Ghrelin inhibits autonomic response to gastric distension in rats by acting on vagal pathway.

Ghrelin is the only orexigenic peptide currently known and a potent prokinetic by promoting gastric motility but novel insights suggest that its role extends beyond satiety regulation. Whereas ghrelin was shown to provide somatic and colonic antinociception, its impact on gastric sensitivity is unknown even though stomach is a major ghrelin secreting tissue. Autonomic response to gastric mechanosensitivity was estimated by measuring blood pressure variation as a surrogate marker in response to gastric distension (GD) before and after ghrelin (or vehicle) administration. Involvement of spinal and vagal pathways in the ghrelin effect was studied by performing celiac ganglionectomy and subdiaphragmatic vagotomy respectively and by evaluating the expression of phosphorylated extracellular-regulated kinase 1/2 (p-ERK1/2) in dorsal root and nodose ganglia. Finally the phenotype of Ghrelin receptor expressing neurons within the nodose ganglia was determined by in situ hybridization and immunofluorescence. Ghrelin reduced blood pressure variation in response to GD except in vagotomized rats. Phosphorylated-ERK1/2 levels indicated that ghrelin reduced neuronal activation induced by GD in nodose ganglion. The effect of ghrelin on gastric mechanosensitivity was abolished by pre-treatment with antagonist [D-Lys3]-GHRP-6 (0.3 mg/kg i.v.). Immunofluorescence staining highlights the colocalization of Ghrelin receptor with ASIC3 and TRPV1 within gastric neurons of nodose ganglion. Ghrelin administration reduced autonomic response to gastric distension. This effect likely involved the Ghrelin receptor and vagal pathways.

Citrulline decreases hepatic endotoxin-induced injury in fructose-induced non-alcoholic liver disease: an ex vivo study in the isolated perfused rat liver.

Steatosis can sensitise the liver to various challenges and favour the development of non-alcoholic fatty liver disease (NAFLD). In this context, fructose feeding promotes endotoxin translocation from the gut, contributing to disease progression via an inflammatory process. Citrulline is protective against fructose-induced NAFLD; we hypothesised that this property might be related to its anti-inflammatory and antioxidative action against endotoxin-induced hepatic injuries. This hypothesis was evaluated in a model of perfused liver isolated from NAFLD rats. Male Sprague-Dawley rats (n 30) were fed either a standard rodent chow or a 60 % fructose diet alone, or supplemented with citrulline (1 g/kg per d) for 4 weeks. After an evaluation of their metabolic status, fasted rats received an intraperitoneal injection of lipopolysaccharide (LPS) (2·5 mg/kg). After 1 h, the livers were isolated and perfused for 1 h to study liver function and metabolism, inflammation and oxidative status. In vivo, citrulline significantly decreased dyslipidaemia induced by a high-fructose diet and insulin resistance. In the isolated perfused rat livers, endotoxaemia resulted in higher cytolysis (alanine aminotransferase release) and higher inflammation (Toll-like receptor 4) in livers of fructose-fed rats, and it was prevented by citrulline supplementation. Oxidative stress and antioxidative defences were similar in all three groups. Amino acid exchanges and metabolism (ammonia and urea release) were only slightly different between the three groups. In this context of mild steatosis, our results suggest that fructose-induced NAFLD leads to an increased hepatic sensitivity to LPS-induced inflammation. Citrulline-induced restriction of the inflammatory process may thus contribute to the prevention of NAFLD.

Sex-related effects of nutritional supplementation of Escherichia coli: relevance to eating disorders.

OBJECTIVES: The biological background of sex-related differences in the development of eating disorders (EDs) is unknown. Recent data showed that gut bacteria Escherichia coli induce autoantibodies against anorexigenic α-melanocyte-stimulating hormone (α-MSH) associated with psychopathology in ED. The aim of this study was to compare the effects of E. coli on feeding and autoantibodies against α-MSH and adrenocorticotropic hormone (ACTH), between female and male rats. METHODS: Commensal E. coli K12 were given in a culture medium daily to adult Wistar rats by intragastric gavage over a 3-wk period; control rats received culture medium only. RESULTS: Before gavage, E. coli K12 DNA was detected in feces of female but not male rats. E. coli provision was accompanied by an increase in body weight gain in females, but a decrease in body weight gain and food intake in males. Independent of E. coli treatment, plasma levels of anti-α-MSH and ACTH immunoglobulin (Ig)G were higher in female than male rats. Females responded to E. coli by increasing α-MSH IgG levels and affinity, but males by increasing α-MSH IgM levels. Affinity of IgG for ACTH was increased in both E. coli-treated females and males, although with different kinetics. IgG from females stimulated more efficiently α-MSH-induced cyclic adenosine monophosphate production by melanocortin 4 receptor-expressing cells compared with IgG from males. DISCUSSION: Sex-related response to how E. coli affects feeding and anti-melanocortin hormone antibody production may depend on the presence of these bacteria in the gut before E. coli supplementation. These data suggest that sex-related presence of certain gut bacteria may represent a risk factor for ED development.

