Peripheral Corticotropin-releasing Factor Research Articles

Neuroendocrine Control of the Gut During Stress: Corticotropin-Releasing Factor Signaling Pathways in the Spotlight

Stress affects the gastrointestinal tract as part of the visceral response. Various stressors induce similar profiles of gut motor function alterations, including inhibition of gastric emptying, stimulation of colonic propulsive motility, and hypersensitivity to colorectal distension. In recent years, substantial progress has been made in our understanding of the underlying mechanisms of stress's impact on gut function. Activation of corticotropin-releasing factor (CRF) signaling pathways mediates both the inhibition of upper gastrointestinal (GI) and the stimulation of lower GI motor function through interaction with different CRF receptor subtypes. Here, we review how various stressors affect the gut, with special emphasis on the central and peripheral CRF signaling systems.

Immediate and Sustained Blood Pressure Lowering by Urocortin 2

Recently, novel corticotropin-releasing factor-related peptides, named urocortin 1, 2, and 3, and a distinct cardiac and peripheral vascular receptor (corticotropin-releasing factor receptor 2) were described being part of a peripheral corticotropin-releasing factor system modulating cardiovascular function in response to stress. Vasorelaxation and blood pressure lowering have been reported after acute administration of these peptides. No data are available on the acute and chronic effects of urocortin 2 on blood pressure in models of arterial hypertension. To test these effects, hypertensive salt-sensitive and normotensive salt-resistant Dahl rats were randomly assigned to twice-daily applications of urocortin 2 or vehicle for 5 weeks. Blood pressure, heart rate, and left ventricular dimension and function were recorded at baseline, after initial application, and, together with cardiac and aortic expression of urocortin 2 and its receptor, after 5 weeks of treatment. Urocortin 2 significantly reduced blood pressure in hypertensive rats without affecting heart rate. Long-term urocortin 2 treatment in hypertensive rats induced sustained blood pressure reduction and diminished the development of hypertension-induced left ventricular hypertrophy and the deterioration of left ventricular contractile function. Corticotropin-releasing factor receptor 2 expression was preserved despite chronic stimulation by urocortin 2. In conclusion, our study shows that, in an animal model of arterial hypertension, urocortin 2 has immediate and sustained blood pressure-lowering effects. Beneficial effects on blood pressure, left ventricular dimension, and function, together with preserved receptor expression, suggest that corticotropin-releasing factor receptor 2 stimulation by urocortin 2 may represent a novel approach to the treatment of arterial hypertension.

Abstract 4394: Chronic Administration of Urocortin 2 Prevents the Development of Heart Failure in an Animal Model of Hypertensive Heart Disease

Background: Recently, novel corticotropin-releasing factor (CRF)-related peptides named Urocortin (Ucn) 1, 2, and 3 were described. Available data suggest that the Ucns are part of a peripheral CRF system modulating cardiovascular function and mediating cardiovascular responses to stress. Chronic Ucn2 administration induced sustained blood pressure lowering and prevented the development of left ventricular hypertrophy (LVH) in an animal model of hypertensive heart disease. However, no data are available whether chronic administration of Ucn2 may prevent the progression from LVH to heart failure. Methods: Experiments were performed in Dahl salt-sensitive rats. Animals were fed a high salt diet containing 4% NaCl to induce arterial hypertension, LVH, and heart failure. From the phase of LVH on (after 7 weeks of high salt diet), animals were injected with either Ucn2 at a dose of 2.5 μg/kg body weight or vehicle b.i.d. Animals underwent repetitive tail cuff blood pressure measurements and echocardiographic analysis of LV dimension and function at baseline (prior to first injection of Ucn2) and after 5 weeks of b.i.d. treatment with Ucn2. Results: Results are given in the following table as mean (± SD). No differences in heart weight/body weight ratios between Ucn2- and vehicle treated animals were found after 5 weeks of treatment. Conclusion: Chronic CRF receptor stimulation by Ucn2 in the severely hypertensive Dahl salt-sensitive rat, an animal model of hypertensive heart disease, prevents the progression from LVH to LV dilatation and the deterioration of LV function. Thus, chronic administration of Ucn2 might represent a novel approach to the prevention of heart failure.

