A crosstalk between muscarinic and CRF2 receptors regulates cellular adhesion properties of human colon cancer cells.

Patients with inflammatory bowel disease often suffer from chronic and relapsing intestinal inflammation that favor the development of colitis associated cancer. An alteration of the epithelial intestinal barrier function observed in IBD is supposed to be a consequence of stress. It has been proposed that corticotrophin-releasing factor receptor (CRF2), one of the two receptors of CRF, the principal neuromediator of stress, acts on cholinergic nerves to induce stress-mediated epithelial barrier dysfunction. Non-neuronal acetylcholine (Ach) and muscarinic receptors (mAchR) also contribute to alterations of epithelial cell functions. In this study, we investigated the mechanisms through which stress and Ach modulate epithelial cell adhesive properties. We show that Ach-induced activation of mAchR in HT-29 cells results in cell dissociation together with changes in cell-matrix contacts, which correlates with the acquisition of invasive potential consistent with a matrix metalloproteinase (MMP) mode of invasion. These processes result from mAchR subsequent stimulation of the cascade of src/Erk and FAK activation. Ach-induced secretion of laminin 332 leads to α3ß1 integrin activation and RhoA-dependent reorganization of the actin cytoskeleton. We show that Ach-mediated effects on cell adhesion are blocked by astressin 2b, a CRF2 antagonist, suggesting that Ach action depends partly on CRF2 signaling. This is reinforced by the fact that Ach-mediated activation of mAchR stimulates both the synthesis and the release of CRF2 ligands in HT-29 cells (effects blocked by atropine). In summary, our data provides evidence for a novel intracellular circuit involving mAchR acting on CRF2-signaling that could mediate colonic mucosal barrier dysfunction and exacerbate mucosal inflammation.

Brain responses to uncertainty about upcoming rectal discomfort in quiescent Crohn's disease - a fMRI study.

BACKGROUND: Patients with Crohn's disease (CD) in remission are exposed to chronic psychological distress, due to the constant risk of relapse. This permanent situation of anticipation and uncertainty can lead to anxiety, which may, in turn, trigger relapse. We aimed to investigate the effects of uncertainty on behavioral and brain responses to anticipation of visceral discomfort in quiescent CD patients. METHODS: Barostat-controlled rectal distensions were preceded by cued uncertain or certain anticipation in nine CD patients and nine matched healthy volunteers. Brain responses obtained before distension across the different anticipation conditions in regions of interest (ROI) involved in (anticipation of) pain were measured using functional magnetic resonance imaging and compared between CD and controls. The association between anxiety-related psychological variables and cerebral anticipatory activity was tested. KEY RESULTS: During uncertainty, CD patients had significantly stronger activations than controls in the cingulate cortex, insula, amygdala, and thalamus with trends in the hippocampus, prefrontal, and secondary somatosensory cortex. In patients, brain responses to uncertainty in the majority of ROI correlated positively with gastrointestinal symptom-specific anxiety, trait-anxiety, and intolerance of uncertainty. CONCLUSIONS & INFERENCES: In a context of uncertainty regarding occurrence of uncomfortable visceral sensations, CD is associated with excessive reactivity in brain regions known to be involved in sensory, cognitive and emotional aspects of pain processing and modulation, and threat appraisal. Our findings contribute to a better understanding of the role of emotional and cognitive processes in CD. This may, in turn, lead to the development of new (psycho)therapeutic approaches for management of symptoms and related anxiety.

Chronic vagus nerve stimulation in Crohn's disease: a 6-month follow-up pilot study.

The vagus nerve (VN) is a link between the brain and the gut. The VN is a mixed nerve with anti-inflammatory properties through the activation of the hypothalamic-pituitary-adrenal axis by its afferents and by activating the cholinergic anti-inflammatory pathway through its efferents. We have previously shown that VN stimulation (VNS) improves colitis in rats and that the vagal tone is blunted in Crohn's disease (CD) patients. We thus performed a pilot study of chronic VNS in patients with active CD. Seven patients under VNS were followed up for 6 months with a primary endpoint to induce clinical remission and a secondary endpoint to induce biological (CRP and/or fecal calprotectin) and endoscopic remission and to restore vagal tone (heart rate variability). Vagus nerve stimulation was feasible and well-tolerated in all patients. Among the seven patients, two were removed from the study at 3 months for clinical worsening and five evolved toward clinical, biological, and endoscopic remission with a restored vagal tone. These results provide the first evidence that VNS is feasible and appears as an effective tool in the treatment of active CD.

