Background & Aims Rifaximin is used to treat patients with functional gastrointestinal disorders, but little is known about its therapeutic mechanism. We propose that rifaximin modulates the ileal bacterial community, reduces subclinical inflammation of the intestinal mucosa, and improves gut barrier function to reduce visceral hypersensitivity. Methods We induced visceral hyperalgesia in rats, via chronic water avoidance or repeat restraint stressors, and investigated whether rifaximin altered the gut microbiota, prevented intestinal inflammation, and improved gut barrier function. Quantitative polymerase chain reaction and 454 pyrosequencing were used to analyze bacterial 16S rRNA in ileal contents from the rats. Reverse transcription, immunoblot, and histologic analyses were used to evaluate levels of cytokines, the tight junction protein occludin, and mucosal inflammation, respectively. Intestinal permeability and rectal sensitivity were measured. Results Water avoidance and repeat restraint stress each led to visceral hyperalgesia, accompanied by mucosal inflammation and impaired mucosal barrier function. Oral rifaximin altered the composition of bacterial communities in the ileum (Lactobacillus species became the most abundant) and prevented mucosal inflammation, impairment to intestinal barrier function, and visceral hyperalgesia in response to chronic stress. Neomycin also changed the composition of the ileal bacterial community (Proteobacteria became the most abundant species). Neomycin did not prevent intestinal inflammation or induction of visceral hyperalgesia induced by water avoidance stress. Conclusions Rifaximin alters the bacterial population in the ileum of rats, leading to a relative abundance of Lactobacillus. These changes prevent intestinal abnormalities and visceral hyperalgesia in response to chronic psychological stress.

The vagal afferent system is strategically positioned to mediate rapid changes in motility and satiety in response to systemic glucose levels. In the present study we aimed to identify glucose-excited and glucose-inhibited neurons in nodose ganglia and characterize their glucose-sensing properties. Whole-cell patch-clamp recordings in vagal afferent neurons isolated from rat nodose ganglia demonstrated that 31/118 (26%) neurons were depolarized after increasing extracellular glucose from 5 to 15 mm; 19/118 (16%) were hyperpolarized, and 68/118 were non-responsive. A higher incidence of excitatory response to glucose occurred in gastric-than in portal vein-projecting neurons, the latter having a higher incidence of inhibitory response. In glucose-excited neurons, elevated glucose evoked membrane depolarization (11 mV) and an increase in membrane input resistance (361 to 437 M ). Current reversed at −99 mV. In glucose-inhibited neurons, membrane hyperpolarization (−13 mV) was associated with decreased membrane input resistance (383 to 293 M ). Current reversed at −97 mV. Superfusion of tolbutamide, a K ATP channel sulfonylurea receptor blocker, elicited identical glucose-excitatory but not glucose-inhibitory responses. Kir6.2 shRNA transfection abolished glucose-excited but not glucose-inhibited responses. Phosphatidylinositol bisphosphate (PIP 2 ) depletion using wortmannin increased the fraction of glucose-excited neurons from 26% to 80%. These results show that rat nodose ganglia have glucose-excited and glucose-inhibited neurons, differentially distributed among gastric-and portal vein-projecting nodose neurons. In glucose-excited neurons, glucose metabolism leads to K ATP channel closure, triggering membrane depolarization, whereas in glucose-inhibited neurons, the inhibitory effect of elevated glucose is mediated by an ATP-independent K + channel. The results also show that PIP 2 can determine the excitability of glucose-excited neurons.

