Gastric organoids: Advancing the study of H. pylori pathogenesis and inflammation.

For decades, traditional in vitro and in vivo models used for the study of Helicobacter pylori infection have relied heavily on the use of gastric cancer cell lines and rodents. Major challenges faced by these methods have been the inability to study cancer initiation in already cancerous cell lines, and the difficulty in translating results obtained in animal models due to genetic differences. These challenges have prevented a thorough understanding of the pathogenesis of disease and slowed the development of cancer therapies and a suitable vaccine against the pathogen. In recent years, the development of gastric organoids has provided great advantages over the traditional in vivo and in vitro models due to their similarities to the human stomach in vivo, their ease of use, and the capacity for long-term culture. This review discusses the advantages and limitations of existing in vivo and in vitro models of H. pylori infection, and how gastric organoids have been applied to study H. pylori pathogenesis, with a focus on how the pathogen interacts with the gastric epithelium, inflammatory processes, epithelial repair, and cancer initiation. The potential applications of organoids to address more complex questions on the role of hormones, vaccine-induced immunity are also discussed.

Homeostasis and Cancer Initiation: Organoids as Models to Study the Initiation of Gastric Cancer.

Gastric cancer represents a significant disease burden worldwide. The factors that initiate cancer are not well understood. Chronic inflammation such as that triggered by H. pylori infection is the most significant cause of gastric cancer. In recent years, organoid cultures developed from human and animal adult stem cells have facilitated great advances in our understanding of gastric homeostasis. Organoid models are now being exploited to investigate the role of host genetics and bacterial factors on proliferation and DNA damage in gastric stem cells. The impact of a chronic inflammatory state on gastric stem cells and the stroma has been less well addressed. This review discusses what we have learned from the use of organoid models to investigate cancer initiation, and highlights questions on the contribution of the microbiota, chronic inflammatory milieu, and stromal cells that can now be addressed by more complex coculture models.

Plasticity of neuroeffector transmission during bowel inflammation1.

Altered gastrointestinal (GI) function contributes to the debilitating symptoms of inflammatory bowel diseases (IBD). Nerve circuits contained within the gut wall and outside of the gut play important roles in modulating motility, mucosal fluid transport, and blood flow. The structure and function of these neuronal populations change during IBD. Superimposed on this plasticity is a diminished responsiveness of effector cells - smooth muscle cells, enterocytes, and vascular endothelial cells - to neurotransmitters. The net result is a breakdown in the precisely orchestrated coordination of motility, fluid secretion, and GI blood flow required for health. In this review, we consider how inflammation-induced changes to the effector innervation of these tissues, and changes to the tissues themselves, contribute to defective GI function in models of IBD. We also explore the evidence that reversing neuronal plasticity is sufficient to normalize function during IBD.

Excitability and Synaptic Transmission in the Enteric Nervous System: Does Diet Play a Role?

Changes in diet are a challenge to the gastrointestinal tract which needs to alter its processing mechanisms to continue to process nutrients and maintain health. In particular, the enteric nervous system (ENS) needs to adapt its motor and secretory programs to deal with changes in nutrient type and load in order to optimise nutrient absorption.The nerve circuits in the gut are complex, and the numbers and types of neurons make recordings of specific cell types difficult, time-consuming, and prone to sampling errors. Nonetheless, traditional research methods like intracellular electrophysiological approaches have provided the basis for our understanding of the ENS circuitry. In particular, animal models of intestinal inflammation have shown us that we can document changes to neuronal excitability and synaptic transmission.Recent studies examining diet-induced changes to ENS programming have opted to use fast imaging techniques to reveal changes in neuron function. Advances in imaging techniques using voltage- or calcium-sensitive dyes to record neuronal activity promise to overcome many limitations inherent to electrophysiological approaches. Imaging techniques allow access to a wide range of ENS phenotypes and to the changes they undergo during dietary challenges. These sorts of studies have shown that dietary variation or obesity can change how the ENS processes information-in effect reprogramming the ENS. In this review, the data gathered from intracellular recordings will be compared with measurements made using imaging techniques in an effort to determine if the lessons learnt from inflammatory changes are relevant to the understanding of diet-induced reprogramming.

Alternative splicing of the TRPC3 ion channel calmodulin/IP3 receptor-binding domain in the hindbrain enhances cation flux.