Factors associated with pregnancy outcome in anti-TNF treated women with inflammatory bowel disease.

BACKGROUND: The safety of anti-tumour necrosis factor (TNF) agents during pregnancy is a major concern for child-bearing women and physicians. AIM: To assess the impact of anti-TNF therapy on adverse pregnancy and foetal outcomes in women with inflammatory bowel disease (IBD). METHODS: Pregnancies occurring during anti-TNF treatment or less than 3 months after its cessation in IBD patients followed in GETAID centres were recorded from January 2009 to December 2010. Ninety-nine pregnancies in women without anti-TNF treatment were identified from the CESAME registry. We compared pregnancy and neonatal outcomes by a case-control study. RESULTS: In the 124 IBD patients followed, 133 pregnancies were reported. At the conception time, 23% of patients had active disease. Eighty-eight per cent (n = 117) of the 133 pregnancies followed until delivery resulted in 118 liveborns (one twin pregnancy). Complications were observed in 47 (35%) women and 24 (20%) newborns. In multivariate analysis, factors associated with pregnancy complications were: current smoking (P = 0.004), a B2 (stenotic) phenotype in CD women (P = 0.004), occurrence of a flare during pregnancy (P = 0.006) and a past history of complicated pregnancy (P = 0.007). Current smoking was the only factor associated with severe (i.e. potentially lethal) pregnancy complications (P = 0.02). Having IBD for more than 10 years prior to conception was associated with newborn complications (P = 0.007). No difference was found with the control group for any of the pregnancy and neonatal outcomes. CONCLUSION: In our series, the safety profile of anti-TNF therapy during pregnancy and the neonatal period appears similar to control group of IBD women not treated with anti-TNF therapy.

The link between negative affect, vagal tone, and visceral sensitivity in quiescent Crohn's disease.

Autonomic dysfunction and mood disorders are frequently described in Crohn's disease (CD) and are known to influence visceral sensitivity. We addressed the link between vagal tone, negative affect, and visceral sensitivity in CD patients without concomitant features of irritable bowel syndrome (IBS). Rectal distensions to a discomfort threshold of 70% and onset of pain were performed in nine CD patients in remission and eight healthy controls. Autonomic parameters were evaluated with heart rate variability and electrodermal reactivity. We showed that CD patients had (i) higher scores of depressive symptomatology (12 ± 3 in patients vs 4 ± 1 in controls on the Center for Epidemiologic Studies-Depression Scale; p = 0.038), (ii) reduced vagal tone (HF 257 ± 84 ms(2) vs 1607 ± 1032 ms(2) , p = 0.043; LF 455 ± 153 ms(2) vs 1629 ± 585 ms(2) , p = 0.047), (iii) decreased sympathetic reactivity during an aversive stimulus, and (iv) higher tolerance to rectal distension pressures (43 ± 3 mmHg vs 30 ± 2 mmHg, p = 0.002) and low sensitivity index scores. In conclusion, our results provide preliminary evidence that patients with quiescent CD, in the absence of IBS, are hyposensate to experimental rectal distension. These data provide further evidence that anxiety and depressive symptomatology in addition to autonomic dysfunction modulate visceral pain perception in quiescent CD patients in the absence of IBS.

Hyperhomocysteinaemia is associated with osteoporosis in patients with Crohn's disease.