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Research has shown that the synergistic interaction between vagal cholecystokinin-A receptors (CCKARs) and leptin receptors (LRbs) mediates short term satiety. We hypothesize that this synergistic interaction is mediated by cross-talk between signaling cascades used by CCKARs and LRbs, which, in turn, activates closure of K ؉ channels, leading to membrane depolar- Leptin, the product of the ob gene, is secreted primarily from white adipocyte tissue; its level in the circulation correlates with the degree of adiposity (1, 2). Circulating leptin crosses the blood-brain barrier via a receptor-mediated transport system (3, 4) and acts on the long form of the leptin receptor (LRb) 2 in the medial hypothalamus to regulate feeding behavior and energy balance (5). Leptin is secreted from several other sites, including the gastric mucosa, brown adipocyte tissue, placenta, mammary gland, ovarian follicles, and brain (5, 6). Leptin mRNA and leptin protein have also been detected in human stomach mucosa (7) and rat gastric fundus (8). Leptin levels in the stomach are altered by nutritional state and by cholecystokinin (CCK) administration. CCK is not, however, a stimulus for leptin release from isolated adipocytes (8). Leptin is the key signaling molecule responsible for long term satiety and energy balance; mutations that cause defective leptin secretion or abnormal leptin receptor signaling result in obesity in ob/ob mice (9, 10) and in humans (11). The leptin receptor belongs to the IL-6 receptor family of class 1 cytokine receptors and mediates the biological effects of leptin via the Janus kinase 2-signal transducer and activator of transcription 3 (JAK2/STAT3) pathway (12-14). Several splice variants of the leptin receptor exist; however, the LRb isoform mediates the leptin effect on satiety (4). CCK is an endogenous peptide found in the gastrointestinal tract and the brain. It is released into the circulation after a meal and acts on neurons both centrally and peripherally (15). The satiety action of CCK appears to be mediated by low affinity CCK-A receptors (CCKARs) on vagal afferent neurons (16). Systemic administration of CCK inhibits food intake in several species, including rats and humans (17), giving credence to the hypothesis that peripheral CCK acts as a satiety signal. CCK cannot penetrate the blood-brain barrier; therefore, systemically administered CCK likely acts at a peripheral site to inhibit feeding (18). In contrast to leptin, the effect of CCK on food intake occurs within 15 min after intraperitoneal administration of CCK-8, suggesting that CCK may act as a meal-related short term satiety signal (19,20).Both CCKARs and LRbs are widely distributed in nodose ganglia (NG) and the vagus nerve (21,22). There is evidence that a synergistic interaction between leptin and CCK leads to the reduction of short term food intake (23)(24)(25). In fact, the satiety action of CCK appears to depend on leptin signaling (26). Currently, the intracellular signaling mechanisms responsible for the synergistic interacti...

Gastrin and its carboxyl-terminal homolog cholecystokinin (CCK) exert a variety of biological actions in the brain and gastrointestinal tract that are mediated in part through one or more G protein-coupled receptors which exhibit similar affinity for both peptides. Genomic clones encoding a human gastrin/CCKB receptor were isolated by screening a human EMBL phage library with a partial-length DNA fragment which was based on the nucleotide sequence of the canine gastrin receptor. The gene contained a 1356-bp open reading frame consisting of five exons interrupted by 4 introns and was assigned to human chromosome llpl5.4. A region of exon 4, which encodes a portion of the putative third intracellular loop, appears to be alternatively spliced to yield two different mRNAs, one containing (452 amino acids; long isoform) and the other lacking (447 amino acids; short isoform) the pentapeptide sequence Gly-Gly-Ala-Gly-Pro. The two receptor isoforms may contribute to functional differences in gastrin-and CCK-mediated signal transduction. (12,13). They encode homologous proteins of similar length (453 and 450 aa, respectively) with seven deduced transmembrane regions, comparable to other G protein-coupled receptors. The recombinant receptors, when transfected into COS-7 cells, manifest high (nanomolar) affinities for both gastrin and CCK and are coupled to an increase in intracellular calcium (12,13). Structurally, the canine gastrin receptor displays nearly 90% amino acid identity with the putative CCKB receptors from human and rat (9-11).The sequence and organization of the full-length genes encoding members of the gastrin/CCK receptor family have not been identified. Such information is essential for analysis of their transcriptional regulation and structural relationships to other receptor genes. We have cloned and sequenced a gastrin/CCKB receptor gene from a human genomic library**. The gene appears to be alternatively spliced to yield two different receptor isoforms which differ by 5 aa in the putative third intracellular loop. The gastrin/CCKB receptor gene was assigned to human chromosome 11p15.4 by fluorescence in situ hybridization. METHODSGenomic Library Screening and DNA Sequencing. A human genomic library in the bacteriophage AEMBL-3 (Clontech) was screened with a 32P-labeled cDNA fragment generated by the polymerase chain reaction (PCR) with primers based on the cDNA sequence of the canine gastrin receptor (see below). Two positive clones were digested with Sst I and the resulting restriction fragments of length 0.7-3.2 kb (Fig. 1) were subcloned into phage M13mpl8 or -mpl9 and sequenced in both directions by the dideoxy method (14). In Abbreviation: CCK, cholecystokinin. tPresent address: