Corticotropin-releasing Factor Receptor Antagonist Research Articles

Role of CRF1 and CRF2 Receptors in Stress-induced Increase of Intestinal Epithelial Permeability in the Mouse Colon

Background and Aims: Stress is known to increase intestinal permeability, which may contribute to the development of irritable bowel syndrome (IBS) and other intestinal disorders. Corticotropin releasing factor (CRF) has been proven to mediate stress-induced changes in gut function, such as increased colonic motility, ion secretion, and intestinal permeability. Actions of CRF are mediated by two G-protein coupled receptors, CRF1 and CRF2; both are expressed in the intestine. However, the roles of the CRF1 and CRF2 receptors in stress-induced increase of intestinal epithelial permeability remain unknown. Methods: Adult male and female C57BL/6 mice were restrained (1h/d) for five consecutive days. The selective CRF1 and CRF2 receptor antagonists were injected intraperitoneally 15 min prior to the restraint stress. Control mice were kept in their home cages without restraint. After the last restraint stress/control session, the proximal colon was removed, and the mucosa/submucosa preparations were dissected and mounted onto the Ussing Chamber System for the measurement of intestinal permeability. Transepithelial resistance (TER) and FITC-inulin (4kD) flux across the colonic epithelium were used as indicators of paracellular permeability. Horseradish peroxidase (HRP, 40kD) flux was used to measure transcellular permeability. Results: Restraint stress did not alter TER or FITC-inulin flux in male mice. Pretreatment with the CRF1 receptor antagonist NBI 27914 or the CRF2 receptor antagonist antisauvagine-30 had no effect on TER or FITC-inulin flux in male mice. In female mice, restraint stress significantly reduced TER and increased FITC-inulin flux in the proximal colon, which were blocked by NBI 27914 but not by antisauvagine-30. Restraint stress significantly increased HRP flux in both male and female mice. In male mice, pretreatment with NBI 27914 significantly reduced stress-evoked increase of HRP flux, while antisauvagine-30 had no effect. In female mice, both NBI 27914 and antisauvagine-30 significantly reduced stress-evoked increase of HRP flux. Conclusions: Stress evokes different changes in epithelial permeability in the proximal colon of male and female mice. In male mice, stress does not influence paracellular permeability but increases transcellular permeability, which is mediated by the CRF1 receptor. In female mice, stress increases both transcellular and paracellular permeability. Stress-induced increase of paracellular permeability was mediated by the CRF1 receptor, but the increased transcellular permeability was mediated by both CRF1 and CRF2 receptors. University of Wisconsin-La Crosse Undergraduate Research Grant (CS and LB). This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

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.

Role of corticotropin-releasing factor neurotransmission in the lateral hypothalamus on baroreflex impairment evoked by chronic variable stress in rats.

Despite the importance of physiological responses to stress in a short-term, chronically these adjustments may be harmful and lead to diseases, including cardiovascular diseases. The lateral hypothalamus (LH) has been reported to be involved in expression of physiological and behavioral responses to stress, but the local neurochemical mechanisms involved are not completely described. The corticotropin-releasing factor (CRF) neurotransmission is a prominent brain neurochemical system implicated in the physiological and behavioral changes induced by aversive threats. Furthermore, chronic exposure to aversive situations affects the CRF neurotransmission in brain regions involved in stress responses. Therefore, in this study, we evaluated the influence of CRF neurotransmission in the LH on changes in cardiovascular function and baroreflex activity induced by chronic variable stress (CVS). We identified that CVS enhanced baseline arterial pressure and impaired baroreflex function, which were followed by increased expression of CRF2, but not CRF1, receptor expression within the LH. Local microinjection of either CRF1 or CRF2 receptor antagonist within the LH inhibited the baroreflex impairment caused by CVS, but without affecting the mild hypertension. Taken together, the findings documented in this study suggest that LH CRF neurotransmission participates in the baroreflex impairment related to chronic stress exposure.

The role of hypothalamic Orexin-A in stress-induced gastric dysmotility: An agonistic interplay with corticotropin releasing factor.