BACKGROUND: A high prevalence of osteoporosis is observed in Crohn's disease. Recent data have shown that homocysteinaemia is an important risk factor in low-bone mineralization and fracture. AIM: To look for an association between homocysteinaemia and low-bone mineralization in Crohn's disease patients. PATIENTS AND METHODS: Ninety-two consecutive patients (sex ratio M/F 0.87; mean age: 36.6 +/- 13.2 years) were recruited between 2003 and 2005. Bone densitometry was performed on inclusion. The following parameters were analysed: age, sex, Crohn's Disease Activity Index, duration and extent of Crohn's disease, smoking status, corticosteroid treatment, immunosuppressive drugs, plasma homocysteine, folate and vitamin B12 concentration. RESULTS: The prevalence of a high homocysteine level (>15 micromol/L) was 60%. Osteoporosis and low-bone mineralization observed in 26 (28%), and 60 (65%) patients, respectively. On a multivariate analysis, associated factors for osteoporosis and low-bone mineralization were respectively: hyperhomocysteinaemia (OR: 61.4; CI: 95: 23-250; P < 0.001), and ileal Crohn's disease [OR: 13.8; CI: 95: 2.5-150; P = 0.036] for osteoporosis and hyperhomocysteinaemia [OR: 63.7; CI: 95: 8.5-250; P < 0.001] and disease duration of at least 5 years [OR: 11.4; CI: 95: 1.31-99; P = 0.039] for low-bone mineralization. Results were similar whichever site osteoporosis was detected. CONCLUSION: Hyperhomocysteinaemia was observed in 60% of our Crohn's disease patients and was strongly associated with low-bone mineralization and osteoporosis (OR: 61.4).

[Factors associated with hyperhomocysteinemia in inflammatory bowel disease: prospective study in 81 patients]. / Hyperhomocystéinémie et facteurs associés au cours des MICI: étude prospective chez 81 patients.

BACKGROUND: A high prevalence (52%) of hyperhomocysteinemia is observed in Crohn disease (CD), however it is not well documented in ulcerative colitis (UC). Furthermore, in the different works studying hyperhomocysteinemia the associated factors are different. AIM: Prospective evaluation of hyperhomocysteinemia in inflammatory bowel disease (IBD) patients, of the risk factors and the determination of a potential risk of colorectal carcinoma in case of hyperhomocysteinemia. PATIENTS AND METHODS: IBD patients followed in our department were prospectively recruited between November 2003-September 2004. To be included patients should have passed a coloscopy in the two years. Patients with kidney failure or drugs supposed, to interfere with homocystéine metabolism (folates, vitamin B12, methotrexate) were excluded from the study. The following parameters were analysed: age, sex, clinical activity indexes (CDAI for Crohn disease and CAI for ulcerative colitis), length-extent and type of the disease (CD or UC), smoking, plasma homocystein concentration, folates and vitamin B12. RESULTS: Eighty-one patients (60 CD, 21 UC, mean age 43.8 +/- 17.3) were included, 30 had an active disease at inclusion and 16 were smokers. The prevalence of high homocystein concentration was 55.6%. In univariate analysis a low rate of folates was the only risk factor for a high homocystein concentration (74 vs. 52.8%; P = 0.018). Smoking was almost an associated factor. In multivariate analysis, a low rate of folate was the only risk factor of hyperhomocysteinemia, OR = 3.59 [1.27-10.17]. Five endoscopic lesions considered as precancerous were described; these patients had all a hyperhomocysteinemia. CONCLUSION: The prevalence of hyperhomocysteinemia is high in UC and in CD. A low folate rate is the only risk factor observed in our study. There is a possible link between colorectal cancer and hyperhomocysteinemia. A high Plasma homocystein concentration must be search in inflammatory bowel disease patients and a substitutive treatment of folates and vitamin B12 is necessary in case of hyperhomocysteinemia.

6-tioguanine monitoring in steroid-dependent patients with inflammatory bowel diseases receiving azathioprine.

