorders and gastric diseases. Chronic stress has been shown to increase the mechanosensitivity of GVAs in mice and to reduce food intake and body weight. Whether the mechanosensitivity of GVAs is modulated by stress hormones is not known. This study aimed to determine the effect of stress hormones on GVA mechanosensitivity. The expression of stress hormone receptors in GVA cell bodies was determined in 8-wk-old male C57BL/6 mice using quantitative RT-PCR combined with laser capture microdissection. The mechanosensitivity of GVAs was determined in the absence and presence of stress hormones using an in vitro single-fiber recording preparation. NR3C1 and CRHR2 (mRNA isoforms of glucocorticoid receptor and CRF2 receptor, respectively) were expressed in GVA neurons. The glucocorticoid receptor agonist corticosterone had no effect on the mechanosensitivity of either tension or mucosal GVAs. Activation of CRF2 receptor by its specific analog, urocortin 3, significantly increased the mechanosensitivity of both tension and mucosal GVAs, an effect prevented by the CRF2 receptor antagonist astressin 2B. In conclusion, activation of CRF2 receptor increases the mechanosensitivity of GVAs. This may contribute to the stress- and CRF2 receptor-associated changes in feeding behavior and gastric function, possibly contributing to the hypersensitivity of GVAs in chronic stress conditions.NEW & NOTEWORTHY Gastric vagal afferents (GVAs) relay food-related signals to the central nervous system, where they are processed, eventually leading to modulation of food intake and gastric function. GVA signaling can be modulated by an array of hormones. Stress has been shown to induce GVA hypersensitivity. This study demonstrates that GVA neurons express subtypes of stress hormone receptors, specifically CRF2. Furthermore, activation of CRF2 receptor increases GVA mechanosensitivity, which could have implications for food intake and gastric function.

Contribution of Corticotropin-Releasing Factor Receptor 1 (CRF1) to Serotonin Receptor 5-HT2CR Function in Amygdala Neurons in a Neuropathic Pain Model.

The amygdala plays a key role in emotional-affective aspects of pain and in pain modulation. The central nucleus (CeA) serves major amygdala output functions related to emotional-affective behaviors and pain modulation. Our previous studies implicated the corticotropin-releasing factor (CRF) system in amygdala plasticity and pain behaviors in an arthritis model. We also showed that serotonin (5-HT) receptor subtype 5-HT2CR in the basolateral amygdala (BLA) contributes to increased CeA output and neuropathic pain-like behaviors. Here, we tested the novel hypothesis that 5-HT2CR in the BLA drives CRF1 receptor activation to increase CeA neuronal activity in neuropathic pain. Extracellular single-unit recordings of CeA neurons in anesthetized adult male rats detected increased activity in neuropathic rats (spinal nerve ligation model) compared to sham controls. Increased CeA activity was blocked by local knockdown or pharmacological blockade of 5-HT2CR in the BLA, using stereotaxic administration of 5-HT2CR short hairpin RNA (shRNA) viral vector or a 5-HT2CR antagonist (SB242084), respectively. Stereotaxic administration of a CRF1 receptor antagonist (NBI27914) into the BLA also decreased CeA activity in neuropathic rats and blocked the facilitatory effects of a 5-HT2CR agonist (WAY161503) administered stereotaxically into the BLA. Conversely, local (BLA) knockdown of 5-HT2CR eliminated the inhibitory effect of NBI27914 and the facilitatory effect of WAY161503 in neuropathic rats. The data suggest that 5-HT2CR activation in the BLA contributes to neuropathic pain-related amygdala (CeA) activity by engaging CRF1 receptor signaling.

Corticotropin Releasing Factor Type 1 and 2 Receptor Signaling in the Medial Prefrontal Cortex Modulates Binge-Like Ethanol Consumption in C57BL/6J Mice.