Canonical transient receptor potential (TRPC3) nonselective cation channels are effectors of G-protein-coupled receptors (GPCRs), activated via phospholipase C-diacylglycerol signaling. In cerebellar Purkinje cells, TRPC3 channels cause the metabotropic glutamate receptor (mGluR)-mediated slow EPSC (sEPSC). TRPC3 channels also provide negative feedback regulation of cytosolic Ca(2+), mediated by a C terminus "calmodulin and inositol trisphosphate receptor binding" (CIRB) domain. Here we report the alternative splicing of the TRPC3 mRNA transcript (designated TRPC3c), resulting in omission of exon 9 (approximately half of the CIRB domain) in mice, rats, and guinea pigs. TRPC3c expression is brain region specific, with prevalence in the cerebellum and brainstem. The TRPC3c channels expressed in HEK293 cells exhibit increased basal and GPCR-activated channel currents, and increased Ca(2+) fluorescence responses, compared with the previously characterized (TRPC3b) isoform when activated via either the endogenous M3 muscarinic acetylcholine receptor, or via coexpressed mGluR1. GPCR-induced TRPC3c channel opening rate (cell-attached patch) matched the maximum activation achieved with inside-out patches with zero cytosolic Ca(2+), whereas the GPCR-induced TRPC3b activation frequency was significantly less. Both TRPC3 channel isoforms were blocked with 2 mm Ca(2+), attributable to CIRB domain regulation. In addition, genistein blocked Purkinje cell (S)-2-amino-2-(3,5-dihydroxyphenyl) acetic acid (mGluR1)-activated TPRC3 current as for recombinant TRPC3c current. This novel TRPC3c ion channel therefore has enhanced efficacy as a neuronal GPCR-Ca(2+) signaling effector, and is associated with sensorimotor coordination, neuronal development, and brain injury.

Increased caveolae density and caveolin-1 expression accompany impaired NO-mediated vasorelaxation in diet-induced obesity.

Diet-induced obesity induces changes in mechanisms that are essential for the regulation of normal artery function, and in particular the function of the vascular endothelium. Using a rodent model that reflects the characteristics of human dietary obesity, in the rat saphenous artery we have previously demonstrated that endothelium-dependent vasodilation shifts from an entirely nitric oxide (NO)-mediated mechanism to one involving upregulation of myoendothelial gap junctions and intermediate conductance calcium-activated potassium channel activity and expression. This study investigates the changes in NO-mediated mechanisms that accompany this shift. In saphenous arteries from controls fed a normal chow diet, acetylcholine-mediated endothelium-dependent vasodilation was blocked by NO synthase and soluble guanylyl cyclase inhibitors, but in equivalent arteries from obese animals sensitivity to these agents was reduced. The expression of endothelial NO synthase (eNOS) and caveolin-3 in rat saphenous arteries was unaffected by obesity, whilst that of caveolin-1 monomer and large oligomeric complexes of caveolins-1 and -2 were increased in membrane-enriched samples. The density of caveolae was increased at the membrane and cytoplasm of endothelial and smooth muscle cells of saphenous arteries from obese rats. Dissociation of eNOS from caveolin-1, as a prerequisite for activation of the enzyme, may be compromised and thereby impair NO-mediated vasodilation in the saphenous artery from diet-induced obese rats. Such altered signaling mechanisms in obesity-related vascular disease represent significant potential targets for therapeutic intervention.

Enhanced contractility in pregnancy is associated with augmented TRPC3, L-type, and T-type voltage-dependent calcium channel function in rat uterine radial artery.