BACKGROUND: 6-Thioguanine (6-tioguanine) nucleotides are the active metabolites of azathioprine. AIM: The aim of the study was to evaluate the rate of clinical remission without steroids in steroid-dependent Crohn's disease and ulcerative colitis patients receiving azathioprine, the medium- and long-term efficacy and the predictive factors of clinical response when monitoring 6-tioguanine. METHODS: Steroid-dependent Crohn's disease and ulcerative colitis patients receiving either azathioprine or not (treated later with a daily dose of 2.5 mg/kg) were prospectively included. 6-tioguanine was monitored at 1 and 2 months and every 3 months thereafter for 1 year. The azathioprine dose was adapted to reach a 6-tioguanine level of >250 pmol/8 x 10(8) red blood cells. Thiopurine methyltransferase genotype/phenotype was evaluated in some patients. RESULTS: A total of 106 patients were prospectively included (70 Crohn's disease, 36 ulcerative colitis). The clinical remission rate without steroids in patients receiving azathioprine, in intention-to-treat analysis, was 72% and 59% at 6 and 12 months, respectively. The remission rate was significantly higher in patients with 6-tioguanine >250 pmol/8 x 10(8) RBC (86% and 69% at 6 and 12 months, respectively; P < 0.01). No significant difference was observed between Crohn's disease and ulcerative colitis patients whether treated by azathioprine or not on inclusion. In the univariate analysis, the absence of Crohn's disease stenosis, a 6-tioguanine level >250 pmol/8 x 10(8) RBC, and an increase of erythrocyte mean corpuscular volume were the factors predictive of a favourable clinical response. In the multivariate analysis, only a 6-tioguanine level of >250 pmol/8 x 10(8) red blood cells was a predictive factor of favourable clinical remission. CONCLUSIONS: Clinical remission without steroids is significantly more likely when monitoring 6-tioguanine so as to reach a level of >250 pmol/8 x 10(8) red blood cells in steroid-dependent Crohn's disease and ulcerative colitis patients receiving azathioprine (86% and 69% at 6 and 12 months, respectively).

Drug interaction between infliximab and azathioprine in patients with Crohn's disease.

BACKGROUND: A drug interaction has been observed between infliximab and methotrexate in rheumatoid arthritis. AIM: To look for an interaction between infliximab and azathioprine in Crohn's disease patients using the active metabolites of azathioprine: 6-tioguanine nucleotides. METHODS: Patients receiving azathioprine who required infliximab for ileo-colonic or ano-perineal Crohn's disease were recruited prospectively. 6-tioguanine nucleotide levels were evaluated before infusion, within 1-3 weeks after the first infusion and 3 months after the first infusion. The clinical outcome was evaluated by the Harvey-Bradshaw index or the closure of ano-perineal fistulas. RESULTS: Thirty-two patients were included (17 received one infusion and 15 received three infusions). The mean 6-tioguanine nucleotide level was comparable before and 3 months after the first infusion, but a significant increase was observed within 1-3 weeks after the first infusion (P < 0.001). In parallel, a significant decrease in leucocyte count and increase in mean corpuscular volume were observed; these modifications were normalized 3 months after infusion. An increase in 6-tioguanine nucleotide level of greater than 400 pmol/8 x 108 erythrocytes was strongly related to good tolerance and a favourable response to infliximab, with a predictive value of 100%. CONCLUSIONS: This prospective study provides evidence for a drug interaction between azathioprine and infliximab.

Tioguanine in patients with Crohn's disease intolerant or resistant to azathioprine/mercaptopurine.

BACKGROUND: Tioguanine (TG) is an antimetabolite which may be regarded as an alternative to azathioprine (AZA)/mercaptopurine (MP) in patients with inflammatory bowel diseases. AIMS: : To evaluate the tolerance and efficacy of TG in patients with Crohn's disease, intolerant or resistant to AZA/MP. METHODS: An open prospective study was made on Crohn's disease patients treated with TG. Intolerance to AZA/MP was defined as a reaction occurring within 1 month after introduction of AZA/MP, including pancreatitis, abdominal pain, fever, arthralgia, myalgia, cutaneous rash, fatigue, alopecia, hepatitis and digestive intolerance. Resistance to AZA/MP was defined as the persistence of activity after at least 3 months of AZA/MP therapy. RESULTS: Forty-nine Crohn's disease patients (36 women, 13 men; intolerance: n = 39; resistance: n= 10) were treated with TG (20 mg/day). Clinical pancreatitis did not recur under TG. Five patients (10%) had to stop TG due to intolerant reactions observed 13-21 days after TG was started. No haematological side-effects were observed under TG. The probability of clinical remission without corticosteroids or infliximab at 6 and 12 months was 46% and 79%, respectively, in the 40 patients with active disease at baseline. The probability of clinical relapse during maintenance TG therapy at 6 and 12 months was 29% and 53%, respectively, in the 28 patients in remission at baseline or who had achieved remission on TG. CONCLUSIONS: TG is a possible alternative treatment in Crohn's disease patients, intolerant (especially for pancreatitis) or resistant to AZA/MP.

Genotypic analysis of thiopurine S-methyltransferase in patients with Crohn's disease and severe myelosuppression during azathioprine therapy.

