Musclin prevents depression-like behavior in male mice by activating urocortin 2 signaling in the hypothalamus.

Introduction: Physical activity is recommended as an alternative treatment for depression. Myokines, which are secreted from skeletal muscles during physical activity, play an important role in the skeletal muscle-brain axis. Musclin, a newly discovered myokine, exerts physical endurance, however, the effects of musclin on emotional behaviors, such as depression, have not been evaluated. This study aimed to access the anti-depressive effect of musclin and clarify the connection between depression-like behavior and hypothalamic neuropeptides in mice. Methods: We measured the immobility time in the forced swim (FS) test, the time spent in open arm in the elevated-plus maze (EPM) test, the mRNA levels of hypothalamic neuropeptides, and enumerated the c-Fos-positive cells in the paraventricular nucleus (PVN), arcuate nucleus (ARC), and nucleus tractus solitarii (NTS) in mice with the intraperitoneal (i.p.) administration of musclin. Next, we evaluated the effects of a selective corticotropin-releasing factor (CRF) type 1 receptor antagonist, selective CRF type 2 receptor antagonist, melanocortin receptor (MCR) agonist, and selective melanocortin 4 receptor (MC4R) agonist on changes in behaviors induced by musclin. Finally we evaluated the antidepressant effect of musclin using mice exposed to repeated water immersion (WI) stress. Results: We found that the i.p. and i.c.v. administration of musclin decreased the immobility time and relative time in the open arms (open %) in mice and increased urocortin 2 (Ucn 2) levels but decreased proopiomelanocortin levels in the hypothalamus. The numbers of c-Fos-positive cells were increased in the PVN and NTS but decreased in the ARC of mice with i.p. administration of musclin. The c-Fos-positive cells in the PVN were also found to be Ucn 2-positive. The antidepressant and anxiogenic effects of musclin were blocked by central administration of a CRF type 2 receptor antagonist and a melanocortin 4 receptor agonist, respectively. Peripheral administration of musclin also prevented depression-like behavior and the decrease in levels of hypothalamic Ucn 2 induced by repeated WI stress. Discussion: These data identify the antidepressant effects of musclin through the activation of central Ucn 2 signaling and suggest that musclin and Ucn 2 can be new therapeutic targets and endogenous peptides mediating the muscle-brain axis.

Mumefural prevents insulin resistance and amyloid-beta accumulation in the brain by improving lowered interstitial fluid pH in type 2 diabetes mellitus.

The present study tried to clarify if mumefural would prevent hyperglycemia, one of the typical symptoms of type 2 diabetes mellitus (T2DM), since mumefural is an extract from Japanese apricots preventing hyperglycemia. To clarify if mumefural would prevent T2DM pathogenesis, we used Otsuka Long-Evans Tokushima fatty (OLETF) rats, T2DM model. Mumefural diminished hyperglycemia, HOMA-IR and plasma triglyceride concentration in OLETF rats under fasting conditions. In addition, mumefural elevated protein expression of sodium-coupled monocarboxylate transporter 1 (SMCT1) in the distal colon participating in absorption of weak organic acids, which behave as bases but not acids after absorption into the body. Mumefural also increased the interstitial fluid pH around the brain hippocampus lowered in OLETF rats compared with non-T2DM LETO rats used as control for OLETF rats. Amyloid-beta accumulation in the brain decreased in accordance with the pH elevation. On the one hand, mumefural didn't affect plasma concentrations of glucagon, GLP-1, GIP or PYY under fasting conditions. Taken together, these observations indicate that: 1) mumefural would be a useful functional food improving hyperglycemia, insulin resistance and the lowered interstitial fluid pH in T2DM; 2) the interstitial fluid pH would be one of key factors influencing the accumulation of amyloid-beta.

Intraperitoneal administration of nesfatin­1 stimulates glucagon­like peptide­1 secretion in fasted mice.