In pregnancy, α-adrenoceptor-mediated vasoconstriction is augmented in uterine radial arteries and is accompanied by underlying changes in smooth muscle (SM) Ca(2+) activity. This study aims to determine the Ca(2+) entry channels associated with altered vasoconstriction in pregnancy, with the hypothesis that augmented vasoconstriction involves transient receptor potential canonical type-3 (TRPC3) and L- and T-type voltage-dependent Ca(2+) channels. Immunohistochemistry showed TRPC3, L-type Cav1.2 (as the α1C subunit), T-type Cav3.1 (α1G), and Cav3.2 (α1H) localization to the uterine radial artery SM. Fluorescence intensity of TRPC3, Cav1.2, and Cav3.2 was increased, and Cav3.1 decreased in radial artery SM from pregnant rats. Western blot analysis confirmed increased TRPC3 protein expression in the radial artery from pregnant rats. Pressure myography incorporating pharmacological intervention to examine the role of these channels in uterine radial arteries showed an attenuation of phenylephrine (PE)-induced constriction with Pyr3 {1-[4-[(2,3,3-trichloro-1-oxo-2-propen-1-yl)amino]phenyl]-5-(trifluoromethyl)-1H-pyrazole-4-carboxylic acid}-mediated TRPC3 inhibition or with nifedipine-mediated L-type channel block alone in vessels from pregnant rats; both effects of which were diminished in radial arteries from nonpregnant rats. Combined TRPC3 and L-type inhibition attenuated PE-induced constriction in radial arteries, and the residual vasoconstriction was reduced and abolished with T-type channel block with NNC 55-0396 in arteries from nonpregnant and pregnant rats, respectively. With SM Ca(2+) stores depleted and in the presence of PE, nifedipine, and NNC 55-0396, blockade of TRPC3 reversed PE-induced constriction. These data suggest that TRPC3 channels act synergistically with L- and T-type channels to modulate radial artery vasoconstriction, with the mechanism being augmented in pregnancy.

Pregnancy-induced remodelling and enhanced endothelium-derived hyperpolarization-type vasodilator activity in rat uterine radial artery: transient receptor potential vanilloid type 4 channels, caveolae and myoendothelial gap junctions.

In pregnancy, the vasculature of the uterus undergoes rapid remodelling to increase blood flow and maintain perfusion to the fetus. The present study determines the distribution and density of caveolae, transient receptor potential vanilloid type 4 channels (TRPV4) and myoendothelial gap junctions, and the relative contribution of related endothelium-dependent vasodilator components in uterine radial arteries of control virgin non-pregnant and 20-day late-pregnant rats. The hypothesis examined is that specific components of endothelium-dependent vasodilator mechanisms are altered in pregnancy-related uterine radial artery remodelling. Conventional and serial section electron microscopy were used to determine the morphological characteristics of uterine radial arteries from control and pregnant rats. TRPV4 distribution and expression was examined using conventional confocal immunohistochemistry, and the contribution of endothelial TRPV4, nitric oxide (NO) and endothelium-derived hyperpolarization (EDH)-type activity determined using pressure myography with pharmacological intervention. Data show outward hypertrophic remodelling occurs in uterine radial arteries in pregnancy. Further, caveolae density in radial artery endothelium and smooth muscle from pregnant rats was significantly increased by ~94% and ~31%, respectively, compared with control, whereas caveolae density did not differ in endothelium compared with smooth muscle from control. Caveolae density was significantly higher by ~59% on the abluminal compared with the luminal surface of the endothelium in uterine radial artery of pregnant rats but did not differ at those surfaces in control. TRPV4 was present in endothelium and smooth muscle, but not associated with internal elastic lamina hole sites in radial arteries. TRPV4 fluorescence intensity was significantly increased in the endothelium and smooth muscle of radial artery of pregnant compared with control rats by ~2.6- and 5.5-fold, respectively. The TRPV4 signal was significantly higher in the endothelium compared with the smooth muscle in radial artery of both control and pregnant rats, by ~5.7- and 2.7-fold, respectively. Myoendothelial gap junction density was significantly decreased by ~37% in radial artery from pregnant compared with control rats. Pressure myography with pharmacological intervention showed that NO contributes ~80% and ~30%, and the EDH-type component ~20% and ~70% of the total endothelium-dependent vasodilator response in radial arteries of control and pregnant rats, respectively. TRPV4 plays a functional role in radial arteries, with a greater contribution in those from pregnant rats. The correlative association of increased TRPV4 and caveolae density and role of EDH-type activity in uterine radial artery of pregnant rats is suggestive of their causal relationship. The decreased myoendothelial gap junction density and lack of TRPV4 density at such sites is consistent with their having an integral, albeit complex, interactive role in uterine vascular signalling and remodelling in pregnancy.

Myoendothelial contacts, gap junctions, and microdomains: anatomical links to function?