PERIPHERAL PLASMA CORTICOTROPIN‐RELEASING FACTOR CONCENTRATION DOES NOT CORRELATE WITH AUGMENTED COLONIC MOTILITY IN RESPONSE TO RESTRAINT STRESS IN RATS

1. Corticotropin-releasing factor (CRF) alters colonic motor function in response to restraint stress. It remains unclear whether peripheral CRF released by restraint stress correlates with stress-mediated colonic motility. 2. One strain gauge transducer was sutured on the serosal surface of the mid colon and a silastic cannula was inserted into the jugular vein. Seven days after surgery, the wires from the transducers were connected to the recording system and colonic motility was recorded. Rats were subjected to restraint stress for 90 min and blood samples were obtained every 30 min. 3. Immediately after the start of stress loading, phasic contractions were observed that lasted during restraint stress loading. Intracisternal injection of CRF elicited rapid augmentation of colonic motility, similar to that of restraint stress. 4. The plasma CRF concentration (4 pg/mL) remained unaltered within the first 30 min after stress loading, but increased gradually to 23 pg/mL at 90 min after the beginning of restraint stress. There was no positive correlation observed between plasma CRF concentrations and restraint stress-stimulated colonic motility. 5. These findings indicate that central CRF has a predominant role in stress-induced augmentation of colonic motility. It remains unclear whether a delayed response of plasma CRF concentrations in response to restraint stress is involved in mediating colonic motility.

Peripheral corticotropin releasing factor (CRF) and a novel CRF1 receptor agonist, stressin1‐A activate CRF1 receptor expressing cholinergic and nitrergic myenteric neurons selectively in the colon of conscious rats

Intraperitoneal (i.p.) corticotropin releasing factor (CRF) induced a CRF(1) receptor-dependent stimulation of myenteric neurons and motility in the rat proximal colon. We characterize the colonic enteric nervous system response to CRF in conscious rats. Laser capture microdissection combined with reverse transcriptase polymerase chain reaction (RT-PCR) and immunohistochemistry in longitudinal muscle myenteric plexus whole-mount colonic preparations revealed CRF(1) receptor expression in myenteric neurons. CRF (i.p., 10 microg kg(-1)) induced Fos immunoreactivity (IR) (cells per ganglion) selectively in myenteric plexus of proximal (18.3 +/- 2.4 vs vehicle: 0.0 +/- 0.0) and distal colon (16.8 +/- 1.2 vs vehicle: 0.0 +/- 0.0), but not in that of gastric corpus, antrum, duodenum, jejunum and ileum. The selective CRF(1) agonist, stressin(1)-A (i.p., 10 microg kg(-1)) also induced Fos IR in myenteric but not in submucosal plexus of the proximal and distal colon. Fos IR induced by CRF was located in 55 +/- 1.9% and 53 +/- 5.1% of CRF(1) receptor-IR myenteric neurons and in 44 +/- 2.8% and 40 +/- 3.9% of cholinergic neurons with Dogiel type I morphology, and in 20 +/- 1.6% and 80 +/- 3.3% of nitrergic neurons in proximal and distal colon respectively. CRF and stressin(1)-A elicit defecation and diarrhoea. These data support that one mechanism through which peripherally injected CRF ligands stimulate colonic function involves a direct action on colonic cholinergic and nitrergic myenteric neurons expressing CRF(1) receptor.

Abstract 670: Immediate And Sustained Blood Pressure Lowering by CRF-receptor Stimulation: A Novel Approach To Antihypertensive Therapy?