BACKGROUND & AIMS: Myelosuppression in patients with Crohn's disease (CD) treated with azathioprine has been attributed to low activity of thiopurine S-methyltransferase (TPMT). Allelic variants of the TPMT gene responsible for changes in the enzyme activity have been characterized. We investigated the distribution of mutant alleles associated with TPMT deficiency in patients with CD and myelosuppression during azathioprine/6-mercaptopurine therapy. METHODS: Forty-one patients with CD were included. They developed leukopenia or thrombocytopenia during azathioprine or 6-mercaptopurine treatment. Polymerase chain reaction-based methods were used to search for mutations associated with TPMT deficiency. RESULTS: Four patients (10%) had 2 mutant alleles associated with TPMT deficiency, 7 (17%) had 1 mutant allele, and 30 (73%) had no known TPMT mutation. The delay between administration of the drug and occurrence of bone marrow toxicity was less than 1.5 months in the 4 patients with 2 mutant alleles, and ranged from 1 to 18 months in patients with 1 mutant allele and from 0.5 to 87 months in patients with normal genotype. CONCLUSIONS: Twenty-seven percent of patients with CD and myelosuppression during azathioprine therapy had mutant alleles of the TPMT gene associated with enzyme deficiency. Myelosuppression is more often caused by other factors. Continued monitoring of blood cell counts remains mandatory in patients treated with azathioprine.

Fedotozine, a kappa-opioid agonist, prevents spinal and supra-spinal Fos expression induced by a noxious visceral stimulus in the rat.

Fedotozine, a kappa opioid agonist, reverses digestive ileus caused by acetic acid (AA)-induced visceral pain in rats. The aims of this study were: to map, in conscious rats, central pathways activated by AA using Fos as a marker of neuronal activation; to characterize primary afferent fibres involved in this activation; and to investigate the effect of fedotozine on AA-induced Fos expression. AA (0.6%; 10 mL kg-1) was injected i.p. in conscious rats either untreated; pretreated 14 days before with capsaicin; pretreated 20 min previously with fedotozine; or pretreated 2 h prior to fedotozine with the kappa-antagonist nor-binaltorphimine (nor-BNI). Controls received the vehicle alone. 60 min after injection of AA, rats were processed for Fos immunohistochemistry. Visceral pain was assessed by counting abdominal cramps. AA induced Fos in the thoraco-lumbar spinal cord (laminae I, V, VII and X) and numerous brain structures such as the nucleus tractus solitarius, and paraventricular nucleus (PVN) of the hypothalamus, whereas almost no Fos labelling was observed in controls. Capsaicin pretreatment blocked AA-induced Fos in all structures tested. Fedotozine significantly decreased AA-induced abdominal cramps and Fos immunoreactivity in the spinal cord and PVN, this effect being reversed by nor-BNI pretreatment. AA induces Fos in the spinal cord and numerous brain nucuei, some of which are involved in the control of digestive motility in rats. This effect is mediated through capsaicin-sensitive afferent fibres and prevented by fedotozine most likely through a peripheral action on visceral afferents.

Effect of a chronic stress on CRF neuronal activity and expression of its type 1 receptor in the rat brain.

The purpose of this study was to compare the effect of an acute versus a chronic immobilization stress on the genetic expression of c-fos and corticotropin-releasing factor type 1 receptor (CRF1 receptor) in the paraventricular nucleus (PVN) of the rat hypothalamus. Male Sprague-Dawley rats were exposed to either a single 90-min immobilization stress or the same session for 11 consecutive days. Animals were deeply anesthetized before (control); immediately, 1.5, 3, 6, or 12 h after the acute stress; or after the last session of the repeated exposures to immobilization. Coronal frozen sections (30 micrometers) of the brains were cut and mRNAs encoding the rat c-fos and CRF1 receptor were assayed by in situ hybridization histochemistry using 35S-labeled riboprobes. Localization of these transcripts within PVN CRF-immunoreactive (ir) neurons was also determined. The expression of the mRNA encoding either c-fos or CRF1 receptor was barely detectable to low in the PVN of control animals, but the acute stress session induced a robust signal for both transcripts in this endocrine nucleus. Numerous CRF-ir neurons were positive for the gene encoding either c-fos or CRF1 receptor in the PVN of acutely stressed animals. In contrast, the PVN of chronically stressed animals displayed a significantly lower CRF1 receptor mRNA signal after the last stress session. In these animals, stress-induced transcription of c-fos mRNA occurred in the magnocellular PVN 90 min after the end of the last stress session but only a low signal was detected in the parvocellular division. Moreover, very few CRF-ir neurons of the PVN expressed either the CRF1 receptor or c-fos transcript in chronically stressed rats. These data provide evidence for an adaptive cellular mechanism involving an attenuated action of CRF within the PVN in response to repeated homotypic stress exposures.