Central Orexin-A (OXA) modulates gastrointestinal (GI) functions and stress response. This study aimed to investigate whether OXA and CRF interact at hypothalamic level. Solid gastric emptying (GE), fecal output (FO), plasma corticosterone (CORT), and postprandial antro-pyloric motility were assessed in rats that underwent acute restraint stress (ARS) and pretreated with central OX1R and/or CRF receptor antagonists SB-334867 and alpha-helical CRF9,41 . Microdialysis was performed to assess ARS-induced release of OXA and CRF in PVN and LHA, respectively. Immunofluorescence labeling was performed to detect the stress-induced changes in OXA and to assess the hypothalamic distribution of OX1R and CRF1/2 receptors. ARS-induced c-Fos immunoreactivity was evaluated in PVN and LHA of rats received OX1R and CRF receptor antagonists. ARS delayed GE by disturbing the coordination of antro-pyloric contractions while stimulating FO and CORT secretion. ARS-induced alterations in GE, FO, plasma CORT, and antro-pyloric motility were attenuated by OX1R and/or CRF receptor antagonists, however, these changes were completely restored in rats received both antagonists. ARS stimulated release of OXA and CRF which were significantly attenuated by α-CRF9,41 and SB-334867, respectively. The OX1R was detected in CRF-immunoreactive cells, whereas dense expression of CRF2 receptor but not CRF1 was observed in LHA. ARS remarkably increased OXA immunoreactivity in LHA. ARS-induced c-Fos expression in LHA and PVN was abolished by α-CRF9,41 and SB-334867, respectively. Our findings suggest a reciprocal contribution of OXA and CRF which seems to be involved in the mediation of stress-induced alterations in neuroendocrine and GI motor functions.

The Impact of Heroin Self-Administration and Environmental Enrichment on Ventral Tegmental CRF1 Receptor Expression.

There is a strong link between chronic stress and vulnerability to drug abuse and addiction. Corticotropin releasing factor (CRF) is central to the stress response that contributes to continuation and relapse to heroin abuse. Chronic heroin exposure can exacerbate CRF production, leading to dysregulation of the midbrain CRF-dopamine-glutamate interaction. Here we investigated the role of midbrain CRF1 receptors in heroin self-administration and assessed neuroplasticity in CRF1 receptor expression in key opioid addiction brain regions. Infusions of antalarmin (a CRF1 receptor antagonist) into the ventral tegmental area (VTA) dose dependently reduced heroin self-administration in rats but had no impact on food reinforcement or locomotor activity in rats. Using RNAscope in situ hybridization, we found that heroin, but not saline, self-administration upregulated CRF1 receptor mRNA in the VTA, particularly on dopamine neurons. AMPA GluR1 and dopamine reuptake transporter mRNA in VTA neurons were not affected by heroin. The western-blot assay showed that CRF1 receptors were upregulated in the VTA and nucleus accumbens. No significant changes in CRF1 protein expression were detected in the prefrontal cortex, insula, dorsal hippocampus, and substantia nigra. In addition, we found that 15 days of environmental enrichment implemented after heroin self-administration does not reverse upregulation of VTA CRF1 receptor mRNA but it downregulates dopamine transporter mRNA. Overall, these data suggest that heroin self-administration requires stimulation of VTA CRF1 receptors and upregulates their expression in brain regions involved in reinforcement. Such long-lasting neuroadaptations may contribute to continuation of drug use and relapse due to stress exposure and are not easily reversed by EE exposure.

The effects of neuropeptide W on food consumption and feeding behavior in neonatal meat-type chicks: Role of CRF1/CRF2 and NPY1 receptors