Background: Recently, novel corticotropin-releasing factor (CRF)-related peptides, named urocortin I (UcnI), UcnII, and UcnIII were described. Available data suggest that the Ucns are part of a peripheral CRF system modulating cardiovascular function and mediating cardiovascular responses to stress. Blood pressure (BP) lowering effects have been described after administration of UcnI. However, no data are available on effects of UcnII on BP in an animal model of systemic arterial hypertension. Methods: Experiments were performed in Dahl salt-sensitive (DSS) and salt-resistant rats (DSR, control). Animals were fed a diet containing 4% NaCl (high salt) to induce arterial hypertension in DSS rats. At the end of week 2 of high salt diet, both DSS and DSR rats were randomly assigned to i.p. injections of either UcnII (2.5 μg/kg body weight) or vehicle b.i.d. for five weeks. Animals underwent repetitive tail cuff BP measurements at baseline (prior to first injection), at 5 and 15 minutes after the first injection and at week 1, 2, and 5 of b.i.d. treatment. At week 5 animals were sacrificed to determine heart weight /body weight ratio. Results: Systolic BP (SBP, mmHg) and heart rate (HR, min −1 ) are given in the following table as mean ± SD (n=10 per group). Conclusions: In hypertensive DSS rats, acute CRF-receptor stimulation by UcnII immediately lowered BP to the range observed in DSR rats. Compared to vehicle-treated DSS rats, sustained BP reduction was observed with further chronic administration of UcnII. No severe reflex tachycardia was observed after administration of UcnII. Thus, CRF-receptor stimulation might represent a novel approach to the treatment of arterial hypertension.

Gastric Vagal Efferent Inhibition Evoked by Intravenous CRF Is Unrelated to Simultaneously Recorded Vagal Afferent Activity in Urethane-Anesthetized Rats

Corticotropin-releasing factor (CRF) injected peripherally or released in response to stressful challenges to the organism reduces gastric tone and contractility, in part by vagal pathways. However, information on the changes in gastric vagal impulse activity evoked by peripheral CRF administration is entirely lacking. Using a novel "dual recording" method in urethane-anesthetized rats, vagal efferent (VE) and afferent (VA) impulse activities were recorded simultaneously from separate, fine bundles dissected from the ventral gastric vagus nerve branch innervating the glandular stomach. Activity records for 38 VA single units (SUs) and 33 VE SUs were sorted from multiunit records obtained from 13 preparations. Intravenous (iv) administration of saline had no effect on multiunit VE activity, whereas CRF (1 microg/kg, iv) immediately inhibited VE activity, reaching a nadir of 54 +/- 8.0% of preinjection levels at 3.0 min postinjection. CRF (1 microg/kg, iv) inhibited 25/33 (75.8%) VE SUs and excited three of 33 (9.1%) VE SUs. In contrast to potent effects on VE activity, iv CRF did not alter multiunit VA activity. Single-unit analysis, however, revealed five of 38 (13.1%) VA SUs excited by iv CRF at widely varying latencies (suggesting an indirect mode of action) and one inhibited VA SU. VA SUs excited after iv CRF did not respond during gastric distention and vice versa. These experiments are the first to use simultaneous recording of gastric VA and VE units. The data demonstrate a predominantly inhibitory influence of iv CRF on VE outflow to the hindstomach, not driven by gastric vagovagal reflex activity.

CRF1 Receptors as a Therapeutic Target for Irritable Bowel Syndrome

The characterization of the corticotropin-releasing factor (CRF) family of neuroendocrine regulatory peptides, the cloning and pharmacological characterization of two CRF receptor subtypes (CRF(1) and CRF(2)), and the development of selective CRF receptor antagonists provided new insight to unravel the mechanisms of stress and the potential involvement of the CRF system in different pathophysiological conditions, including functional gastrointestinal disorders, mainly irritable bowel syndrome (IBS), and psychopathologies such as anxiety/depression. Compelling pre-clinical data showed that brain CRF administration mimics acute stress-induced colonic responses and enhances colorectal distension-induced visceral pain in rats through CRF(1) receptors. Similarly, peripheral CRF reduced the pain threshold to colonic distension and increased colonic motility in humans and rodents. These observations mimic the manifestations of IBS, characterized by abdominal bloating/discomfort and altered bowel habits. Moreover, CRF-CRF(1) pathways have been implicated in the development of anxiety/depression. These psychopathologies, together with stressful life events, have high comorbidity with IBS, and are considered significant components of the disease. From these observations, CRF(1) receptors have been suggested as a target to treat IBS. Peripherally acting CRF(1) antagonists might directly improve IBS symptoms, as related to motility, secretion and immune response. On the other hand, central actions will be beneficial as to prevent the psychopathologies that co-exist with IBS and as a way to modulate the central processing of stress- and visceral pain-related signals. Here, we review the pre-clinical and clinical data supporting these assumptions, and address the efforts done at a pharmaceutical level to develop effective therapies targeting CRF(1) receptors for functional gastrointestinal disorders.