Increasing endogenous secretion of glucagon­like peptide (GLP)­1 is considered a promising therapeutic approach for type 2 diabetes because decreased GLP­1 plasma concentrations have been observed in patients with this condition. Nesfatin­1, which is a central and peripheral anorexigenic peptide, has been reported to release GLP­1 from enteroendocrine STC­1 cells, although whether nesfatin­1 stimulates GLP­1 secretion in vivo remains to be elucidated. Previous studies have indicated that nesfatin­1 has glucose­lowering and insulinotropic effects in mice and rats; however, the in vivo mechanism remains unclear. The present study aimed to investigate whether peripheral administration of nesfatin­1 increased blood concentrations of GLP­1 and insulin in food­deprived mice. Nesfatin­1 was administered intraperitoneally to 18­h fasted mice. Plasma GLP­1 and insulin concentrations in the mice administered 2.5 µmol/kg nesfatin­1 were higher than those in saline­treated mice. Blood glucose concentrations in mice treated with 1.25 and 2.5 µmol/kg nesfatin­1 were lower than those in saline­treated mice. The mRNA expression of preproglucagon in mouse ilea after treatment with 1.25 µmol/kg nesfatin­1 was higher than that in saline­treated mice. The administration of 1.25 µmol/kg nesfatin­1 raised GLP­1 concentrations at 30 and 60 min and insulin concentrations at 30 and 60 min after injection. Furthermore, the higher level of nesfatin­1­induced insulin was diminished by pre­administration of anti­GLP­1 antiserum. Intraperitoneally administered nesfatin­1 increased insulin concentrations by accelerating GLP­1 secretion. The results are the first in vivo demonstration of promotion of GLP­1 secretion by nesfatin­1 in the mouse, suggesting the developmental potential of nesfatin­1 for GLP­1 release.

Secondary bile acid lithocholic acid attenuates neurally evoked ion transport in the rat distal colon.

The inhibitory action of the secondary bile acid lithocholic acid (LCA) on neurally evoked Cl-/HCO3- secretion was investigated using the Ussing-chambered mucosal-submucosal preparation from the rat distal colon. Electrical field stimulation (EFS) evoked cholinergic and noncholinergic secretory responses in the rat distal colon. The responses were almost completely blocked by TTX (10-6 M) but not atropine (10-5 M) or hexamethonium (10-4 M). The selective antagonist for VIP receptor 1 (VPAC1) greatly reduced the EFS-evoked response. Thus, the rat distal colon may be predominantly innervated by noncholinergic VIP secretomotor neurons. Basolateral addition of 6 × 10-5 M LCA inhibited the EFS-evoked response. The inhibitory action of LCA was partly rescued by the Y2R antagonist BIIE0246. The bile acid receptor TGR5 agonist INT-777 mimicked the LCA-induced inhibitory action. Immunohistochemical staining showed the colocalization of TGR5 and PYY on L cells. TGR5 immunoreactivity was also found in VIP-immunoreactive submucosal neurons which also expressed the PYY receptor, Y2R. These results suggest that LCA inhibits neurally evoked Cl-/HCO3- secretion through the activation of TGR5 on L cells and cholinergic- and VIP-secretomotor neurons in the submucosal plexus. Furthermore, the inhibitory mechanism may involve TGR5-stimulated PYY release from L cells and Y2R activation in VIP-secretomotor neurons.

Rubiscolin­6 rapidly suppresses the postprandial motility of the gastric antrum and subsequently increases food intake via δ­opioid receptors in mice.

Rubiscolin­6 is a food­derived opioid peptide found in Spinacia oleracea that has anti­nociceptive, memory­enhancing, anxiolytic­like and anti­depressant effects. Rubiscolin­6 has been reported to have two opposing effects on food intake. Food intake is closely connected to gut motility; however, to the best of our knowledge, the effect of rubiscolin­6 on gut motility has not been reported. The present study aimed to investigate the effect of rubiscolin­6 on postprandial motility of the gastric antrum in conscious mice. A catheter was implanted in the gastric antrum of male C57BL/6J mice. Manometric measurements were performed in fasted male mice and chow was then provided to assess motility in the fed state. Rubiscolin­6, the δ­opioid receptor antagonist naltrindole, a mixture of rubiscolin­6 and naltrindole, or vehicle was administered intraperitoneally 30 min after eating. The percentage motor index (%MI) was then calculated. Cumulative food intake was measured in both ad libitum­fed and overnight­fasted mice. The %MI was significantly lower in mice treated with rubiscolin­6 compared with that in the other groups, but normalized by treatment with the rubiscolin­6/naltrindole mixture. The decrease in %MI induced by rubiscolin­6 remained for 1 h after administration. Cumulative food intake was significantly higher 4 and 6 h after rubiscolin­6 administration in ad libitum­fed mice but was normalized by the rubiscolin­6/naltrindole mixture. Food intake 30 min after rubiscolin­6 administration was normal, but was higher in mice treated with the rubiscolin­6/naltrindole mixture. Thus, rubiscolin­6 may have a rapid effect to reduce postprandial antral motility and may subsequently increase food intake after this inhibitory effect disappears. These effects were revealed to be mediated through δ­opioid receptors. The orexigenic effect of rubiscolin­6 may be applicable to the treatment of anorexia and cachexia.

