Objectives:The objective of this study was to investigate comparatively the influence of proton-pump inhibitors (PPI) administration on three bacterial communities in the oral cavity, stomach, and colon along the alimentary tract.Methods:Forty-five subjects including 18 patients taking PPI were enrolled. Stimulated saliva, gastric fluid (GF), and feces were obtained from each subject for the microbiota analysis through bacterial 16S rRNA gene profiling using the pyrosequencing method.Results:The species richness (alpha diversity) was similar among these three microbiota, whereas the interindividual diversity (beta diversity) was much higher in the fecal microbiota compared with that in the others. The UniFrac analysis indicated that the salivary and GF microbiota were similar to one another; however, both differed greatly from the fecal microbiota in the overall bacterial community structure. In the comparison between PPI-users and PPI-nonusers, a bacterial cell number increase of ~1,000 times was found in the GF of PPI-users using culturing methods, whereas the bacterial number and composition were nearly identical between the two groups using quantitative PCR and a similarity search based on 16S profiling. The beta diversity significantly increased in both the salivary and GF microbiota of PPI-users compared with PPI-nonusers.Conclusions:These results suggest that the GF microbiota has recently moved from the saliva. Bacterial overgrowth in the GF by PPI administration may be due to a lack of killing rather than proliferation of the bacteria in the acid-suppressed stomach. The biological significance of the increase in beta diversity by PPI administration remains unclear.

ObjectiveTo investigate the structure of the gastric microbiota in functional dyspepsia (FD) and its role in the pathophysiology.DesignWe compared the basic physiological properties of the gastric fluid (GF) and the structure of the microbiota in the GF of 44 healthy control (HC) participants and 44 patients with FD. We then treated the patients with FD with a yogurt containing a probiotic strain of Lactobacillus gasseri OLL2716 (LG21 yogurt) and investigated the effects on the bacteriological parameters and symptoms to examine the relationship between them.ResultsThe volume of GF recovered from the stomach after overnight fasting was greater in the patients with FD than in the HCs, and decreased in the patients with FD whose symptoms were improved by the LG21 yogurt treatment. An analysis using a terminal restriction fragment polymorphism method demonstrated that the overall structure of the bacterial community and the abundance of genus Prevotella in the GF of the patients in the FD group were significantly different from those in the HC group. In the patients with FD, this bacteriological change was restored by treatment with LG21 yogurt. A significant inverse correlation was found between the abundance of Prevotella and the severity of postprandial distress-like symptoms in patients with FD who received LG21 yogurt.ConclusionsSignificant dysbiosis was found in the GF microbiota of patients with FD and considered to be involved in the pathogenesis. The abundance of genus Prevotella in the GF may be used as a biomarker of the efficacy of the treatment of FD.Trial registration numberUMINCTR000022026.

ObjectiveTo investigate the influence of antimicrobials on both the gut microbiota structure and the plasma ghrelin level using Helicobacter pylori-infected patients who underwent eradication therapy.DesignTwenty H. pylori-infected patients (mean age 68.3 years old) who underwent eradication therapy participated in the study. For the therapy, patients had 1 week of triple therapy consisting of amoxicillin, clarithromycin and proton-pump inhibitors. Stool and blood samples were obtained before (S1), immediately after (S2) and/or 3 months after (S3) the therapies. The concentrations of ghrelin and leptin in the blood were assayed using an ELISA. The V3-V4 region of the 16S rRNA gene was amplified using bacterial DNA from the stool, and about 50 000 high-quality amplicons per sample were grouped into operational taxonomic units for bacteriological analyses.ResultsThe Bacteroidetes:Firmicutes (B:F) ratio was significantly greater at S3 than S1 (P<0.01). This increase in the B:F ratio between S3 and S1 was found in 15 out of 20 patients. A significant decrease in the concentration of active ghrelin (P=0.003) in the plasma was observed between S3 and S1. There was a statistically significant correlation between the rate of patients whose B:F ratio increased and that of patients whose active ghrelin level decreased between S3 and S1 according to Fisher’s exact probability test (P=0.03).ConclusionsChanges in the gut microbiota, such as the B:F ratio after treatment with antimicrobials, might cause a change in the plasma ghrelin level, as the direct and earliest target of antimicrobials would be the microbiota rather than the hormone-secreting system.