Corticotropin‐releasing factor (CRF) receptor type 2 in the human stomach: Protective biological role by inhibition of apoptosis

Corticotropin-releasing factor agonists exert inhibitory effects in stomach functions possibly through peripheral routes. We have previously reported the expression of Urocortin (Ucn) I, an endogenous ligand of both CRF receptor types CRF(1) and CRF(2), in the human stomach. We examined CRF(1) and CRF(2) expression in the same tissue. Using RT-PCR, CRF(2) but not CRF(1) transcripts were detected in RNA extracts from normal human stomach. In addition, immunohistochemical analysis revealed receptor protein in epithelial gastric cells. In order to investigate the biological role of CRF(2) in these cells, an in vitro model was established, using the gastric cancer cell line AGS transiently transfected to express functional CRF(2). The effect of the CRF(2) endogenous ligands CRF, Ucns I and II on the growth parameters of the AGS/CRF(2) was examined. After 1 day of exposure, all three ligands reduced the degree of apoptosis (16%-19%, n = 9, P < 0.05) compared to non-treated controls and this effect was observed for 3 days of treatment. No such effect was detected in non-transfected cells, suggesting mediation through CRF(2) receptors. Administration of CRF, Ucns I and II had no effect on the proliferation rate of AGS/CRF(2) cells or on the release of PGE(2) by them. Our results demonstrate CRF(2) expression in the human gastric mucosa and indicate a physiological role of this receptor type in regulating apoptosis, an important parameter of gastric cell regeneration. Paracrine effects exerted by locally expressed endogenous ligands, such as Ucn I, support a significant role of the peripheral CRF system in gastric physiology. J. Cell. Physiol. 209: 905-911, 2006. (c) 2006 Wiley-Liss, Inc.

Peripherally administered CRF stimulates colonic motility via central CRF receptors and vagal pathways in conscious rats

Corticotropin releasing factor (CRF) is one of the most important factors in the mechanism of stress-induced stimulation of colonic motility. However, it is controversial whether stress-induced stimulation of colonic motility is mediated via central or peripheral CRF receptors. We investigated the hypothesis that peripherally injected CRF accelerates colonic motility through the central CRF receptor, but not the peripheral CRF receptor. A strain gauge transducer was sutured on the serosal surface of the proximal colon. Colonic motility was monitored before and after the peripheral injection of CRF. An in vitro muscle strip study was also performed to investigate the peripheral effects of CRF. Subcutaneous injection of CRF (30-100 microg/kg) stimulated colonic motility in a dose-dependent manner. The stimulatory effect of peripherally administered CRF on colonic motility was abolished by truncal vagotomy, hexamethonium, atropine, and intracisternal injection of astressin (a CRF receptor antagonist). No responses to CRF (10(-9) -10(-7) M) of the muscle strips of the proximal colon were observed. These results suggest that the stimulatory effect of colonic motility in response to peripheral administration of CRF is mediated by the vagus nerve, nicotinic receptors, muscarinic receptors, and CRF receptors of the brain stem. It is concluded that peripherally administered CRF reaches the area postrema and activates the dorsal nucleus of vagi via central CRF receptors, resulting in stimulation of the vagal efferent and cholinergic transmission of the proximal colon.