In several studies, the regulatory role of the neuropeptide W (NPW) system in food intake has been demonstrated. Considering the lack of avian studies in this field, the current research was conducted to evaluate the effects of intracerebroventricular (ICV) infusion of NPW and its interferences with corticotropin, melanocortin, and neuropeptide Y (NPY) receptors on meal consumption and feeding behaviors of broilers. In the first experiment, birds were injected with NPW (0.75, 1.5, and 3 nmol) in addition to saline. In the second experiment, saline, CRF1 receptor antagonist (NBI35965, 30 μg), NPW (3 nmol), and simultaneous injections of NBI35965 and NPW were performed. Experiments 3–8 were identical to experiment 2, except that CRF2 receptor antagonist (K41498, 30 μg), MC3/MC4 receptor antagonist (SHU9119, 0.5 nmol), MC4 receptor antagonist (HS024, 0.5 nmol), NPY1 receptor antagonist (BMS193885, 1.25 nmol), NPY2 receptor antagonist (CYM9484, 1.25 nmol), and NPY5 receptor (antagonist L-152,804, 1.25 nmol) were administrated instead of NBI35965. After that, cumulative feed intake and feeding behavior were monitored for 2 h and 30 min after injections, respectively. Following the infusion of NPW (1.5 and 3 nmol), there was a significant stimulation of meal consumption in chickens (P < 0.05). Concomitant injection of NBI35965 and K41498 with NPW enhanced the appetite-increasing effect of NPW (P < 0.05); while BMS193885 suppressed this effect of NPW (P < 0.05). Injection of SHU9119, HS024, CYM9484, and L-152804 with NPW at the same time, had no significant effect on NPW-induced hyperphagia (P > 0.05). NPW also significantly decreased the standing period and the number of jumps, steps, and exploratory pecks, and led to an increase in sitting period and feeding pecks (P < 0.05). Based on the observations, it seems that NPW-induced hyperphagia could be mediated through CRF1, CRF2, and NPY1 receptors in neonatal broilers.

Interactions between corticotropin releasing factor signaling and prophylactic antibiotics on measures of intestinal function in weaned and transported pigs.

The study objective was to evaluate the interaction between corticotrophin releasing factor (CRF) receptor signaling and prophylactic antibiotic administration on intestinal physiology in newly weaned and transported pigs. Pigs (n = 56; 5.70 ± 1.05kg) were weaned (20.49 ± 0.64d), a blood sample was taken, and then pigs were given an intraperitoneal injection of saline (SAL; n = 28 pigs) or a CRF receptor antagonist (CRFA; n = 28 pigs; 30μg/kg body weight; Astressin B), and then were transported in a livestock trailer for 12 h and 49min. A second and third intraperitoneal injection was given at 4h 42 min and 11h 36min into the transport process, respectively. Following transport, 4 SAL and 4 CRFA pigs were blood sampled and euthanized. The remaining 48 pigs were individually housed and given dietary antibiotics [AB; n = 12 SAL and 12 CRFA pigs; chlortetracycline (441ppm) + tiamulin (38.6ppm)] or no dietary antibiotics (NAB; n = 12 SAL and 12 CRFA pigs) for 14d post-transport. Blood was collected at 12h and on d 3, 7, and 14, and then pigs were euthanized on d 7 (n = 24) and d 14 (n = 24) post-weaning and transport. Circulating cortisol was reduced (p = 0.05) in CRFA pigs when compared to SAL pigs post-weaning and transport. On d 7, jejunal villus height and crypt depth was greater overall (p < 0.05) in AB-fed pigs versus NAB-fed pigs. On d 14, ileal crypt depth was reduced (p = 0.02) in CRFA pigs when compared to SAL pigs. Jejunal CRF mRNA abundance tended to be reduced (p = 0.09) on d 7 in CRFA pigs versus SAL pigs. On d 14, jejunal tumor necrosis factor-alpha was reduced (p = 0.01) in AB-fed pigs versus NAB-fed pigs. On d 7, change in glucose short-circuit current tended to be increased (p = 0.07) in CRFA pigs fed the AB diet when compared to CRFA pigs fed the NAB diet. In conclusion, CRFA pigs and pigs fed AB had some similar biological intestinal function measures post-weaning and transport.

Crinecerfont, a CRF1 receptor antagonist, lowers adrenal androgens in adolescents with congenital adrenal hyperplasia

Crinecerfont, a CRF1 receptor antagonist, lowers adrenal androgens in adolescents with congenital adrenal hyperplasia

A Brain Region-Dependent Alteration in the Expression of Vasopressin, Corticotropin-Releasing Factor, and Their Receptors Might Be in the Background of Kisspeptin-13-Induced Hypothalamic-Pituitary-Adrenal Axis Activation and Anxiety in Rats.

