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it is known that urocortin 1 (UCN) acts on both corticotropinreleasing factor receptors (CRF 1 and CRF2), the mechanisms underlying UCN-induced anorexia remain unclear. In contrast, ghrelin, the endogenous ligand for the growth hormone secretagogue receptor, stimulates food intake. In the present study, we examined the effects of CRF 1 and CRF2 receptor antagonists (CRF1a and CRF2a) on ghrelin secretion and synthesis, c-fos mRNA expression in the caudal brain stem, and food intake following intracerebroventricular administration of UCN. Eight-week-old, male Sprague-Dawley rats were used after 24-h food deprivation. Acylated and des-acylated ghrelin levels were measured by enzyme-linked immunosorbent assay. The mRNA expressions of preproghrelin and c-fos were measured by real-time RT-PCR. The present study provided the following important insights into the mechanisms underlying the anorectic effects of UCN: 1) UCN increased acylated and des-acylated ghrelin levels in the gastric body and decreased their levels in the plasma; 2) UCN decreased preproghrelin mRNA levels in the gastric body; 3) UCN-induced reduction of plasma ghrelin and food intake were restored by CRF 2a but not CRF1a; 4) UCN-induced increase of c-fos mRNA levels in the caudal brain stem containing the nucleus of the solitary tract (NTS) was inhibited by CRF 2a; and 5) UCN-induced reduction of food intake was restored by exogenous ghrelin and rikkunshito, an endogenous ghrelin secretion regulator. Thus, UCN increases neuronal activation in the caudal brain stem containing NTS via CRF 2 receptors, which may be related to UCN-induced inhibition of both ghrelin secretion and food intake.corticotropin-releasing factor receptors; dorsal vagal complex; rikkunshito; digestive system hormone; anorexia STRESSFUL LIFE EVENTS have been associated with the onset or symptom exacerbation of several functional gastrointestinal disorders including functional dyspepsia and irritable bowel syndrome (24). Corticotropin-releasing factor (CRF) is a key mediator in the central nervous system and is secreted as an adaptive response to stress. In rodents, central CRF administration induces stress-like behaviors, including increased depression, decreased rearing activity, and suppression of food intake (5, 50). Urocortin 1 (UCN), a member of the mammalian CRF family, bears 45% sequence identity to CRF and acts as an endogenous ligand for CRF receptors (49). Although CRF and UCN mediate their actions through the activation of CRF 1 and CRF 2 receptors, CRF and UCN display different binding affinities for these receptors. CRF preferentially binds to CRF 1 receptors, whereas UCN shows high affinity for both receptors; we therefore believe that UCN is useful for examining the actions of both CRF 1 and CRF 2 receptors. Central UCN administration suppresses feeding in rats and mice, and the suppression was shown to be at least partly CRF 2 mediated in studies that used selective antagonists (7) or antisense oligonucleotides (38). In addition, the initial effect of food intake in...

BackgroundColor perception is important for fish to survive and reproduce in nature. Visual pigments in the retinal photoreceptor cells are responsible for receiving light stimuli, but the function of the pigments in vivo has not been directly investigated in many animals due to the lack of color-blind lines and appropriate color-perception tests.MethodsIn this study, we established a system for producing color-blind fish and testing their spectral sensitivity. First, we disrupted long-wavelength-sensitive (LWS) opsins of medaka (Oryzias latipes) using the CRISPR/Cas9 system to make red-color-blind lines. Single guide RNAs were designed using the consensus sequences between the paralogous LWSa and LWSb genes to simultaneously introduce double-frameshift mutations. Next, we developed a non-invasive and no-prior-learning test for spectral sensitivity by applying an optomotor response (OMR) test under an Okazaki Large Spectrograph (OLS), termed the O-O test. We constructed an electrical-rotary cylinder with black/white stripes, into which a glass aquarium containing one or more fish was placed under various monochromatic light conditions. The medaka were irradiated by the OLS every 10 nm, from wavelengths of 700 nm to 900 nm, and OMR was evaluated under each condition.ResultsWe confirmed that the lws − medaka were indeed insensitive to red light (protanopia). While the control fish responded to wavelengths of up to 830 nm (λ = 830 nm), the lws − mutants responded up to λ = 740 nm; however, this difference was not observed after adaptation to dark: both the control and lws − fish could respond up to λ = 820 ~ 830 nm.ConclusionsThese results suggest that the lws − mutants lost photopic red-cone vision, but retained scotopic rod vision. Considering that the peak absorption spectra (λmax) of medaka LWSs are about 560 nm, but the light-adapted control medaka could respond behaviorally to light at λ = 830 nm, red-cone vision could cover an unexpectedly wide range of wavelengths, and behavioral tests could be an effective way to measure spectral sensitivity. Using the CRISPR/Cas9 and O-O systems, the establishment of various other color-blind lines and assessment of their spectra sensitivity could be expected to proceed in the future.Electronic supplementary materialThe online version of this article (doi:10.1186/s12863-017-0477-7) contains supplementary material, which is available to authorized users.