Role of peripheral corticotropin-releasing factor and urocortin II in intestinal inflammation and motility in terminal ileum

Corticotropin-releasing factor (CRF) and the closely related family of neuropeptides urocortins (Ucns) are ancient paracrine-signaling peptides secreted in both the central and peripheral neural circuits. CRF and Ucns released from the CNS (central) regulate a plethora of physiological processes that include food intake, inflammation, and bowel motility and permeability. In the gastrointestinal tract, CRF actions are largely proinflammatory, whereas the effects of the Ucn subtypes can be either pro- or antiinflammatory. Central (intracerebroventricular) or peripheral (i.p.) administration of CRF or Ucns inhibits gastric emptying and promotes colonic motility. To ascertain the role of peripherally expressed CRF and UcnII in gastrointestinal inflammation and motility, we generated ileum-specific phenotypic knockouts of these peptides by using RNA interference. Long dsRNA effectively silenced basal expression of CRF and UcnII in ileum. Control dsRNA or saline treatment did not affect CRF or UcnII expression. In an experimental model of toxin-induced intestinal inflammation, inhibition of CRF ablated the inflammatory response (measured by epithelial damage, mucosal edema, and neutrophil infiltration). UcnII dsRNA treatment did not alter the inflammatory response to toxin. Furthermore, ileal motility was increased after site-specific inhibition of both CRF and UcnII. Thus, we demonstrate that ileal-specific CRF promotes inflammation and both CRF and UcnII modulate bowel motility.

CRF1 receptor signaling pathways are involved in stress-related alterations of colonic function and viscerosensitivity: implications for irritable bowel syndrome.

1. The characterization of corticotropin releasing factor (CRF) and, more recently, the discovery of additional CRF-related ligands, urocortin 1, urocortin 2 and urocortin 3, the cloning of two distinct CRF receptor subtypes, 1 (CRF(1)) and 2 (CRF(2)), and the development of selective CRF receptor antagonists provided new insight to unravel the mechanisms of stress. Activation of brain CRF(1) receptor signaling pathways is implicated in stress-related endocrine response and the development of anxiety-like behaviors. 2. Compelling evidence in rodents showed also that both central and peripheral injection of CRF and urocortin 1 mimic acute stress-induced colonic response (stimulation of motility, transit, defecation, mucus and watery secretion, increased ionic permeability and occurrence of diarrhea) in rodents. Central CRF enhances colorectal distention-induced visceral pain in rats. Peripheral CRF reduced pain threshold to colonic distention and increased colonic motility in humans. 3. Nonselective CRF(1)/CRF(2) antagonists and selective CRF(1) antagonists inhibit exogenous (central or peripheral) CRF- and acute stress-induced activation of colonic myenteric neurons, stimulation of colonic motor function and visceral hyperalgesia while selective CRF(2) antagonists have no effect. None of the CRF antagonists influence basal or postprandial colonic function in nonstressed animals. 4. These findings implicate CRF(1) receptors in stress-related stimulation of colonic function and hypersensitivity to colorectal distention. Targeting CRF(1)-dependent pathways may have potential benefit against stress or anxiety-/depression-related functional bowel disorders.

Corticotropin-releasing factor and interleukin-1beta are involved in the electroacupuncture-induced analgesic effect on inflammatory pain elicited by carrageenan.

Electroacupuncture (EA) is used to relieve various kinds of pain. However, the mechanistic basis of electroacupuncture analgesia (EAA) in inflammatory pain remains unclear. In the present study, we investigated whether endogenous peripheral corticotropin-releasing factor (CRF) or interleukin-1beta (IL-1) participated in EAA during hyperalgesia elicited by carrageenan-induced inflammation. Carrageenan was subcutaneously administered by intraplantar (i.pl.) injection of the left hind paw to induce inflammation. Nociceptive thresholds were measured using the paw pressure threshold (PPT) (Randall Sellito Test). Rats received 3 Hz EA in the left anterior tibial muscles for 1 hour after carrageenan injection. The selective CRF antagonist, alpha-helical CRF, or the recombinant IL-1 receptor antagonist, IL-1ra, was administered by i.pl. injection of the inflamed paw or by intravenous (i.v.) injection 1 hour before EA. PPT decreased significantly 3 hours after carrageenan injection. This decrease persisted at least 24 hours after carrageenan injection. EA resulted in significant increases of PPT, moreover, PPT elevations lasted 24 hours after carrageenan injection. By contrast, PPT elevations produced by EA were dose-dependently antagonized by local i.pl. injection of alpha-helical CRF or IL-1ra. This PPT elevation was not influenced by i.v. injection of alpha-helical CRF or IL-1ra. These findings suggest that peripheral CRF or IL-1 participate in EAA during hyperalgesia. The release of CRF or IL-1 elicited by EA may trigger the release of opioid peptides within inflamed tissue which may activate peripheral opioid receptors and inhibit the pain.