Previously, we reported that intracerebroventricularly administered kisspeptin-13 (KP-13) induces anxiety-like behavior and activates the hypothalamic-pituitary-adrenal (HPA) axis in rats. In the present study, we aimed to shed light on the mediation of KP-13's stress-evoking actions. The relative gene expressions of the corticotropin-releasing factor (Crf, Crfr1, and Crfr2) and arginine vasopressin (Avp, Avpr1a, and Avpr1b) systems were measured in the amygdala and hippocampus of male Wistar rats after icv KP-13 treatment. CRF and AVP protein content were also determined. A different set of animals received CRF or V1 receptor antagonist pretreatment before the KP-13 challenge, after which either an open-field test or plasma corticosterone levels measurement was performed. In the amygdala, KP-13 induced an upregulation of Avp and Avpr1b expression, and a downregulation of Crf. In the hippocampus, the mRNA level of Crf increased and the level of Avpr1a decreased. A significant rise in AVP protein content was also detected in the amygdala. KP-13 also evoked anxiety-like behavior in the open field test, which the V1 receptor blocker antagonized. Both CRF and V1 receptor blockers reduced the KP-13-evoked rise in the plasma corticosterone level. This suggests that KP-13 alters the AVP and CRF signaling and that might be responsible for its effect on the HPA axis and anxiety-like behavior.

Visceral hypersensitivity induced by mild traumatic brain injury via the corticotropin-releasing hormone receptor: An animal model.

Mild blast-induced traumatic brain injury (bTBI) induces various gut symptoms resembling human irritable bowel syndrome (IBS) as one of mental and behavioral disorders. However, the underlying mechanisms remain unclear. We investigated whether the extremely localized brain impact extracranially induced by laser-induced shock wave (LISW) evoked IBS-like phenomenon including visceral hypersensitivity and intestinal hyperpermeability in rats. The rats were subjected to LISW on the scalp to shock the entire brain. Visceral hypersensitivity was evaluated by the threshold pressure of abdominal withdrawal reflex (AWR) using a colorectal distension test. Permeability was evaluated by the concentration of penetrating FITC-dextran from intestine and the mRNA expression levels of tight junction family proteins. Involvement of corticotropin-releasing factor receptor (CRFR) 1 and 2 was examined by evaluating mRNA expression and modulating CRFR function with agonist, recombinant CRF (10 μg/kg), and antagonist, astressin (33 μg/kg). High-throughput sequencing of the gut microbiota was performed by MiSeqIII instrument and QIIME tool. The thresholds of the AWR were significantly lowered after LISW. Permeability was increased in small intestine by LISW along with decreased expression of tight junction ZO-1. LISW significantly increased CRFR1 expression and decreased CRFR2 expression. Visceral hypersensitivity was significantly aggravated by CRFR agonist and suppressed by CRFR antagonist. The α- and β-diversity of the fecal microbiota was altered after LISW. LISW provoked visceral hypersensitivity, small intestinal hyperpermeability, altered expression of CRFRs and changes in the microbiota, suggesting that genuine bTBI caused by LISW can induce a pathophysiology comparable to that of human IBS.

Hypophagia induced by intracerebroventricular injection of apelin-13 is mediated via CRF1/CRF2 and MC3/MC4 receptors in neonatal broiler chicken

Previous studies have shown the role of apelin and its receptors in the regulation of food intake. In the present study, we investigate the mediating role of melanocortin, corticotropin, and neuropeptide Y systems in apelin-13- induced food intake in broilers. Eight trials were run in the current investigation to ascertain the relationships between the aforementioned systems and apelin-13 on food intake and behavioral changes after apelin-13 administration. In experiment 1, hens were given an intracerebroventricular administration of a solution for control in addition to apelin-13 (0.25, 0.5, and 1 µg). Astressin-B (a CRF1/CRF2 receptor antagonist, 30 µg), apelin-13 (1 µg), and administration of astressin-B and apelin-13 concurrently, were all injected into the birds in experiment 2. Experiments 3 through 8 were quite similar to experiment 2, with the exception of astressin2-B (CRF2 receptor antagonist, 30 µg), SHU9119 (MC3/MC4 receptor antagonist, 0.5 nmol), MCL0020 (MC4 receptor antagonist, 0.5 nmol), BIBP-3226 (NPY1 receptor antagonist, 1.25 nmol), BIIE 0246 (NPY2 receptor antagonist, 1.25 nmol), and CGP71683A (NPY5 receptor antagonist, 1.25 nmol) were injected instead of astressin-B. After then, total food consumption was monitored for 6 h. Apelin-13 injections of 0.5 and 1 µg decreased feeding (P < 0.05). The hypophagic effects of apelin were attenuated following the simultaneous administration of Astressin-B and Astressin2-B with apelin-13 (P > 0.05). Co-infusion of SHU9119 and apelin-13 reduced the appetite-decreasing effects of apelin-13 (P > 0.05). When MCL0020 and apelin-13 were injected at the same time, the hypophagia that apelin-13 induced was eliminated (P > 0.05). BIBP-3226, BIIE 0246, and CGP71683A had no effect on the hypophagia brought on by apelin-13 (P > 0.05). Also, apelin-13 significantly increased number of steps, jumps, exploratory food, pecks and standing time while decreased siting time (P < 0.05). These findings suggest that apelin-13-induced hypophagia in hens may involve the CRF1/CRF2 and MC3/MC4 receptors.