Relationship between adiponectin multimer levels and subtypes of cerebral infarction.

AIM: Serum adiponectin levels are decreased in patients with cerebral infarction. Adiponectin in circulation exists in three isoforms: high molecular weight (HMW), medium molecular weight (MMW), and low molecular weight (LMW) adiponectin. We measured serum levels of total adiponectin and adiponectin multimers (HMW, MMW, and LMW) in patients with cerebral infarction and compared the serum levels of the three adiponectin multimers in stroke subtypes. We also evaluated the clinical value of adiponectin multimer levels as a biomarker for cerebral infarction. METHODS: We assessed a total of 132 patients with cerebral infarctions. The serum levels of total and adiponectin multimers were measured using enzyme-linked immunosorbent assay (ELISA). RESULTS: The total and HMW adiponectin levels were significantly lower in atherothrombotic infarction (AI) than in cerebral embolism (CE) (total, p < 0.05; HMW, p < 0.05). In male patients, the MMW adiponectin level was significantly lower in the lacunar infarction (LI) group than in the AI group (p < 0.05). The LMW adiponectin level was significantly lower in the AI group than in the LI and CE groups (LI, p < 0.001; CE, p = 0.001). However, there were no significant differences in adiponectin multimer levels among the stroke subtypes in female subjects. Additionally, in female patients with AI and LI, the LMW adiponectin levels were negatively associated with C-reactive protein (CRP; AI, p < 0.05; LI, p < 0.05). CONCLUSION: These findings suggest that a decrease in adiponectin is associated with AI and that serum LMW adiponectin level represents a potential biomarker for AI.

Minimum biological domain of xenin-25 required to induce anion secretion in the rat ileum.

Xenin-25 has a variety of physiological functions in the gastrointestinal tract, including ion transport and motility. Xenin-25 and neurotensin show sequence homology, especially near their C-terminal regions. The sequence similarity between xenin-25 and neurotensin indicates that the effects of xenin-25 is mediated by the neurotensin receptor but some biological actions of xenin-25 are independent. We have previously reported that xenin-25 modulates intestinal ion transport and colonic smooth muscle activity. However, minimal biological domain of xenin-25 to induce ion transport was not clear. To improve the mechanistic understanding of xenin-25 and to gain additional insights into the functions of xenin-25, the present study was designed to determine the minimal biological domain of xenin-25 required for ion transport in the rat ileum using various truncated xenin fragments and analogues in an Ussing chamber system. The present results demonstrate that the minimum biological domain of xenin-25 to induce Cl-/HCO3- secretion in the ileum contains the C-terminal pentapeptide. Furthermore, Arg at position 21 is important to retain the biological activity of xenin-25 and induces Cl-/HCO3- secretion in the rat ileum.

Men's lacrosse protective equipment increases strain during exercise in the heat.