Differential profile of CRF receptor distribution in the rat stomach and duodenum assessed by newly developed CRF receptor antibodies.

Peripheral corticotropin-releasing factor (CRF) receptor ligands inhibit gastric acid secretion and emptying while stimulating gastric mucosal blood flow in rats. Endogenous CRF ligands are expressed in the upper gastrointestinal (GI) tissues pointing to local expression of CRF receptors. We mapped the distribution of CRF receptor type 1 (CRF1) and 2 (CRF2) in the rat upper GI. Polyclonal antisera directed against the C-terminus of the CRF receptor protein were generated in rabbits and characterized by western blotting and immunofluorescence using CRF1- and CRF2-transfected cell lines and in primary cultured neurons from rat brain cortex. A selective anti-CRF1 antiserum (4467a-CRF1) was identified and used in parallel with another antiserum recognizing both CRF1 and CRF2 (4392a-CRF1&2) to immunostain gastric tissue sections. Antiserum 4467a-CRF1 demonstrated specific immunostaining in a narrow zone in the upper oxyntic gland within the stomach corpus. Conversely, 4392a-CRF1&2 labeled cells throughout the oxyntic gland and submucosal blood vessels. Pre-absorption with the specific antigen peptide blocked immunostaining in all experiments. Doublestaining showed co-localization of 4392a-CRF1&2 but not 4467a-CRF1 immunoreactivity with H/K-ATPase and somatostatin immunostaining in parietal and endocrine cells of the oxyntic gland. No specific staining was observed in the antrum with either antisera, whereas only antiserum 4392a-CRF1&2 showed modest immunoreactivity in the duodenal mucosa. Finally, co-localization of CRF2 and urocortin immunoreactivity was found in the gastric glands. These results indicate that both CRF receptor subtypes are expressed in the rat upper GI tissues with a distinct pattern and regional differences suggesting differential function.

Role of corticotropin-releasing factor receptors type 1 and 2 in modulating the rat adrenocorticotropin response to stressors.

We investigated the contribution of corticotropin-releasing factor (CRF) receptors types 1 and 2 (CRF(1) and CRF(2)) in mediating the ACTH response to shock, alcohol injection, or endotoxemia in the rat. Peptidic (Astressin B and Astressin(2)-B) and nonpeptidic (NBI 30775) CRF antagonists were injected iv before the stressors at doses previously shown to be effective in blocking the corresponding receptors. Because NBI 30775, which specifically blocks CRF(1), penetrates the brain following systemic injection, we also compared its effect with that of Astressin B, which primarily, though not exclusively, targets CRF(1) but does not cross the blood-brain barrier. Shocks, alcohol (4.5 g/kg, intragastrically) or lipopolysaccharide (LPS, 1 micro g/kg, iv) all significantly released ACTH. Astressin B or NBI 30775 markedly decreased the effect of shocks or alcohol and also interfered, though less significantly so, with the influence of LPS. In contrast, specific blockade of CRF(2) with Astressin(2)-B, although not significantly altering the overall ACTH response to shocks, alcohol, or LPS, slightly enhanced ACTH levels during the early phase of some of these responses. Interestingly, combined administration of NBI 30775 and Astressin(2)-B decreased ACTH levels more than NBI 30775 alone, although this difference did not reach statistical significance. Finally, blockade of CRF(1) and/or CRF(2) augmented LPS- induced TNF-alpha and IL-6 release. Collectively, there results confirm the critical role played by CRF(1) in mediating the ACTH response to shocks, alcohol and LPS, whereas the influence of CRF(2) remains subtle. Finally, we showed that peripheral endogenous CRF restrains the ability of LPS to release cytokines.