In several species and in many vascular beds, ultrastructural studies describe close contact sites between the endothelium and smooth muscle of <∼20nm. Such sites are thought to facilitate the local action of signaling molecules and/or the passage of current, as metabolic and electrical coupling conduits between the arterial endothelium and smooth muscle. These sites have the potential for bidirectional communication between the endothelium and smooth muscle, as a key pathway for coordinating vascular function. The aim of this brief review is to summarize the literature on the ultrastructural anatomy and distribution of key components of MECC sites in arteries. In addition to their traditional role of facilitating electrical coupling between the two cell layers, data on the role of MECC sites in arteries, as signaling microdomains involving a spatial localization of channels, receptors and calcium stores are highlighted. Diversity in the density and specific characteristics of MECC sites as signaling microdomains suggests considerable potential for functional diversity within and between arteries in health and disease.

Analysis of real-time serotonin (5-HT) availability during experimental colitis in mouse.

Serotonin (5-HT)-containing enterochromaffin (EC) cells of the intestine transduce chemical and mechanical stimuli from the intestinal lumen by releasing 5-HT on to afferent nerve terminals. Dysfunctional mucosal 5-HT signaling has been implicated in heightened visceral sensitivity and altered motility in patients with inflammatory bowel disease and in animal models. Our aim was to characterize the release and uptake of 5-HT in the mouse dextran sulfate sodium (DSS; 5% wt/vol) model of colitis. We made electrochemical recordings and used an ELISA assay to determine mucosal 5-HT release and uptake in untreated mice and mice with DSS-induced colitis. Peak and steady-state 5-HT concentrations were measured before and during blockade of the serotonin reuptake transporter (SERT) with 1 microM fluoxetine. Electrochemical recordings showed that colons from DSS-treated mice had roughly twice the steady-state levels of extracellular 5-HT and compression-evoked 5-HT release compared with untreated mice. Fluoxetine doubled the compression-evoked and steady-state 5-HT levels in control and DSS mice. These data were supported by ELISA assays, which showed enhanced 5-HT release during colitis, by immunohistochemical analyses, which showed increases in EC cell numbers, and by real-time PCR, which identified a decrease in SERT mRNA expression in the mucosa during colitis. These data are the first to demonstrate 5-HT release close to its release site and near its site of action during DSS-colitis. We conclude that DSS-colitis increases 5-HT availability primarily by an increase in the numbers of EC cells and/or of content of 5-HT in these EC cells.

Blockade of Pannexin-1 Channels and Purinergic P2X7 Receptors Shows Protective Effects Against Cytokines-Induced Colitis of Human Colonic Mucosa.

Introduction: The pannexin-1 (Panx1) channels are found in many cell types, and ATP released from these channels can act on nearby cells activating purinergic P2X7 receptors (P2X7R) which lead to inflammation. Although Panx1 and P2X7R are implicated in the process of inflammation and cell death, few studies have looked at the role they play in inflammatory bowel disease in human. Hence, the aim of the present study was to investigate the function of Panx1 and P2X7R in an ex vivo colitis model developed from human colonic mucosal explants. Materials and Methods: Healthy human colonic mucosal strips (4 × 10 mm) were incubated in carbogenated culture medium at 37°C for 16 h. Proinflammatory cytokines TNFα and IL-1ß (each 10 ng/mL) were used to induce colitis in mucosal strips, and the effects of Panx1 and P2X7R on cytokines-induced tissue damage were determined in the presence of the Panx1 channel blocker 10Panx1 (100 µM) and P2X7R antagonist A438079 (100 µM). The effects of 10Panx1 and A438079 on cytokines-enhanced epithelial permeability were also studied using Caco-2 cells. Results: Histological staining showed that the mucosal strips had severe structural damage in the cytokines-only group but not in the incubation-control group (P < 0.01). Compared to the cytokines-only group, crypt damage was significantly decreased in groups receiving cytokines with inhibitors (10Panx1, A438079, or 10Panx1 + A438079, P < 0.05). The immunoreactive signals of tight junction protein zonula occludens-1 (ZO-1) were abundant in all control tissues but were significantly disrupted and lost in the cytokines-only group (P < 0.01). The diminished ZO-1 immunoreactivity induced by cytokines was prevented in the presence of 10Panx1 (P = 0.04). Likewise, 10Panx1 significantly attenuated the cytokines-evoked increase in paracellular permeability of Caco-2 cells. Although the inhibition of P2X7R activity by A438079 diminished cytokines-induced crypt damage, its effect on the maintenance of ZO-1 immunoreactivity and Caco-2 epithelial cell integrity was less evident. Conclusion: The blockade of Panx1 and P2X7R reduced the inflammatory cytokines-induced crypt damage, loss of tight junctions and increase in cell permeability. Thus, Panx1 and P2X7R may have roles in causing mucosal damage, a common clinical feature of inflammatory bowel disease.