OBJECTIVES: The purpose of this study was to determine thermoregulatory and cardiovascular effects of wearing men's lacrosse protective equipment during simulated lacrosse activities in the heat. DESIGN: We conducted a randomized, controlled, crossover study. METHODS: Thirteen healthy men (22 ±â€¯3 y, 76.2 ±â€¯8.9 kg, 181 ±â€¯6 cm, 16.06 ±â€¯6.16% body fat) completed two matched exercise trials in the heat (WBGT: 25.5 ±â€¯0.8 °C). In randomized order, participants donned full men's lacrosse equipment (helmet, shoulder/elbow pads, and gloves) in one trial while the other included no equipment. Participants completed a topography body scan to determine specific body surface area covered with equipment. Rectal temperature (Tre), heart rate (HR), and mean weighted skin temperature (Tsk) were measured throughout trials. Whole body sweat rate was assessed for trial comparisons. RESULTS: The equipment covered 32.62 ±â€¯2.53% body surface area in our participants. Post-exercise Tre was significantly greater with equipment (39.36 ±â€¯0.04 °C) compared to control (38.98 ±â€¯0.49 °C; p = .007). The overall rate of rise of Tre was significantly greater with equipment (0.043 ±â€¯0.015 °C·min-1) compared to control (0.031 ±â€¯0.008 °Cmin-1; p = .041). Regardless of time point, HR and Tsk were significantly elevated with equipment compared to control trial (p ≤ .026). Sweat rates were elevated with equipment (1.76 ±â€¯0.74 L·h-1) compared to shorts and t-shirt (1.13 ±â€¯0.26 L·h-1), but this difference was not significant (p = .058). CONCLUSIONS: Our data indicate impairments in heat dissipation and increased cardiovascular strain imposed by men's lacrosse equipment.

The effect of Xenin25 on spontaneous circular muscle contractions of rat distal colon in vitro.

Xenin25 has a variety of physiological functions in the Gastrointestinal (GI) tract, including ion transport and motility. However, the motility responses in the colon induced by Xenin25 remain poorly understood. Therefore, the effect of Xenin25 on the spontaneous circular muscle contractions of the rat distal colon was investigated using organ bath chambers and immunohistochemistry. Xenin25 induced the inhibition followed by postinhibitory spontaneous contractions with a higher frequency in the rat distal colon. This inhibitory effect of Xenin25 was significantly suppressed by TTX but not by atropine. The inhibitory time (the duration of inhibition) caused by Xenin25 was shortened by the NTSR1 antagonist SR48692, the NK1R antagonist CP96345, the VPAC2 receptor antagonist PG99-465, the nitric oxide-sensitive guanylate-cyclase inhibitor ODQ, and the Ca2+ -dependent K+ channel blocker apamin. The higher frequency of postinhibitory spontaneous contractions induced by Xenin25 was also attenuated by ODQ and apamin. SP-, NOS-, and VIP-immunoreactive neurons were detected in the myenteric plexus (MP) of the rat distal colon. Small subsets of the SP-positive neurons were also Calbindin positive. Most of the VIP-positive neurons were also NOS positive, and small subsets of the NK1R-positive neurons were also VIP positive. Based on the present results, we propose the following mechanism. Xenin25 activates neuronal NTSR1 on the SP neurons of IPANs, and transmitters from the VIP and apamin-sensitive NO neurons synergistically inhibit the spontaneous circular muscle contractions via NK1R. Subsequently, the postinhibitory spontaneous contractions are induced by the offset of apamin-sensitive NO neuron activation via the interstitial cells of Cajal. In addition, Xenin25 also activates the muscular NTSR1 to induce relaxation. Thus, Xenin25 is considered to be an important modulator of post prandial circular muscle contraction of distal colon since the release of Xenin25 from enteroendocrine cells is stimulated by food intake.

Red rice koji extract alleviates hyperglycemia by increasing glucose uptake and glucose transporter type 4 levels in skeletal muscle in two diabetic mouse models.