Crinecerfont, a CRF1 Receptor Antagonist, Lowers Adrenal Androgens in Adolescents With Congenital Adrenal Hyperplasia.

Crinecerfont, a corticotropin-releasing factor type 1 receptor antagonist, has been shown to reduce elevated adrenal androgens and precursors in adults with congenital adrenal hyperplasia (CAH) due to 21-hydroxylase deficiency (21OHD), a rare autosomal recessive disorder characterized by cortisol deficiency and androgen excess due to elevated adrenocorticotropin. To evaluate the safety, tolerability, and efficacy of crinecerfont in adolescents with 21OHD CAH. This was an open-label, phase 2 study (NCT04045145) at 4 centers in the United States. Participants were males and females, 14 to 17 years of age, with classic 21OHD CAH. Crinecerfont was administered orally (50 mg twice daily) for 14 consecutive days with morning and evening meals. The main outcomes were change from baseline to day 14 in circulating concentrations of ACTH, 17-hydroxyprogesterone (17OHP), androstenedione, and testosterone. 8 participants (3 males, 5 females) were enrolled; median age was 15 years and 88% were Caucasian/White. After 14 days of crinecerfont, median percent reductions from baseline to day 14 were as follows: ACTH, -57%; 17OHP, -69%; and androstenedione, -58%. In female participants, 60% (3/5) had ≥50% reduction from baseline in testosterone. Adolescents with classic 21OHD CAH had substantial reductions in adrenal androgens and androgen precursors after 14 days of oral crinecerfont administration. These results are consistent with a study of crinecerfont in adults with classic 21OHD CAH.

Ocimum tenuiflorum extract (HOLIXERTM): Possible effects on hypothalamic-pituitary-adrenal (HPA) axis in modulating stress.

Ocimum tenuiflorum is a sacred medicinal plant bestowed with multiple health benefits. This plant is traditionally considered an adaptogen. Many scientific studies have indicated the anti-stress potential of Ocimum tenuiflorum but with higher doses. The present study investigated the effects of HolixerTM (a clinically studied standardized Ocimum tenuiflorum extract) on modulating stress using two in vivo models, namely the swim endurance study in mice and forced swim test in rats. In addition, we explored the mechanism of action of HolixerTM on the HPA axis using two in vitro cell-based assays to check for its inhibitory effect on cortisol release and CRF1 receptor antagonistic activity. Ocimum tenuiflorum extract enhanced the swimming time in mice, reduced the stress-induced increase in immobility time, and prevented the increase in corticosterone in rats subjected to the forced swim test. Further, Ocimum tenuiflorum extract inhibited cortisol release and exhibited a significant CRF1 receptor antagonist activity. Thus, Ocimum tenuiflorum extract was found effective in managing stress, and the effect could be due to the inhibition of cortisol release and the antagonistic effect on the CRF1 receptors.

Comparisons of stress-related neuronal activation induced by restraint in adult male rat offspring with prenatal exposure to buprenorphine, methadone, or morphine.