Effects of rabbit anti-α-melanocyte-stimulating hormone (α-MSH) immunoglobulins on α-MSH signaling related to food intake control.

Anti-α-melanocyte-stimulating hormone (α-MSH) polyclonal antibodies have been used for α-MSH neutralization in functional studies, but the results are sometime inconsistent with the antibody expected blocking properties. The present study aimed to determine if rabbit (Rb) anti-α-MSH immunoglobulins (Ig) may inhibit or enhance α-MSH signaling on melanocortin receptor type 4 (MC4R) and α-MSH-induced anorexigenic effect if presented as immune complexes with α-MSH. Polyclonal Rb anti-α-MSH IgG were commercially available and their ability to bind α-MSH has been confirmed by the immunohistochemical detection of α-MSH neurons in the rat hypothalamus. In vitro assay of the cyclic-adenosine mono-phosphate (cAMP) secreted by cells transfected with MC4R was performed to analyze effect of Rb IgG on α-MSH-induced cAMP production. We found that adding Rb IgG to α-MSH resulted in stimulation of cAMP detected at lower peptide concentrations as compared to α-MSH alone. To determine effects of Rb IgG on food intake, rats were injected into the arcuate hypothalamic nucleus with either α-MSH, Rb IgG alone or Rb IgG preincubated with α-MSH. During 2 days after injections, food intake was increased in both groups of rats receiving Rb IgG. However, during following 4 days when food was restricted to 1h/day, only the Rb IgG group displayed higher food intake. Furthermore, after the refeeding, 24h food intake was lower in rats receiving Rb IgG - α-MSH immune complexes. This group of rats was also characterized by higher number of immunopositive neurons in the arcuate nucleus expressing α-MSH and agouti-related protein but not tyrosine hydroxylase. Taken together, these results show that Rb anti-α-MSH antisera, although efficient for immunohistochemical detection of α-MSH, does not always display α-MSH blocking properties but, in contrast, may enhance α-MSH binding to MC4R and increase α-MSH anorexigenic effects when presented as immune complexes with the peptide.

Glutamine supplementation, but not combined glutamine and arginine supplementation, improves gut barrier function during chemotherapy-induced intestinal mucositis in rats.

BACKGROUND & AIMS: Increased intestinal permeability occurs during chemotherapy-induced intestinal mucositis. Previous data suggest that glutamine and arginine may have additive or synergic effects to limit intestinal damage. The present study aimed to evaluate the effects of glutamine and arginine, each alone or in combination, on gut barrier function during methotrexate (MTX)-induced mucositis in rats. METHODS: Eighty Sprague Dawley rats received during 7 days (d) standard chow supplemented with protein powder (PP), glutamine (G, 2%), arginine (A, 1.2%) or glutamine plus arginine (GA). All diets were isonitrogenous. Rats received subcutaneous injections of MTX (2.5 mg/kg) from d0 to d2. The intestinal permeability and tight junction proteins were assessed at d4 and d9 in the jejunum by FITC-dextran and by western blot and immunohistochemistry, respectively. RESULTS: At d4, intestinal permeability was increased in MTX-PP, MTX-A and MTX-GA rats compared with controls but not in MTX-G rats. The expression of claudin-1, occludin and ZO-1 was decreased in MTX-PP group compared with controls but was restored in MTX-G and MTX-A rats. In MTX-GA rats, occludin expression remained decreased. These effects could be explained by an increase of erk phosphorylation and a decrease of IκBα expression in MTX-PP and MTX-GA rats. At d9, Intestinal permeability remained higher only in MTX-GA rats. This was associated with a persistent decrease of occludin expression. CONCLUSIONS: Glutamine prevents MTX-induced gut barrier disruption by regulating occludin and claudin-1 probably through erk and NF-κB pathways. In contrast, combined glutamine and arginine has no protective effect in this model.

Alteration of intestinal barrier function during activity-based anorexia in mice.