BACKGROUND: Red rice koji (RRK), prepared by growing Monascus species on steamed rice, has been reported to lower blood glucose levels in diabetic animal models. However, the action mechanism is not yet completely understood. OBJECTIVE: The objective of this study was to examine the mechanism underlying the hypoglycemic action of RRK extract in two diabetic animal models: the insulin-deficiency mice, where the insulin deficiency was induced by streptozotocin (STZ), and insulin-resistance mice, where the insulin resistance was induced by a high-fat diet (HFD). DESIGN: Low (12.5 mg/kg body weight [BW]) and high (50.0 mg/kg BW) doses of RRK extract were orally administered to the mice for 10 successive days (0.25 mL/day/mouse). The protein expression levels of glucose transporter type 4 (GLUT4) in the skeletal muscle and glucose transporter type 2 (GLUT2) in the liver were measured. Blood glucose (BG) levels of STZ-treated mice in insulin tolerance test (ITT) and BG and insulin levels of HFD-fed mice in intraperitoneal glucose tolerance test (IPGTT) were investigated. RESULTS: In the STZ-treated mice, oral administration of RRK extract lowered BG levels and food intake but increased plasma 1,5-anhydroglucitol level. Moreover, the RRK extract lowered the BG levels of STZ-treated mice as measured by ITT. In the HFD-fed mice, we confirmed that the orally administered RRK extract lowered the BG and the homeostasis model assessment index for insulin resistance. Furthermore, the RRK extract lowered the BG and insulin levels of HFD-fed mice in IPGTT. Regarding the protein levels of GLUT, the orally administered RRK extract increased the GLUT4 level in the skeletal muscle; however, the RRK extract did not alter the GLUT2 level in the liver of either the STZ-treated or the HFD-fed mice. DISCUSSION: Our study demonstrates that RRK extract can improve impaired glucose tolerance in mouse models of diabetes by enhancing GLUT4 expression in skeletal muscle. CONCLUSION: These results suggest that RRK extract could potentially be a functional food for the treatment of diabetes mellitus.

Lipopolysaccharides transport during fat absorption in rodent small intestine.

Lipopolysaccharides (LPS) are potent pro-inflammatory molecules that enter the systemic circulation from the intestinal lumen by uncertain mechanisms. We investigated these mechanisms and the effect of exogenous glucagon-like peptide-2 (GLP-2) on LPS transport in the rodent small intestine. Transmucosal LPS transport was measured in Ussing-chambered rat jejunal mucosa. In anesthetized rats, the appearance of fluorescein isothiocyanate (FITC)-LPS into the portal vein (PV) and the mesenteric lymph was simultaneously monitored after intraduodenal perfusion of FITC-LPS with oleic acid and taurocholate (OA/TCA). In vitro, luminally applied LPS rapidly appeared in the serosal solution only with luminal OA/TCA present, inhibited by the lipid raft inhibitor methyl-ß-cyclodextrin (MßCD) and the CD36 inhibitor sulfosuccinimidyl oleate (SSO), or by serosal GLP-2. In vivo, perfusion of FITC-LPS with OA/TCA rapidly increased FITC-LPS appearance into the PV, followed by a gradual increase of FITC-LPS into the lymph. Rapid PV transport was inhibited by the addition of MßCD or by SSO, whereas transport into the lymph was inhibited by chylomicron synthesis inhibition. Intraveous injection of the stable GLP-2 analog teduglutide acutely inhibited FITC-LPS transport into the PV, yet accelerated FITC-LPS transport into the lymph via Nω-nitro-l-arginine methyl ester (l-NAME)- and PG97-269-sensitive mechanisms. In vivo confocal microscopy in mouse jejunum confirmed intracellular FITC-LPS uptake with no evidence of paracellular localization. This is the first direct demonstration in vivo that luminal LPS may cross the small intestinal barrier physiologically during fat absorption via lipid raft- and CD36-mediated mechanisms, followed by predominant transport into the PV, and that teduglutide inhibits LPS uptake into the PV in vivo.NEW & NOTEWORTHY We report direct in vivo confirmation of transcellular lipopolysaccharides (LPS) uptake from the intestine into the portal vein (PV) involving CD36 and lipid rafts, with minor uptake via the canonical chylomicron pathway. The gut hormone glucagon-like peptide-2 (GLP-2) inhibited uptake into the PV. These data suggest that the bulk of LPS absorption is via the PV to the liver, helping clarify the mechanism of LPS transport into the PV as part of the "gut-liver" axis. These data do not support the paracellular transport of LPS, which has been implicated in the pathogenesis of the "leaky gut" syndrome.

Inactivation kinetics modeling of Escherichia coli in concentrated urine for implementing predictive environmental microbiology in sanitation safety planning.