Intravenous urocortin II decreases blood pressure through CRF 2 receptor in rats

Urocortin II (Ucn II) is a new member of the corticotropin-releasing factor (CRF) family that binds selectively to the CRF subtype 2 receptor (CRF 2). CRF or urocortin injected intravenously (iv) induced hypotension. We investigated the influence of iv human Ucn II (hUcn II) on basal mean blood pressure (MAP) and on the sympathetic mediated hypertensive response to TRH analog, RX-77368 injected intracisternally (ic) 20 min after hUcn II in urethane-anesthetized rats. Ucn II (3, 10, and 30 μg/kg, iv) significantly decreased basal MAP from baseline by −20.9±6.5, −21.3±5.4 and −46.8±6.5 mm Hg, respectively, after 10 min. RX-77368 (30 ng, ic) elevated MAP for over 90 min with a maximal hypertensive response at 20 min. Ucn II (3, 10, and 30 μg/kg, iv) did not alter the 20 min net rise in MAP induced by RX-77368 (35.7±7.1, 32.6±3.3 and 24.6±6.9 mm Hg, respectively) compared with vehicle (33.6±4.3 mm Hg). The selective CRF 2 antagonist, astressin 2-B (60 μg/kg, iv) abolished hUcn II hypotensive action while having no effect on basal MAP. These data show that iv hUcn II induces hypotension through peripheral CRF 2 receptor while not altering the responsiveness to sympathetic nervous system-mediated rise in MAP.

Differential actions of peripheral corticotropin-releasing factor (CRF), urocortin II, and urocortin III on gastric emptying and colonic transit in mice: role of CRF receptor subtypes 1 and 2.

Peripheral CRF inhibits gastric emptying and stimulates colonic motor function in rats. We investigated the role of CRF(1) and CRF(2) receptors in i.p. CRF-induced alterations of gut transit in conscious mice using selective CRF(1) and CRF(2) ligands injected i.p. Gastric emptying 2 h after ingestion of a solid chow meal and colonic transit (time to expel a bead inserted into the distal colon) were determined simultaneously. Rat/human (r/h)CRF, which has CRF(1) > CRF(2) binding affinity, decreased distal colonic transit time at lower doses (6-12 microg/kg) than those inhibiting gastric emptying (20-60 microg/kg). Ovine CRF, a preferential CRF(1) receptor agonist (6-60 microg/kg), reduced significantly the colonic transit time without altering gastric emptying, whereas the selective CRF(2) receptor agonists mouse urocortin II (20-60 microg/kg) and urocortin III (120 microg/kg) inhibited significantly gastric emptying without modifying colonic transit. The CRF(1)/CRF(2) receptor antagonist, astressin (30-120 microg/kg), dose dependently prevented r/hCRF (20 microg/kg)-induced inhibition of gastric emptying and reduction of colonic transit time. The selective CRF(1) receptor antagonists, NBI-27914 (C(18)H(20)Cl(4)N(4)C(7)H(8)O(3)S) and CP-154,526 (butyl-[2,5-dimethyl-7-(2,4,6-trimethylphenyl)-7H-pyrrolo[2,3-d]pyrimidin-4-yl]ethylamine) (5-30 mg/kg), dose dependently blocked r/hCRF action on the colon without influencing the gastric response, whereas the CRF(2) receptor antagonist, antisauvagine-30 (30-100 microg/kg), dose dependently abolished r/hCRF-induced delayed gastric emptying and had no effect on colonic response. These data show that i.p. r/hCRF-induced opposite actions on upper and lower gut transit in conscious mice are mediated by different CRF receptor subtypes: the activation of CRF(1) receptors stimulates colonic propulsive activity, whereas activation of CRF(2) receptors inhibits gastric emptying.