Corticotropin releasing factor (CRF) signaling via limbic CRF1 and 2 receptors (CRF1R and CRF2R, respectively) is known to modulate binge-like ethanol consumption in rodents. Though CRF signaling in the medial prefrontal cortex (mPFC) has been shown to modulate anxiety-like behavior and ethanol seeking, its role in binge ethanol intake is unknown. Here, we used “drinking-in-the-dark” (DID) procedures in male and female C57BL/6J mice to address this gap in the literature. First, the role of CRF1R and CRF2R signaling in the mPFC on ethanol consumption was evaluated through site-directed pharmacology. Next, we evaluated if CRF1R antagonist reduction of binge-intake was modulated in part through CRF2R activation by co-administration of a CRF1R and CRF2R antagonist. Intra-mPFC inhibition of CRF1R and activation of CRF2R resulted in decreased binge-like ethanol intake. Further, the inhibitory effect of the CRF1R antagonist was attenuated by co-administration of a CRF2R antagonist. We provide novel evidence that (1) inhibition of CRF1R or activation of CRF2R in the mPFC reduces binge-like ethanol intake; and (2) the effect of CRF1R antagonism may be mediated via enhanced CRF2R activation. These observations provide the first direct behavioral pharmacological evidence that CRF receptor activity in the mPFC modulates binge-like ethanol consumption.

Corticotropin-releasing factor depolarizes rat lateral vestibular nuclear neurons through activation of CRF receptors 1 and 2

Corticotropin-releasing factor (CRF) is a neuropeptide mainly synthesized in the hypothalamic paraventricular nucleus and has been traditionally implicated in stress and anxiety. Intriguingly, genetic or pharmacological manipulation of CRF receptors affects locomotor activity as well as motor coordination and balance in rodents, suggesting an active involvement of the central CRFergic system in motor control. Yet little is known about the exact role of CRF in central motor structures and the underlying mechanisms. Therefore, in the present study, we focused on the effect of CRF on the lateral vestibular nucleus (LVN) in the brainstem vestibular nuclear complex, an important center directly contributing to adjustment of muscle tone for both postural maintenance and the alternative change from the extensor to the flexor phase during locomotion. The results show that CRF depolarizes and increases the firing rate of neurons in the LVN. Tetrodotoxin does not block the CRF-induced depolarization and inward current on LVN neurons, suggesting a direct postsynaptic action of the neuropeptide. The CRF-induced depolarization on LVN neurons was partly blocked by antalarmin or antisauvagine-30, selective antagonists for CRF receptors 1 (CRFR1) and 2 (CRFR2), respectively. Furthermore, combined application of antalarmin and antisauvagine-30 totally abolished the CRF-induced depolarization. Immunofluorescence results show that CRFR1 and CRFR2 are co-localized in the rat LVN. These results demonstrate that CRF excites the LVN neurons by co-activation of both CRFR1 and CRFR2, suggesting that via the direct modulation on the LVN, the central CRFergic system may actively participate in the central vestibular-mediated postural and motor control.

Pretreatment with a CRF antagonist amplifies feeding inhibition induced by fourth ventricular cocaine- and amphetamine-regulated transcript peptide

BackgroundPre-treatment with the corticotropin-releasing factor antagonist α-helical CRF9-41 prevents inhibition of gastric emptying by cocaine-and amphetamine-regulated transcript peptide at a dorsal hindbrain level, but its inhibition of sucrose intake is not affected. This is suggestive of separable underlying mechanisms of action in the caudal brainstem for cocaine-and amphetamine-regulated transcript peptide with regard to food intake and gastrointestinal functions. Here we further examine cocaine-and amphetamine-regulated transcript peptide—corticotropin-releasing factor receptor interactions in caudal brainstem controls of solid food intake. Injections of combinations of vehicle, cocaine-and amphetamine-regulated transcript peptide (0.5 μg or 1 μg) or α-helical CRF9-41 were given into the fourth cerebral ventricle of rats. Nocturnal solid food intake was recorded over 22 h.ResultsPre-treatment with α-helical CRF9-41 into the fourth ventricle significantly increased the responsivity to cocaine-and amphetamine-regulated transcript peptide on hypophagia. In a separate control experiment, α-helical CRF9-41 pre-treatment blocked CRF-induced food intake inhibition indicative of its antagonistic effectiveness.ConclusionsWe conclude that an endogenous Corticotropin-releasing factor agonist may modulate suppression of food intake caused by cocaine-and amphetamine-regulated transcript peptide at a dorsal hindbrain level in the absence of stress. A potential caudal brainstem mechanism whereby cocaine-and amphetamine-regulated transcript peptide effects on food intake is attenuated via corticotropin-releasing factor receptor activity causing tonic inhibition, is suggested.