Corticotropin-releasing factor and systemic capsaicin-sensitive afferents are involved in abdominal surgery-induced Fos expression in the paraventricular nucleus of the hypothalamus.

We previously reported that abdominal surgery induces Fos expression in specific hypothalamic and medullary nuclei and also causes gastric stasis. The gastric ileus is reduced by systemic capsaicin and abolished by central injection of corticotropin-releasing factor (CRF) antagonist. We studied the influence of systemic capsaicin and intracerebroventricular (i.c.v.) injection of the CRF antagonist, alpha-helical CRF9-41, on Fos expression in the brain 1 h after abdominal surgery in conscious rats using immunocytochemical detection. In control groups (vehicle s.c. or i.c.v.), abdominal surgery (laparotomy with cecal manipulation) performed under 7-8 min of enflurane anesthesia induced Fos staining in neurons of the spinal trigeminal, C1/A1 group, ventrolateral medulla, central amygdala, parabrachial nucleus, cuneate nucleus, nucleus tractus solitarii (NTS), paraventricular nucleus of the hypothalamus (PVN) and supraoptic nucleus (SON). Capsaicin (125 mg/kg s.c., 2 weeks before) or alpha-helical CRF9-41 (50 microg i.c.v., before surgery) reduced the number of Fos-positive cells by 50% in the PVN while not modifying the number of Fos-labelled cells in the other nuclei. These results indicate that capsaicin-sensitive primary afferents and brain CRF receptors are part of the pathways and biochemical coding through which abdominal surgery activates PVN neurons 1 h post surgery.

Association between myelodysplastic syndromes and inflammatory bowel diseases. Report of seven new cases and review of the literature.

We report seven patients with both myelodysplastic syndrome (MDS) and inflammatory bowel disease (IBD): Crohn's disease in six cases, ulcerative colitis in one case. We describe their characteristics, and those of 10 previously published similar cases are presented here. Median age at diagnosis of IBD (61 years) was high, as compared to the usual age at diagnosis of IBD. IBD was diagnosed first in nine cases, MDS first in one patient, and both diseases were diagnosed simultaneously in seven cases. Concerning IBD, there was a strong predominance of Crohn's disease (15/17 cases), with an unusually high frequency of colonic involvement (11/15 cases). MDS, in 12/17 cases, showed no excess of marrow blasts. Cytogenetic analysis was abnormal in five of the 13 evaluable cases. These observations suggest that the association between MDS and IBD may not be fortuitous in some cases, and that, in particular, patients with IBD and anemia of nonobvious origin should be evaluated for MDS. The pathogenesis of those associations, however, remains unclear.

Neuronal pathways involved in abdominal surgery-induced gastric ileus in rats.

The 20-min rate of gastric emptying of a noncaloric solution and c-fos expression detected by immunohistochemistry in the brain were monitored 3 h after abdominal surgery performed under 10-min enflurane anesthesia in rats. Abdominal surgery (laparotomy and 1-min manipulation of the cecum) decreased gastric emptying from 60.8 +/- 3.4 to 25.9 +/- 3.4%. Capsaicin applied to the celiac/superior mesenteric ganglia 2 wk before the experiment reduced the delay in gastric emptying induced by abdominal surgery (46.3 +/- 3.4%), whereas perivagal capsaicin application had no effect (23.6 +/- 7.9%). The corticotropin-releasing factor (CRF) antagonist [D-Phe12, Nle21,38,C alpha MeLeu37]CRF-(12--41) injected intracisternally (10-20 micrograms) prevented postoperative gastroparesis induced by surgery, while having no effect on basal gastric emptying. Abdominal surgery increased the number of Fos-positive cells in brain nuclei regulating autonomic outflow: the nucleus of the solitary tract, locus ceruleus, paraventricular nucleus, and supraoptic nucleus of the hypothalamus. These data indicate that capsaicin-sensitive splanchnic afferent fibers and activation of CRF receptors in the brain are part of the neuronal circuitry mediating gastric stasis 3 h after abdominal surgery.