Prenatal opioid exposure may impede the development of adaptive responses to environmental stimuli by altering the stress-sensitive brain circuitry located at the paraventricular nucleus of the hypothalamus (PVH) and locus coeruleus (LC). Corticotropin-releasing factor (CRF) released from neurons in the PVH has emerged as a key molecule to initiate and integrate the stress response. Methadone (Meth) and buprenorphine (Bu) are two major types of synthetic opioid agonists for first-line medication-assisted treatment of opioid (e.g., morphine, Mor) use disorder in pregnant women. No studies have compared the detrimental effects of prenatal exposure to Meth versus Bu on the stress response of their offspring upon reaching adulthood. In this study, we aimed to compare stress-related neuronal activation in the PVH and LC induced by restraint (RST) stress in adult male rat offspring with prenatal exposure to the vehicle (Veh), Bu, Meth, or Mor. CFos-immunoreactive cells were used as an indicator for neuronal activation. We found that RST induced less neuronal activation in the Meth or Mor exposure groups compared with that in the Bu or Veh groups; no significant difference was detected between the Bu and Veh exposure groups. RST-induced neuronal activation was completely prevented by central administration of a CRF receptor antagonist (α-helical CRF9-41, 10 μg/3 μL) in all exposure groups, suggesting the crucial role of CRF in this stress response. In offspring without RST, central administration of CRF (0.5 μg/3 μL)-induced neuronal activation in the PVH and LC. CRF-induced neuronal activation was lessened in the Meth or Mor exposure groups compared with that in the Bu or Veh groups; no significant difference was detected between the Bu and Veh exposure groups. Moreover, RST- or CRF-induced neuronal activation in the Meth exposure group was comparable with that in the Mor exposure group. Further immunohistochemical analysis revealed that the Meth and Mor exposure groups displayed less CRF neurons in the PVH of offspring with or without RST compared with the Bu or Veh groups. Thus, stress-induced neuronal activation in the PVH and LC was well preserved in adult male rat offspring with prenatal exposure to Bu, but it was substantially lessened in those with prenatal exposure to Meth or Mor. Lowered neuronal activation found in the Meth or Mor exposure groups may be, at least in part, due to the reduction in the density of CRF neurons in the PVH.

Anxiolytic reversal of classically conditioned / chronic stress-induced gene expression and learning in the Stress Alternatives Model

Social decision-making is critically influenced by neurocircuitries that regulate stress responsiveness. Adaptive choices, therefore, are altered by stress-related neuromodulatory peptide systems, such as corticotropin releasing factor (CRF). Experimental designs that take advantage of ecologically salient fear-inducing stimuli allow for revelation of neural mechanisms that regulate the balance between pro- and anti-stress responsiveness. To accomplish this, we developed a social stress and conditioning protocol, the Stress Alternatives Model (SAM), that utilizes a simple dichotomous choice, and produces distinctive behavioral phenotypes (Escape or Stay). The experiments involve repeated social aggression, a potent unconditioned stimulus (US), from a novel larger conspecific (a 3X larger Rainbow trout). Prior to the social interaction, the smaller test fish is presented with an auditory conditioning stimulus (water off = CS). During the social aggression, an escape route is available, but is only large enough for the smaller test animal. Surprisingly, although the new aggressor provides vigorous attacks each day, only 50% of the test fish choose Escape. Stay fish, treated with the CRF1 antagonist antalarmin, a potent anxiolytic drug, on day 4, promotes Escape behavior for the last 4 days of the SAM protocol. The results suggest that the decision to Escape, required a reduction in stress reactivity. The Stay fish that chose Escape following anxiolytic treatment, learned how to use the escape route prior to stress reduction, as the Escape latency in these fish was significantly faster than first time escapers. In Escape fish, the use of the escape route is learned over several days, reducing the Escape latency over time in the SAM. Fear conditioning (water off + aggression) resulted in elevated hippocampal (DL) Bdnf mRNA levels, with coincident reduction in the AMPA receptor subunit Glua1 expression, a result that is reversed following a one-time treatment (during SAM aggression on day 4) with the anxiolytic CRF1 receptor antagonist antalarmin.