Esophageal Dysmotility in Patients following Total Laryngectomy.

Objectives Dysphagia is common in total laryngectomees, with some symptoms suggesting esophageal dysmotility. Tracheoesophageal (TE) phonation requires effective esophagopharyngeal air passage. Hence, esophageal dysmotility may affect deglutition or TE phonation. This study aimed to determine (1) the characteristics of esophageal dysmotility in laryngectomees, (2) whether clinical history is sensitive in detecting esophageal dysmotility, and (3) the relationship between esophageal dysmotility and TE prosthesis dysfunction. Study Design Multidisciplinary cross-sectional study. Setting Tertiary academic hospital. Subjects and Methods For 31 participants undergone total laryngectomy 1 to 12 years prior, clinical histories were taken by a gastroenterologist and a speech pathologist experienced in managing dysphagia. Esophageal high-resolution manometry was performed and analyzed using Chicago Classification v3.0. Results Interpretable manometric studies were obtained in 23 (1 normal manometry). Esophageal dysmotility patterns included achalasia, esophagogastric junction outflow obstruction, diffuse esophageal spasm, and other major (30%) and minor (50%) peristaltic disorders. The sensitivity of predicting any esophageal dysmotility was 28%, but it is noteworthy that patients with achalasia and diffuse esophageal spasm (DES) were predicted. Two of 4 participants with TE puncture leakage had poor esophageal clearance. Of 20 TE speakers, 12 had voice problems, no correlation between poor voice, and any dysmotility pattern. Conclusions Peristaltic and lower esophageal sphincter dysfunction are common in laryngectomees. Clinical history, while not predictive of minor motor abnormalities, predicted correctly cases with treatable spastic motor disorders. Dysmotility was not associated with poor phonation, although TE puncture leakage might be linked to poor esophageal clearance. Esophageal dysmotility should be considered in the laryngectomees with persisting dysphagia or leaking TE puncture.

Teaching basic gastrointestinal physiology using classic papers by Dr. Walter B. Cannon.

The movements of the gastrointestinal tract, as described by Walter B. Cannon 100 years ago, reveal much about the functions of this unique organ and how it is controlled by the body. Two classic papers by Cannon provide a rare glimpse into the hidden functions of the body and give students a great example of the scientific method in action. In this essay, we describe the basic movements and functions of the gastrointestinal tract as revealed by X-rays, contrast meals, and Cannon's careful observations. It is Cannon's experimental care and obvious interest in his subject matter that can provide your students with a taste of the excitement of discovery and insight into the processes by which experimental science moves forward.

Purinergic Signaling in Gut Inflammation: The Role of Connexins and Pannexins.

Purinergic receptors play an important role in inflammation, and can be activated by ATP released via pannexin channels and/or connexin hemichannels. The purinergic P2X7 receptor (P2X7R) is of interest since it is involved in apoptosis when activated. Most studies focus on the influence of pannexin-1 (Panx1) and connexin 43 (Cx43) on ATP release and how it affects P2X7R function during inflammation. Inflammatory bowel disease (IBD) is characterized by uncontrolled inflammation within the gastrointestinal system. At present, the pathophysiology of this disease remains largely unknown but it may involve the interplay between P2X7R, Panx1, and Cx43. There are two main types of IBD, ulcerative colitis and Crohn's disease, that are classified by their location and frequency of inflammation. Current research suggests that alterations to normal functioning of innate and adaptive immunity may be a factor in disease progression. The involvement of purinergic receptors, connexins, and pannexins in IBD is a relatively novel notion in the context of gastrointestinal inflammation, and has been explored by various research groups. Thus, the present review focuses on the current research involving connexins, pannexins, and purinergic receptors within the gut and enteric nervous system, and will examine their involvement in inflammation and the pathophysiology of IBD.