Peripheral CRF activates myenteric neurons in the proximal colon through CRF1 receptor in conscious rats

Corticotropin-releasing factor (CRF) injected peripherally induces clustered spike-burst activity in the proximal colon through CRF(1) receptors in rats. We investigated the effect of intraperitoneal CRF on proximal colon ganglionic myenteric cell activity in conscious rats using Fos immunohistochemistry on the colonic longitudinal muscle/myenteric plexus whole mount preparation. In vehicle-pretreated rats, there were only a few Fos immunoreactive (IR) cells per ganglion (1.2 +/- 0.6). CRF (10 microg/kg ip) induced Fos expression in 19.6 +/- 2.1 cells/ganglion. The CRF(1)/CRF(2) antagonist astressin (33 microg/kg ip) and the selective CRF(1) antagonist CP-154,526 (20 mg/kg sc) prevented intraperitoneal CRF-induced Fos expression in the proximal colon (number of Fos-IR cells/ganglion: 2.7 +/- 1.2 and 1.0 +/- 1.0, respectively), whereas atropine (1 mg/kg sc) had no effect. Double labeling of Fos with protein gene product 9.5 revealed the neuronal identity of activated cells that were encircled by varicose fibers immunoreactive to vesicular acetylcholine transporter. Fos immunoreactivity was mainly present in choline acetyltransferase-IR nerve cell bodies but not in the NADPH-diaphorase-positive cells. These results indicate that peripheral CRF activates myenteric cholinergic neurons in the proximal colon through CRF(1) receptor.

Peripheral corticotropin-releasing factor induces diarrhea in rats: role of CRF 1 receptor in fecal watery excretion

Systemic injection of corticotropin-releasing factor (CRF) stimulates colonic secretory and motor functions, and CRF receptors play a role in stress-related alterations of colonic functions. Stress has also been reported to induce diarrhea and we investigated if peripheral injection of CRF can mimic this response in conscious rats. Intravenous (i.v.) injection of CRF (3, 10 or 30 μg/kg) caused diarrhea in 13%, 63% and 75% of rats, respectively, and dose dependently increased the fecal fluid content by 5.1-, 8.6- and 10.8-fold, while the dried solid weight was increased by 5.2-, 4.9- and 5.8-fold, respectively, compared to the i.v. saline group. CRF actions were rapid in onset and blocked by the CRF 1 receptor, antagonist CP-154,526 (butyl-[2,5-dimethyl-7-(2,4,6-trimethylphenyl)-7 H-pyrrolo[2,3- d]pyrimidin-4-yl]ethylamine). These results demonstrate that peripheral CRF induces watery diarrhea, primarily through the activation of CRF 1 receptor suggesting a possible role for these pathways in colonic responses to stress.

Peripheral corticotropin-releasing factor and stress-stimulated colonic motor activity involve type 1 receptor in rats

Background & Aims: Corticotropin-releasing factor (CRF) exerts its action through CRF receptors 1 and 2 (CRF-R1 and CRF-R2). CRF has preferential affinity for CRF-R1, whereas urocortin displays high affinity for both. We investigated changes in colonic motor function after intraperitoneal (IP) injection of CRF-related peptides. Methods: Colonic motility was recorded in vivo in conscious rats equipped with electrodes chronically implanted in the cecum and proximal colon or in vitro in distal colon; fecal output was monitored in naive rats. Results: Rat CRF, rat urocortin, and amphibian sauvagine (10 μg/kg, IP) induced a new pattern of cecocolonic myoelectric activity characterized by clustered spike bursts of long duration; the percentage of occurrence was highest after CRF. The rank order of potency to increase fecal pellet output after IP peptide injection (0.3–10 μg/kg, IP) was CRF > urocortin = sauvagine. The CRF-R1/R2 antagonist astressin (33 μg/kg, IP) and the CRF-R1 antagonist CP-154,526 (20 mg/kg, subcutaneously) inhibited IP CRF-induced changes in cecocolonic myoelectric activity and IP CRF- and water avoidance stress–induced fecal output. In vitro, CRF injected into the inferior mesenteric artery increased distal colonic myoelectric activity compared with saline injection. Conclusions: These results demonstrate that CRF acts peripherally to stimulate colonic motility and that CRF-R1 is primarily involved in mediating IP CRF/urocortin– and water avoidance stress–induced colonic motor response.GASTROENTEROLOGY 2000;119:1569-1579