Ventral tegmental area (VTA) dopaminergic neurons are key components of the reward pathway, and their activity is powerfully controlled by a diverse array of inhibitory GABAergic inputs. Two major sources of GABAergic nerve terminals within the VTA are local VTA interneurons and neurons in the rostromedial tegmental nucleus (RMTg). Here, using optogenetics, we compared synaptic properties of GABAergic synapses on VTA dopamine neurons using selective activation of afferents that originate from these two cell populations. We found little evidence of co-release of glutamate from either input, but RMTg-originating synaptic currents were reduced by strychnine, suggesting co-release of glycine and GABA. VTA-originating synapses displayed a lower initial release probability, and at higher frequency stimulation, short-term depression was more marked in VTA- but not RMTg-originating synapses. We previously reported that nitric oxide (NO)-induced potentiation of GABAergic synapses on VTA dopaminergic cells is lost after exposure to drugs of abuse or acute stress; in these experiments, multiple GABAergic afferents were simultaneously activated by electrical stimulation. Here we found that optogenetically-activated VTA-originating synapses on presumptive dopamine neurons also exhibited NO-induced potentiation, whereas RMTg-originating synapses did not. Despite providing a robust inhibitory input to the VTA, RMTg GABAergic synapses are most likely not those previously shown by our work to be persistently altered by addictive drugs and stress. Our work emphasises the idea that dopamine neuron excitability is controlled by diverse inhibitory inputs expected to exert varying degrees of inhibition and to participate differently in a range of behaviours.

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The trigeminal nucleus caudalis ( TN c) receives extensive afferent innervation from peripheral sensory neurons of the trigeminal ganglion (TG), and is the first central relay in the circuitry underpinning orofacial pain. Despite the initial characterization of the neurons in the superficial laminae, many questions remain. Here we report on electrophysiological properties of 535 superficial lamina I/ II TN c neurons. Based on their firing pattern, we assigned these cells to five main groups, including (1) tonic, (2) phasic, (3) delayed, (4) H‐current, and (5) tonic‐phasic neurons, groups that exhibit distinct intrinsic properties and share some similarity with groups identified in the spinal dorsal horn. Driving predominantly nociceptive TG primary afferents using optogenetic stimulation in TRPV 1/ChR2 animals, we found that tonic and H‐current cells are most likely to receive pure monosynaptic input, whereas delayed neurons are more likely to exhibit inputs that appear polysynaptic. Finally, for the first time in TN c neurons, we used unsupervised clustering analysis methods and found that the kinetics of the action potentials and other intrinsic properties of these groups differ significantly from one another. Unsupervised spectral clustering based solely on a single voltage response to rheobase current was sufficient to group cells with shared properties independent of action potential discharge pattern, indicating that this approach can be effectively applied to identify functional neuronal subclasses. Together, our data illustrate that cells in the TN c with distinct patterns of TRPV 1/ChR2 afferent innervation are physiologically diverse, but can be understood as a few major groups of cells having shared functional properties.

In patients with functional upper gastrointestinal disorders such as gastroesophageal reflux disease and functional dyspepsia, the presence of symptoms is thought to occur in the absence of any organic diseases and the mechanisms behind this remain unclear. We therefore examined the relationship between stomach-related biomarker levels and symptoms. Twenty-four outpatients who had taken proton-pump inhibitors every day were enrolled in this study. The subjects consumed yogurt containing 109 colony-forming units of Lactobacillus gasseri OLL2716 (LG21) every day for three months. They underwent four clinical examinations in total. Each examination consisted of answering a questionnaire with a frequency scale for the symptoms of GERD (FSSG), and included measurements of the serum gastrin, ghrelin, and pepsinogens I and II levels. As a result, the FSSG score and the PGI value showed a decrease and an increase, respectively, after LG21 treatment when analyzed without age adjustment. A multiple regression analysis with additional adjustments for gender and age revealed a strong association between the PGI value and the FSSG symptom scores. Therefore either the PGI level itself or the factors regulating the PGI level might be involved in the etiology of these symptoms.

The opsins have been studied extensively for their functions in visual phototransduction; however, the mechanisms underlying extraocular opsin signaling remain poorly understood. The first mammalian extraocular opsin to be discovered, opsin 3 (OPN3), was found in the brain more than two decades ago, yet its function remains unknown. A significant hindrance to studying OPN3 has been a lack of specific antibodies against mammalian OPN3, resulting in an incomplete understanding of its expression in the brain. Although Opn3 promoter-driven reporter mice have been generated to examine general OPN3 localization, they lack the regulated expression of the endogenous protein and the ability to study its subcellular localization. To circumvent these issues, we have created a knock-in OPN3-mCherry mouse model in which the fusion protein OPN3-mCherry is expressed Significance Statement Before the current report, there had been a significant lack of encephalic opsin 3 (OPN3) protein characterization, likely driven by the absence of murine OPN3 antibodies. This study describes a novel OPN3-mCherry knock-in mouse that we used to analyze endogenous OPN3 protein expression across the brain. We have uncovered new aspects of OPN3: localization to previously undocumented brain subregions, expression in GABAergic neurons and non-neuronal cells, and punctate subcellular localization in the soma. Our OPN3 expression map is an invaluable step toward discovering its elusive encephalic functions. The OPN3-mCherry mouse will facilitate investigation of OPN3 not only in the brain, but also across the entire organism, a useful feature as OPN3 is emerging as a possible mediator of phototransduction outside the brain.