Influence of Tryptophan and Serotonin on Mood and Cognition with a Possible Role of the Gut-Brain Axis.

The serotonergic system forms a diffuse network within the central nervous system and plays a significant role in the regulation of mood and cognition. Manipulation of tryptophan levels, acutely or chronically, by depletion or supplementation, is an experimental procedure for modifying peripheral and central serotonin levels. These studies have allowed us to establish the role of serotonin in higher order brain function in both preclinical and clinical situations and have precipitated the finding that low brain serotonin levels are associated with poor memory and depressed mood. The gut-brain axis is a bi-directional system between the brain and gastrointestinal tract, linking emotional and cognitive centres of the brain with peripheral functioning of the digestive tract. An influence of gut microbiota on behaviour is becoming increasingly evident, as is the extension to tryptophan and serotonin, producing a possibility that alterations in the gut may be important in the pathophysiology of human central nervous system disorders. In this review we will discuss the effect of manipulating tryptophan on mood and cognition, and discuss a possible influence of the gut-brain axis.

TRPV3 expression and vasodilator function in isolated uterine radial arteries from non-pregnant and pregnant rats.

This study investigated the expression and function of transient receptor potential vanilloid type-3 ion channels (TRPV3) in uterine radial arteries isolated from non-pregnant and twenty-day pregnant rats. Immunohistochemistry (IHC) suggested TRPV3 is primarily localized to the smooth muscle in arteries from both non-pregnant and pregnant rats. IHC using C' targeted antibody, and qPCR of TRPV3 mRNA, suggested pregnancy increased arterial TRPV3 expression. The TRPV3 activator carvacrol caused endothelium-independent dilation of phenylephrine-constricted radial arteries, with no difference between vessels from non-pregnant and pregnant animals. Carvacrol-induced dilation was reduced by the TRPV3-blockers isopentenyl pyrophosphate and ruthenium red, but not by the TRPA1 or TRPV4 inhibitors HC-030031 or HC-067047, respectively. In radial arteries from non-pregnant rats only, inhibition of NOS and sGC, or PKG, enhanced carvacrol-mediated vasodilation. Carvacrol-induced dilation of arteries from both non-pregnant and pregnant rats was prevented by the IKCa blocker TRAM-34. TRPV3 caused an endothelium-independent, IKCa-mediated dilation of the uterine radial artery. NO-PKG-mediated modulation of TRPV3 activity is lost in pregnancy, but this did not alter the response to carvacrol.

ATP as a putative sensory mediator: activation of intrinsic sensory neurons of the myenteric plexus via P2X receptors.

The mucosal terminals of sensory neurons intrinsic to the wall of the intestine are sensitive to the chemical environment within the lumen. Lumenal stimuli probably release sensory mediators from the mucosal epithelium, which then activate the nerve terminals indirectly. Here, we tested the idea that ATP activates intrinsic sensory nerve terminals in a way consistent with its being a sensory mediator. We made intracellular recordings from intrinsic sensory neurons located in the myenteric plexus [identified as AH neurons, which are neurons with a long-lasting afterhyperpolarization following the action potential (AP)], located within 1 mm of intact mucosa. Focal electrical stimulation of the mucosa was used to locate and map regions innervated by each neuron. Application of ATP (1-2 mm in the pressure pipette) to these regions elicited trains of APs that originated at the sensory terminals. ATP-gamma-S produced a similar response, but alpha,beta-methylene ATP and 2-methylthio-ATP were only weakly active. The P2 receptor antagonist pyridoxalphosphate-6-azophenyl-2',5'-disulphonic acid (PPADS) (60 microm in the bath) abolished the APs evoked by ATP and ATP-gamma-S but spared similar responses evoked by 5-hydroxytryptamine (5-HT). Another P2 receptor antagonist suramin (100 microm in the bath) did not significantly change the number of APs evoked by ATP. Either ATP or alpha,beta-methylene ATP desensitized the ATP-evoked APs; 50% recovery occurred after approximately 5 sec. The number of APs evoked by ATP was reduced, but not abolished, by the selective 5-HT3 receptor antagonist granisetron (1 microm in the bath). ATP was applied to the cell bodies of sensory neurons to investigate whether the cell bodies express the same P2X receptor as the terminals. ATP evoked a fast depolarization associated with a reduction in input resistance and a reversal potential of -11 mV. This depolarization was potentiated by suramin and blocked by PPADS. We conclude that activation of an atypical excitatory P2X receptor by ATP triggers AP generation in the mucosal processes of the sensory neurons; endogenous 5-HT release may also contribute to activation of the nerve terminals. A similar P2X receptor exists on the cell body of the sensory neuron. Together, these data are consistent with a role for ATP as a sensory mediator in gastrointestinal chemosensory transduction.