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Glycolaldehyde, an intermediate of the Maillard reaction, and fructose, which is mainly derived from the polyol pathway, rapidly inactivate human Cu,Zn-superoxide dismutase (SOD) at the physiological concentration. We employed this inactivation with these carbonyl compounds as a model glycation reaction to investigate whether carnosine and its related compounds could protect the enzyme from inactivation. Of eight derivatives examined, histidine, Gly-His, carnosine and Ala-His inhibited the inactivation of the enzyme by fructose (p<0.001), and Gly-His, Ala-His, anserine, carnosine, and homocarnosine exhibited a marked protective effect against the inactivation by glycolaldehyde (p<0.001). The carnosine-related compounds that showed this highly protective effect against the inactivation by glycolaldehyde had high reactivity with glycolaldehyde and high scavenging activity toward the hydroxyl radical as common properties. On the other hand, the carnosine-related compounds that had a protective effect against the inactivation by fructose showed significant hydroxyl radical-scavenging ability. These results indicate that carnosine and such related compounds as Gly-His and Ala-His are effective anti-glycating agents for human Cu,Zn-SOD and that the effectiveness is based not only on high reactivity with carbonyl compounds but also on hydroxyl radical scavenging activity.

Metabolites of cytochrome P-450 are produced in cells when arachidonic acid cascade is activated. Fever genesis depends largely on the cyclooxygenase branch of arachidonic acid cascade, which is caused by many stimuli, such as interleukin (IL)-1, IL-6, and interferon-alpha. To assess the significance of cytochrome P-450 branch in fever, murine recombinant IL-1 beta was bilaterally microinjected (1 ng/microliter) into the medial preoptic area and anterior hypothalamus in conscious rats treated 60 min previously with or without the cytochrome P-450 inhibitor econazole (15 mg/kg im). The IL-1 beta-induced rise in colonic temperature was enhanced after the plateau phase of fever (from 240 min after IL-1 beta) in econazole-pretreated rats (P < 0.001). Another cytochrome P-450 inhibitor, clotrimazole (15 mg/kg im), also enhanced IL-1 beta-induced fever from 160 min after IL-1 beta injection (P < 0.001). Econazole also enhanced the fever when it was given 120 min before injection of IL-1 beta (P < 0.001). The cytochrome P-450 inhibitor, however, did not affect the fever when given 10 min after IL-1 beta (P = 0.95). Econazole and clotrimazole did not alter normal body temperature (P = 0.65 and 0.73, respectively). The results suggest that the metabolite(s) of cytochrome P-450 affect the falling phase after the plateau phase of fever and act as putative endogenous antipyretic(s).