Corticotropin releasing factor neurons in the visual cortex mediate long-term changes in visual function induced by early adversity

Early adversity can cause malfunction of the visual system in adulthood. Adult female but not male mice undergoing early chronic mild stress (ECMS) maintain ocular dominance (OD) plasticity after the critical period. How early stressful experiences have a long-term impact on it is largely unknown. Here, we observed a wide distribution of corticotropin-releasing factor (CRF)-positive neurons, which mainly colocalized with a subpopulation of GABAergic interneurons in the mouse primary visual cortex (V1). Optogenetic activation of CRF-positive neurons transfected with AAV-ChR2 evoked inhibitory currents in nearby pyramidal cells. ECMS induced a reduction in the expression of CRF mRNA in adult mouse V1. Chemogenetic activation of V1 CRF neurons impaired OD plasticity in adult ECMS females. We further showed that local administration of the corticotropin releasing factor receptor 1 (CRFR1) antagonist via an osmotic minipump into the visual cortex mimicked OD plasticity in adult ECMS females. Whole-cell recording in layer 2/3 pyramidal neurons revealed that the CRFR1 antagonist reduced the short-term depression (STD) of evoked inhibitory postsynaptic current (IPSC) in females but not in males. Likewise, CRF agonists have the opposite effect. In summary, our findings indicate that the local CRF-CRFR1 system within V1 may mediate the long-term and sex-dependent effect of early stress experiences on visual plasticity and provide a target for the prevention of visual deficits in adults with a history of early-life adversity.

Corticotropin-releasing factor is involved in acute stress-induced analgesia and antipruritus.

Under the condition of stress, the hypothalamic-pituitary-adrenal axis (HPA axis) is activated and causes the secretion of corticotropin-releasing factor (CRF). Previous studies have demonstrated that CRF is involved in the regulation of pain and itch. Thus, it remains worthy to explore whether the desensitization of pain and itch under high-intensity acute stress (such as high fear and tension) is related to the sharp increase of CRF. Forced swimming was used to simulate acute stress. ELISA and pharmacological methods were conducted to observe the effects of forced swimming on acute pain or itch and the relationship between blood CRF content and itch or pain behavior. Intracerebroventricular (ICV) administration of CRF was conducted to examine the effects of CRF on acute pain or itch. Intrathecal administration of CRF receptor agonist or antagonist was conducted to examine the receptor mechanisms of the regulatory role of CRF in pain and itch. ELISA experiment showed that the serum CRF in mice reached its peak within 5-10 min after acute stress (forced swimming). Behavioral data showed that the scratching behavior induced by itch agents decreased after acute swimming, while the mechanical pain threshold increased significantly. The inhibitory effect of acute stress on pain and itch is mediated by CRF receptor2 (CRFR2). Then, ICV injection of CRF was used to simulate the massive release of CRF under acute stress, and we observed that the scratching behavior induced by histamine or chloroquine was significantly inhibited after ICV injection of CRF. The above effects of CRF are mainly mediated by CRFR2. These results suggest that 5-10min after acute stress, a large amount of CRF is released into the blood from the hypothalamus, which significantly inhibits acute pain and itch by acting on CRFR2. ICV injection of CRF can replicate the antipruritus effects of acute stress. The present study investigated the mechanism of acute stress-induced analgesia and antipruritus and provided theoretical support for the treatment of pain and itch.

Corticotropin releasing factor and drug seeking in substance use disorders: Preclinical evidence and translational limitations

The neuropeptide, corticotropin releasing factor (CRF), has been an enigmatic target for the development of medications aimed at treating stress-related disorders. Despite a large body of evidence from preclinical studies in rodents demonstrating that CRF receptor antagonists prevent stressor-induced drug seeking, medications targeting the CRF-R1 have failed in clinical trials. Here, we provide an overview of the abundant findings from preclinical rodent studies suggesting that CRF signaling is involved in stressor-induced relapse. The scientific literature that has defined the receptors, mechanisms and neurocircuits through which CRF contributes to stressor-induced reinstatement of drug seeking following self-administration and conditioned place preference in rodents is reviewed. Evidence that CRF signaling is recruited with repeated drug use in a manner that heightens susceptibility to stressor-induced drug seeking in rodents is presented. Factors that may determine the influence of CRF signaling in substance use disorders, including developmental windows, biological sex, and genetics are examined. Finally, we discuss the translational failure of medications targeting CRF signaling as interventions for substance use disorders and other stress-related conditions. We conclude that new perspectives and research directions are needed to unravel the mysterious role of CRF in substance use disorders.