The effect of low estrogen state on serotonin transporter function in mouse hippocampus: a behavioral and electrochemical study.

Defects in serotonergic transmission, including serotonin transporter (SERT) function, have been implicated in depression, anxiety disorders and some aspects of schizophrenia. The sex steroid hormone estrogen is known to modulate functional SERT activity, but whether it is up- or down-regulated is unclear. The aim of the present study was to examine the effect of a low estrogen state in mice on the behavioral effect of drugs acting through the SERT, serotonin uptake kinetics and SERT density in the hippocampus. We compared control mice, ovariectomized (OVX) C57BL/6J mice and aromatase knockout (ArKO) mice that are unable to produce estrogen. Fluoxetine treatment, but not fenfluramine treatment, significantly increased prepulse inhibition (PPI), a measure of sensorimotor gating, in C57BL/6J mice. The effect of fluoxetine was greater in OVX compared to sham-operated mice. In ArKO and J129 wild-type mice, fluoxetine increased PPI to the same extent while fenfluramine increased PPI more in ArKO mice compared to controls. Measurement of the time-course for diffusion and reuptake of exogenous serotonin in the CA3 region of the hippocampus showed that, in OVX mice, the fluoxetine-induced slowing of signal decay after application of serotonin was enhanced when compared to sham-operated controls. Similarly, in ArKO mice, the effect of fluoxetine was enhanced, suggesting that SERT function was greater than in J129 wild-type controls. Measurement of SERT density by [3H]-citalopram autoradiography, revealed an 18% decrease in hippocampus of OVX mice compared to intact controls. SERT density was also significantly reduced in nucleus accumbens (26%) but not in other regions, such as the raphe nuclei. Together, these results suggest that a low estrogen state increases SERT activity in the hippocampus despite an apparent reduction in SERT density. The behavioral consequences of these changes depend on the model of estrogen state used.

Short term exendin-4 treatment reduces markers of metabolic disorders in female offspring of obese rat dams.

OBJECTIVES: Maternal obesity imposes significant health risks in the offspring including diabetes and dyslipidemia. We previously showed that the hypoglycaemic agent exendin-4 (Ex-4) administered from weaning can reverse the maternal impact of 'transmitted disorders' in such offspring. However daily injection for six-weeks was required and the beneficial effect may lapse upon drug withdrawal. This study aimed to investigate whether short term Ex-4 treatment during suckling period in a rodent model can reverse transmitted metabolic disorders due to maternal obesity. METHODS: Maternal obesity was induced in female Sprague Dawley rats by high-fat diet feeding for 6 weeks, throughout gestation and lactation. Female offspring were treated with Ex-4 (5µg/kg/day) between postnatal day (P) 4 and 14. Female offspring were harvested at weaning (P20). Lipid and glucose metabolic markers were measured in the liver and fat. Appetite regulators were measured in the plasma and hypothalamus. RESULTS: Maternal obesity significantly increased body weight, fat mass, and liver weight in the offspring. There was an associated inhibition of peroxisomal proliferator activated receptor gamma coactivator 1α (PGC1α), increased fatty acid synthase (FASN) expression in the liver, and reduced adipocyte triglyceride lipase (ATGL) expression. It also increased the plasma gut hormone ghrelin and reduced glucagon-like peptide-1. Ex-4 treatment partially reversed the maternal impact on adiposity and impaired lipid metabolism in the offspring, with increased liver PGC1α and inhibition of FASN mRNA expression. Ex-4 treatment also increased the expression of a novel fat depletion gene a2-zinc-glycoprotein 1 in the fat tissue. CONCLUSION: Short term Ex-4 treatment during the suckling period significantly improved the metabolic profile in the offspring from the obese mothers at weaning. Long-term studies are needed to follow such offspring to adulthood to examine the sustained effects of Ex-4 in preventing the development of metabolic disease.