Abdominal surgery induces Fos immunoreactivity in the rat brain.

Previous neuropharmacological studies indicate that brain peptides are involved in mediating gastric stasis induced by abdominal surgery. Central pathways activated by abdominal surgery were investigated in the rat by using Fos protein as a marker of neuronal activation. Abdominal surgery (laparotomy alone or combined with cecal manipulation) was performed under brief enflurane anesthesia (7-8 minutes), and 1 hour later rats were killed and brains processed for Fos immunoreactivity. Double labeling with Fos and arginine vasopressin, oxytocin, or tyrosine hydroxylase antibodies was also performed. Abdominal surgery induced Fos staining in the nucleus tractus solitarii, paraventricular and supraoptic nuclei of the hypothalamus, locus coeruleus, and ventrolateral medulla. After abdominal surgery, 18-25% of vasopressin and 18-33% of oxytocin-labeled cells were found to be Fos positive in the paraventricular nucleus and 15% of activated cells in the nucleus tractus solitarii were positive for tyrosine hydroxylase immunoreactivity. Enflurane alone induced c-fos expression in the same brain area; however, the number of Fos-positive cells and double-labeled cells were decreased two- to fivefold and three- to eightfold, respectively, compared with the abdominal surgery groups. These data show that abdominal surgery induced activation of specific hypothalamic, pontine, and medullary neurons. These findings may have implications for the understanding of central mechanisms involved in mediating gastric ileus following abdominal surgery.

Induction of Fos immunoreactivity in the rat brain after cold-restraint induced gastric lesions and fecal excretion.

Cold-restraint alters gastrointestinal function through vagal pathways. Immunohistochemical detection of the nuclear phosphoprotein Fos (Fos-IR) was used to map brain neuronal pathways activated by cold exposure for 3 h in fasted rats maintained individually in semi-cylindrical restraining cages. Gastric lesions and fecal pellet output were also monitored. In rats exposed to cold (4 degrees C) restraint for 3 h, numerous Fos-positive nuclei were observed in the dorsal motor nucleus of the vagus, raphe pallidus, locus coeruleus, and paraventricular nucleus of the hypothalamus, and, to a lesser extent, in the raphe obscurus, parapyramidal region, and medullary noradrenergic region, bed nucleus of the stria terminalis and septum. Fecal pellet output was increased by 8 fold and gastric lesions covered 19.5 +/- 1.1% of the corpus mucosa. Rats restrained at room temperature under otherwise same conditions had little or no Fos-positive cells in these brain nuclei, no gastric erosion and a low pellet output (1.3 +/- 0.5 nb/3 h). These data, in addition to previous functional studies, provide anatomic support for the involvement of neurons in the caudal raphe nuclei, dorsal motor nucleus of the vagus and paraventricular nucleus of the hypothalamus in the autonomic and endocrine responses to cold-restraint.

Water-avoidance stress-induced c-fos expression in the rat brain and stimulation of fecal output: role of corticotropin-releasing factor.

Immunohistochemical detection of the immediate-early gene c-fos was used to determine the pattern of neuronal activity in the rat brain after exposure to water-avoidance stress known to stimulate fecal output in rats. Avoidance to water for 1 h by standing on a small platform increases pellet output and induces numerous Fos-positive cells in the parvocellular part of the paraventricular nucleus of the hypothalamus (PVN), locus coeruleus (LC) and, to a lesser extent, in the bed nucleus of the stria terminalis, lateral septum, dorsal raphe nucleus and A5 and A1 noradrenergic neurons. The corticotropin-releasing factor (CRF) antagonist, alpha-helical CRF9-41 (50 micrograms i.c.v.) reduced water-avoidance stress-induced c-fos expression mainly in the PVN and the LC (44 and 60%, respectively) and decreased by 60% the stimulated fecal output. These data indicate that water-avoidance stress activates PVN and LC neurons through CRF pathways which contribute to the stimulation of colonic motor function.