Urine concentration (condensation) leads to the inactivation of pathogens in urine owing to a hyperosmotic environment. This study proposed an inactivation kinetic model of Escherichia coli (E. coli), a surrogate of human bacterial pathogens, in concentrated synthetic urine. The model parameters were obtained under an assumption that the inactivation rate of E. coli followed a binomial distribution, which made it possible to accurately simulate the time-course decay of E. coli in synthetic urine. The inactivation rate constant values obtained in concentrated urine samples, ammonium buffer solutions and carbonate buffer solutions indicated that the osmotic pressure was a relatively predominant cause for the inactivation of E. coli. The appropriate storage time was estimated using the approach of quantitative microbial risk assessment, which indicated that the 5-fold concentrated urine could be safely collected after 1-day storage when urea was hydrolyzed, whereas 91-hour storage was required for non-concentrated urine. The occupational risk was not negligible even with 6-month storage at 20 °C when urea was not hydrolyzed, which suggested that the urine storage styles should be clarified more minutely. The present study highlights the importance of "predictive environmental microbiology," which deals with inactivation kinetic models of microorganisms under varied environmental conditions to fully implement the hazard analysis and critical control point (HACCP) approach for the safe use of human excreta in agriculture.

Potent Antibacterial Activity of Synthetic Peptides Designed from Salusin-ß and HIV-1 Tat(49-57).

Salusin-ß is an endogenous bioactive peptide that was identified in a human full-length enriched cDNA library using bioinformatics analyses. In our previous study, we found that synthetic salusin-ß exhibits antibacterial activity against only Gram-positive microorganisms such as Staphylococcus aureus NBRC 12732. Salusin-ß has an ability to depolarize the cytoplasmic membrane of this bacterium, and this phenomenon may be linked to the antibacterial activity of this peptide. A cell-penetrating peptide (CPP), human immunodeficiency virus (HIV)-1 transactivator of transcription (Tat) (49-57) is a short cationic peptide that can traverse cell membranes. In this report, synthetic peptide conjugates of salusin-ß and HIV-1 Tat(49-57) showed potent antibacterial activities against both Gram-positive Staphylococcus aureus NBRC 12732 and Gram-negative Escherichia coli NBRC 12734. The synthetic peptides also depolarized the cytoplasmic membrane of Escherichia coli NBRC 12734 as well as Staphylococcus aureus NBRC 12732. These results suggested that HIV-1 Tat(49-57) is a protein transduction domain or CPP that changes the interaction mode between salusin-ß and the cell membrane of Escherichia coli NBRC 12734. By binding to HIV-1 Tat(49-57), salusin-ß showed a broad antibacterial spectrum regardless of whether the target was a Gram-positive or Gram-negative bacterium.

Correction: The effects of intra-stomach obestatin administration on intestinal contractility in neonatal piglets fed milk formula.

[This corrects the article DOI: 10.1371/journal.pone.0230190.].

The effects of intra-stomach obestatin administration on intestinal contractility in neonatal piglets fed milk formula.

A 23-amino acid peptide named obestatin is derived from the ghrelin gene. The aim of the experiment was to study the effects of enteral obestatin administration for a 6-day period on intestinal contractility in piglets fed milk formula. Pigs were treated with 0.9% NaCl (group C) or varying doses of obestatin: 2 µg/kg body weight (BW) (group O2), 10 µg/kg BW (O10) or 15 µg/kg BW (O15) every 8 hours via a stomach tube. Blood was sampled for assessment of obestatin concentration. Duodenal and middle jejunum whole-thickness preparations were studied in an organ bath for isometric recording under electric field stimulation (EFS) and increasing doses of acetylcholine (ACh), and in the presence of atropine and tetrodotoxin (TTX). Additionally, the measurement of intestinal muscularis layer and the immunodetection of Muscarinic Acetylcholine Receptors (M1 and M2) were performed. In comparison to C animals, the obestatin concentration in blood plasma was significantly increased in groups O10 and O15. In both studied intestinal segments, significant increases in the frequency and amplitude of spontaneous contractions were observed in O15 and C groups. In the duodenum and middle jejunum significant differences in responsiveness to EFS (0.5, 5 and 50 Hz) were observed between the groups. The addition of 10-4 M ACh to the duodenum significantly increased the responsiveness in tissues. In contrast, in the middle jejunum a significant increase in the amplitude of contraction was observed after the addition of 10-9 and 10-6 M ACh (groups O15 and O10, respectively). Pretreatment with atropine and TTX resulted in a significant decrease in the responsiveness of the intestinal preparations from all groups, in both studied segments. The increased contractility was not dependent on the expression of muscarinic receptors. Results indicate the importance of enteral obestatin administration in the regulation of intestinal contractility in neonatal piglets.