SummaryThe autonomic nervous system (ANS) conveys neuronal input from the brain to the stomach. We investigated mechanisms through which urocortin 1 (UCN1) injected intracerebroventricularly (ICV, 300 pmol/rat) inhibits circulating ghrelin in rats. This was achieved by assessing (1) the induction of c-fos gene expression as a marker of neuronal activation in specific hypothalamic and caudal brainstem regulating ANS; (2) the influence of vagotomy and pharmacological blockade of central and peripheral α- and β-adrenergic receptor (AR) on ICV UCN1 -induced reduction of plasma ghrelin levels (determined by ELISA); and (3) the relevance of this pathway in the feeding response to a fast in rats. UCN1 increased c-fos mRNA expression in key brain sites influencing sympathetic activity namely the hypothalamic paraventricular and ventromedial nuclei, locus coeruleus, nucleus of the solitary tract, and rostral ventrolateral medulla, by 16-, 29-, 6-, 37-, and 13-fold, respectively. In contrast, the dorsal motor nucleus of the vagus had little c-fos mRNA expression and ICV UCN1 induced a similar reduction in acylated ghrelin in the sham-operated (31%) and vagotomized (41%) rats. An intraperitoneal (IP) injection of either a non-selective α- or selective α2-AR antagonist reduced, while a selective α2-AR agonist enhanced ICV UCN1-induced suppression of plasma acylated ghrelin levels. In addition, IP injection of a non-selective β- or selective β1-AR agonist blocked, and selective β1-AR antagonist augmented, the ghrelin response to ICV UCN1. The IP injections of a selective α1- or non-selective β or β2-AR antagonists, or any of the pretreatments given ICV had no effect. ICV UCN1 reduced the 2-h food intake in response to a fast by 80%, and this effect was partially prevented by a selective α2-AR antagonist. These data suggest that ICV UCN1 reduces plasma ghrelin mainly through the brain sympathetic component of the ANS and peripheral AR specifically α2-AR activation and inactivation of β1-AR. The α2-AR pathway contributes to the associated reduction in food intake.

The pathogenesis of gastroesophageal reflux disease (GERD) is multifactorial but the main factor is extended exposure to gastric acid. The extent of esophageal mucosal injury is determined by the degree and duration of esophageal exposure to acid. Patients with GERD have been found to have delayed acid clearance times that are 2-3 times longer than those without GERD. 1) Indeed, the process of esophageal acid clearance is an important factor in the worsening of esophageal mucosal injury. 2) Impaired esophageal clearance can be partly caused by peristaltic dysfunction 2) and lower esophageal sphincter (LES) functioning. 3) Esophageal motility is also regulated by the esophageal vagal nerve at the interneurons of the central subnucleus of the solitary tract complex. 4) A recent study demonstrated the effect of baclofen, a gamma aminobutyric acid B receptor agonist, on esophageal motility and transient LES relaxation in GERD patients. 5) Although esophageal acid clearance is regulated by neurotransmitters both peripherally and centrally, the local responsiveness of the muscle is still unknown.The motor innervations of LES and the proximal stomach are known to be under cholinergic control. [6][7][8] A recent study demonstrated that the serotonin-4 (5-hydroxytryptamine 4; 5-HT 4 ) receptor is localized in LES and regulates LES tone. 9) Thus, we hypothesized that a neurotransmitter-induced LES response in GERD rats may be impaired at the esophagealtissue level, leading to the development of GERD pathology. In the present study, we examined this hypothesis by measuring LES motility in GERD rats and the effects of neurotransmitters on contraction and relaxation using isolated LES strips in an in vitro study. In addition, we investigated the effects of activation of the 5-HT 4 receptor on esophageal erosion in GERD rats. MATERIALS AND METHODSTest Substances Acetylcholine (Ach), atropine, 5-HT, sodium nitroprusside (SNP), a-methylserotonin (a-Me-5-HT: a 5-HT 2 receptor agonist), 5-methoxytryptamine (5-MeOT: a nonspecific 5-HT receptor agonist), 1-(3-chlorophenyl)biguanide HCl (a 5-HT 3 receptor agonist), BW723 C86HCl (a 5-HT 2B receptor agonist), m-chlorophenylpiperazine HCl (mCPP: a 5-HT 2C receptor agonist), or SB204070 (a 5-HT 4 receptor antagonist) were all purchased from Sigma Aldrich Corp. U.S.A. Cisapride and mosapride (5-HT 4 receptor agonists) was purchased from Tocris Bioscience. Ondansetron (a 5-HT 3 receptor antagonist) was purchased from GlaxoSmithKline.Test Animals Male Wistar rats aged 8 weeks (CLEA Japan Inc., Tokyo, Japan) were used for all experiments. During testing, 4-5 animals were housed in each cage and were allowed free access to food and water in a room that was illuminated between 07:00 and 19:00 h. Temperature and humidity were maintained at constant levels. For tests that evaluated the motility of the proximal LES, animals were housed singly in cages and deprived of food for 24 h after surgery, after which they were given free access to food. Animals that Sapporo, Japan: and d Gastroenterol...