Peripheral CRF-R1/CRF-R2 antagonist, astressin C, induces a long-lasting blockade of acute stress-related visceral pain in male and female rats

Peptide CRF antagonists injected peripherally alleviate stress-induced visceral hypersensitivity (SIVH) to colorectal distension (CRD) in rodents. Here we further evaluated the dose and time-dependent inhibitory activity of several long-acting peptide CRF receptor antagonists related to astressin on SIVH, focusing on astressin C (AstC), which previously showed high efficacy on stress-related alterations of HPA axis and gut secretomotor functions. Male and female Sprague-Dawley rats pretreated subcutaneously (SC) with AstC were injected intraperitoneally (IP) with CRF 15 min later. The visceromotor responses (VMR) to graded phasic CRD (10, 20, 40 and 60 mmHg) were monitored at basal, 15 min and up to 1–8 days after pretreatment. Two other astressin analogs, hexanoyl-astressin D (Hex-AstD) and [CαMeVal19,32]-AstC, were also tested. The response to IP CRF was sex-dependent with female rats requiring a higher dose to exhibit visceral hyperalgesia. Pretreatment with AstC (30–1000 µg/kg) resulted in a dose-related inhibition of IP CRF-induced SIVH and diarrhea in both sexes. The highest dose prevented SIVH and diarrhea up to 5–7 days after a single SC injection and was lost on day 7 (females) and day 8 (males) but reinstated after a second injection of AstC on day 8 or 9 respectively. [CαMeVal19,32]-AstC and Hex-AstD (1000 µg/kg in males) also prevented SIVH. These data show the potent long-lasting anti-hyperalgesic effect of AstC in an acute model of SIVH in both male and female rats. This highlights the potential of long-acting peripheral CRF antagonists to treat stress-sensitive irritable bowel syndrome.

Estradiol Promotes Social Stress Susceptibility in Female Rats in a Region‐ and CRF‐Specific Manner

Repeated exposure to social stress is a common risk factor for anxiety disorders. Women are twice as likely as men to suffer from anxiety. However, the mechanisms behind this increased risk remain unclear, which limits targeted therapeutic options. Interestingly, this increased susceptibility in females is confined between the onset of puberty and the end of menopause, suggesting that ovarian hormones may contribute to this disproportionate risk of stress‐related disorders. We have previously shown that witness stress (WS), a model of social stress in which a female rat is subjected to witnessing an aggressive social defeat encounter between a male intruder and a novel male resident, produces lasting anxiety‐like behaviors selectively in intact, cycling female rats. Ovariectomized (OVX) females are largely resistant to WS‐evoked behaviors, but estradiol replacement reinstates these anxiety‐like behaviors. Moreover, intact females, but not OVX, exposed to WS exhibit a distinct and long‐lasting increase in corticotropin releasing factor (CRF) expression in the central amygdala (CeA), a stress sensitive brain region. The current study seeks to understand specific 17‐β estradiol (17‐βE)‐induced neural regulation that may contribute to the heightened susceptibility to social stress among females to identify a druggable target to promote stress resilience. Following recovery from OVX or sham surgery, adult female rats were treated with either 17‐βE (10μg/rat, s.c.) or vehicle (0μg/rat, s.c.) and one hour later, were exposed to WS or control handling. WS/control episodes were video recorded to measure burying, freezing, and rearing behaviors, and blood samples were collected one hour after WS/control to assess corticosterone (CORT) levels. Brains were collected two hours after WS/control. To quantify activity of CRF neurons, immunohistochemistry was used to double label for CRF and cFos in various stress sensitive regions. Behavioral responses were shown to be regulated by 17‐βE; WS‐evoked burying evident in intact females was decreased by OVX and reinstated by 17‐βE. However, there was no effect of WS or 17‐βE on freezing or rearing behaviors. Additionally, 17‐βE increased WS‐induced plasma CORT levels. cFos positive CRF expression in the CeA was robustly enhanced by 17‐βE during WS, but not control. This effect was not globally observed as there was no effect of WS or 17‐βE on cFos expression in CRF positive neurons within the paraventricular nucleus of the hypothalamus and the hippocampus. Analysis of stress‐ and CRF‐sensitive brain regions including the locus coeruleus are ongoing. Taken together, these findings suggest that an interaction between 17‐βE and CRF is paramount in facilitating the anxiety‐like responses to WS. Given the controversial impact of CRF1 receptor antagonists to date, this study provides evidence of conditions under which anxiety‐like behaviors are associated with enhanced CRF responses and may serve as a personalized viable target to treat increased sensitivity to stress‐related disorders in females.