GLP-2 Acutely Prevents Endotoxin-Related Increased Intestinal Paracellular Permeability in Rats.

BACKGROUND: Circulating endotoxin (lipopolysaccharide, LPS) increases the gut paracellular permeability. We hypothesized that glucagon-like peptide-2 (GLP-2) acutely reduces LPS-related increased intestinal paracellular permeability by a mechanism unrelated to its intestinotrophic effect. METHODS: We assessed small intestinal paracellular permeability in vivo by measuring the appearance of intraduodenally perfused FITC-dextran 4000 (FD4) into the portal vein (PV) in rats 1-24 h after LPS treatment (5 mg/kg, ip). We also examined the effect of a stable GLP-2 analog teduglutide (TDG) on FD4 permeability. RESULTS: FD4 movement into the PV was increased 6 h, but not 1 or 3 h after LPS treatment, with increased PV GLP-2 levels and increased mRNA expressions of proinflammatory cytokines and proglucagon in the ileal mucosa. Co-treatment with a GLP-2 receptor antagonist enhanced PV FD4 concentrations. PV FD4 concentrations 24 h after LPS were higher than FD4 concentrations 6 h after LPS, reduced by exogenous GLP-2 treatment given 6 or 12 h after LPS treatment. FD4 uptake measured 6 h after LPS was reduced by TDG 3 or 6 h after LPS treatment. TDG-associated reduced FD4 uptake was reversed by the VPAC1 antagonist PG97-269 or L-NAME, not by EGF or IGF1 receptor inhibitors. CONCLUSIONS: Systemic LPS releases endogenous GLP-2, reducing LPS-related increased permeability. The therapeutic window of exogenous GLP-2 administration is at minimum within 6-12 h after LPS treatment. Exogenous GLP-2 treatment is of value in the prevention of increased paracellular permeability associated with endotoxemia.

Rubiscolin-6, a δ-Opioid Peptide from Spinach RuBisCO, Exerts Antidepressant-Like Effect in Restraint-Stressed Mice.

Rubiscolin-6 (Tyr-Pro-Leu-Asp-Leu-Phe) is produced by a pepsin digest of spinach d-ribulose-1,5-bisphosphate carboxylase/oxygenase (RuBisCO) and known to act as an agonist on δ-opioid receptor. Here, we showed that administration of rubiscolin-6 reduced immobility time in the tail suspension test in restraint-stressed mice without effect on locomotor activity. The antidepressant-like effect of rubiscolin-6 was blocked by a δ-opioid receptor antagonist, naltrindole. These results indicate that rubiscolin-6 exerts antidepressant-like effect through activation of δ-opioid receptor.

Xenin-25 induces anion secretion by activating noncholinergic secretomotor neurons in the rat ileum.

Xenin-25 is a neurotensin-like peptide that is secreted by enteroendocrine cells in the small intestine. Xenin-8 is reported to augment duodenal anion secretion by activating afferent neural pathways. The intrinsic neuronal circuits mediating the xenin-25-induced anion secretion were characterized using the Ussing-chambered, mucosa-submucosa preparation from the rat ileum. Serosal application of xenin-25 increased the short-circuit current in a concentration-dependent manner. The responses were abolished by the combination of Cl--free and HCO3- -free solutions. The responses were almost completely blocked by TTX (10-6 M) but not by atropine (10-5 M) or hexamethonium (10-4 M). The selective antagonists for neurotensin receptor 1 (NTSR1), neurokinin 1 (NK1), vasoactive intestinal polypeptide (VIP) receptors 1 and 2 (VPAC1 and VPAC2, respectively), and capsaicin, but not 5-hydroxyltryptamine receptors 3 and 4 (5-HT3 and 5-HT4), NTSR2, and A803467, inhibited the responses to xenin-25. The expression of VIP receptors (Vipr) in rat ileum was examined using RT-PCR. The Vipr1 PCR products were detected in the submucosal plexus and mucosa. Immunohistochemical staining showed the colocalization of NTSR1 and NK1 with substance P (SP)- and calbindin-immunoreactive neurons in the submucosal plexus, respectively. In addition, NK1 was colocalized with noncholinergic VIP secretomotor neurons. Based on the results from the present study, xenin-25-induced Cl-/ HCO3- secretion is involved in NTSR1 activation on intrinsic and extrinsic afferent neurons, followed by the release of SP and subsequent activation of NK1 expressed on noncholinergic VIP secretomotor neurons. Finally, the secreted VIP may activate VPAC1 on epithelial cells to induce Cl-/ HCO3- secretion in the rat ileum. Activation of noncholinergic VIP secretomotor neurons by intrinsic primary afferent neurons and extrinsic afferent neurons by postprandially released xenin-25 may account for most of the neurogenic secretory response induced by xenin-25. NEW & NOTEWORTHY This study is the first to investigate the intrinsic neuronal circuit responsible for xenin-25-induced anion secretion in the rat small intestine. We have found that nutrient-stimulated xenin-25 release may activate noncholinergic vasoactive intestinal polypeptide (VIP) secretomotor neurons to promote Cl-/ HCO3- secretion through the activation of VIP receptor 1 on epithelial cells. Moreover, the xenin-25-induced secretory responses are mainly linked with intrinsic primary afferent neurons, which are involved in the activation of neurotensin receptor 1 and neurokinin 1 receptor.