BACKGROUND & AIMS: Anorexia nervosa is a severe eating disorder often leading to malnutrition and cachexia, but its pathophysiology is still poorly defined. Chronic food restriction during anorexia nervosa may induce gut barrier dysfunction, which may contribute to disease development and its complications. Here we have characterized intestinal barrier function in mice with activity-based anorexia (ABA), an animal model of anorexia nervosa. METHODS: Male C57Bl/6 ABA or limited food access (LFA) mice were placed respectively in cages with or without activity wheel. After 5 days of acclimatization, both ABA and LFA mice had progressively limited access to food from 6 h/d at day 6 to 3 h/d at day 9 and until the end of experiment at day 17. A group of pair-fed mice (PF) was also compared to ABA. RESULTS: On day 17, food intake was lower in ABA than LFA mice (2.0 ± 0.18 g vs. 3.0 ± 0.14 g, p < 0.001) and weight loss was more pronounced in ABA and PF compared to LFA mice (23.6 ± 1.6% and 24.7 ± 0.7% vs. 16.5 ± 1.2%; p < 0.05). Colonic histology showed decreased thickness of the muscularis layer in ABA compared to LFA mice (p < 0.05). Colonic permeability was increased in both ABA and PF compared to LFA mice (p < 0.05) but jejunal paracellular permeability was not affected. Expression of claudin-1 in the colon was lower in the ABA than the LFA group (p < 0.05), whereas occludin expression remained unaffected. CONCLUSION: Increased colonic permeability and histological alterations found in ABA mice suggest that intestinal barrier dysfunction may also occur in anorexia nervosa. The role of these alterations in the pathophysiology of anorexia nervosa should be further evaluated.

Gastric distension activates NUCB2/nesfatin-1-expressing neurons in the nucleus of the solitary tract.

Brainstem structures such as the nucleus of the solitary tract (NTS) and the dorsal motor nucleus of the vagus nerve (DMNX) are essential for the digestive function of the stomach. A large number of neurotransmitters including glutamate and gamma-aminobutyric acid (GABA) are involved in the central control of gastric functions. However, the neuropeptidergic systems implicated in this process remain undetermined. Nesfatin-1 was recently identified as a neuropeptide cleaved from the N-terminal part of NEFA/nucleobindin 2 precursor (NUCB2). Central administration of this neuropeptide inhibits food consumption and gastroduodenal motility in rodents. Interestingly, the NTS and the DMNX contain a dense population of NUCB2/nesfatin-1 cell bodies. These observations led us to investigate the possible involvement of NUCB2/nesfatin-1 neurons in the brainstem neuronal pathways that modulate gastric functions. We observed an activation of NTS NUCB2/nesfatinergic neurons after gastric distention in rats. In addition, we found that several NTS NUCB2/nesfatinergic neurons were GABAergic. Finally, when fluorogold was injected at the stomach level, many retrogradely labeled neurons were observed in the DMNX which were also positive for NUCB2/nesfatin-1. Taken together, these observations suggest for the first time that NUCB2/nesfatin-1 neurons of the NTS are sensitive to gastric distension and then may contribute to the satiety signal.

Gastric electrical stimulation increases the discomfort threshold to gastric distension.

INTRODUCTION: Gastric electrical stimulation (GES) is now considered as a new therapeutic alternative for patients with medically refractory vomiting and/or nausea, although its mechanisms of action remain poorly understood. METHODS AND PATIENTS: Gastric discomfort threshold, measured as the gastric maximal tolerable volume (MTV) to distension, was examined before and after GES, in nine patients implanted for chronic and severe nausea and vomiting. RESULTS: GES increased gastric MTV from 522 ± 64 ml at baseline to 628 ± 60 ml 6 months after the start of GES (P=0.03), whereas gastric emptying remained unchanged. The increase in MTV was correlated with symptoms and quality of life at 6 months, whereas gastric emptying was not. Finally, MTV varied in a similar manner at 6 months in patients with delayed and normal gastric emptying measured before implantation. CONCLUSION: Taken together, these data indicate that modification of gastric sensation to distension, rather than gastric emptying, is associated with symptoms' outcome during GES.

Gastric electrical stimulation decreases gastric distension-induced central nociception response through direct action on primary afferents.

BACKGROUND & AIMS: Gastric electrical stimulation (GES) is an effective therapy to treat patients with chronic dyspepsia refractory to medical management. However, its mechanisms of action remain poorly understood. METHODS: Gastric pain was induced by performing gastric distension (GD) in anesthetized rats. Pain response was monitored by measuring the pseudo-affective reflex (e.g., blood pressure variation), while neuronal activation was determined using c-fos immunochemistry in the central nervous system. Involvement of primary afferents was assessed by measuring phosphorylation of ERK1/2 in dorsal root ganglia. RESULTS: GES decreased blood pressure variation induced by GD, and prevented GD-induced neuronal activation in the dorsal horn of the spinal cord (T9-T10), the nucleus of the solitary tract and in CRF neurons of the hypothalamic paraventricular nucleus. This effect remained unaltered within the spinal cord when sectioning the medulla at the T5 level. Furthermore, GES prevented GD-induced phosphorylation of ERK1/2 in dorsal root ganglia. CONCLUSIONS: GES decreases GD-induced pain and/or discomfort likely through a direct modulation of gastric spinal afferents reducing central processing of visceral nociception.