Rubiscolin-6 activates opioid receptors to enhance glucose uptake in skeletal muscle.

Rubiscolin-6 is an opioid peptide derived from plant ribulose bisphosphate carboxylase/oxygenase (Rubisco). It has been demonstrated that opioid receptors could control glucose homeostasis in skeletal muscle independent of insulin action. Therefore, Rubiscolin-6 may be involved in the control of glucose metabolism. In the present study, we investigated the effect of rubiscolin-6 on glucose uptake in skeletal muscle. Rubiscolin-6-induced glucose uptake was measured using the fluorescent indicator 2-[N-(7-nitrobenz-2-oxa-1,3-diazol-4-yl) amino]-2-deoxyglucose (2-NBDG) in L6 and C2C12 cell lines. The protein expressions of glucose transporter 4 (GLUT4) and AMP-activated protein kinase (AMPK) in L6 cells were observed by Western blotting. The in vivo effects of rubiscolin-6 were characterized in streptozotocin (STZ)-induced diabetic rats. Rubiscolin-6 induced a concentration-dependent increase in glucose uptake levels. The increase of phospho-AMPK (pAMPK) and GLUT4 expressions were also observed in L6 and C2C12 cells. Effects of rubiscolin-6 were blocked by opioid receptor antagonists and/or associated signals inhibitors. Moreover, Rubiscolin-6 produced a dose-dependent reduction of blood glucose and increased GLUT4 expression in STZ-induced diabetic rats. In conclusion, rubiscolin-6 increases glucose uptake, potentially via an activation of AMPK to enhance GLUT4 translocation after binding to opioid receptors in skeletal muscle.

Improvement of Diabetes Mellitus Symptoms by Intake of Ninjin'yoeito.

Diabetes mellitus is a well-known common disease and one of the most serious social problems in the worldwide. Although various types of drugs are developed, the number of patients suffering from diabetes mellitus is still increasing. Ninjin'yoeito (NYT) is one of formulas used in Japanese traditional herbal medicines for improving various types of metabolic disorders. However, the effect of NYT on diabetes mellitus has not yet been investigated. In the present study, we tried to clarify the action of NYT on the serum glucose level in streptozotocin (STZ)-induced diabetic mice. We found that intake of NYT decreased the serum glucose level and increased insulin sensitivity in STZ-induced diabetic mice. NYT treatment also improved acidification of the interstitial fluid around skeletal muscles found in STZ-induced diabetic mice, while the interstitial fluid acidification has been reported to cause insulin resistance. Furthermore, in the proximal colon of STZ-induced diabetic mice, NYT treatment showed a tendency to increase the expression of sodium-coupled monocarboxylate transporter 1 (SMCT1), which has ability to absorb weak organic acids (pH buffer molecules) resulting in improvement of the interstitial fluid acidification. Based on these observations, the present study suggests that NYT is a useful formula to improve hyperglycemia and insulin resistance via elevation of interstitial fluid pH in diabetes mellitus, which might be caused by increased absorption of pH buffer molecules (SMCT1 substrates, weak organic acids) mediated through possibly elevated SMCT1 expression in the proximal colon.