Oxytocin at physiological concentrations evokes adrenocorticotropin (ACTH) release from corticotrophs by increasing intracellular free calcium mobilized mainly from intracellular stores. Oxytocin displays synergistic or additive effects on ACTH-releasing factor or arginine vasopressin-induced ACTH secretion, respectively

The potency of oxytocin (OT) in evoking ACTH secretion by isolated, superfused rat adenohypophyseal corticotrophs and its enhancement by CRF and arginine vasopressin (AVP) were analyzed. Each secretagogue effectively released ACTH from adenohypophyseal cells when added separately in pulsatile fashion in physiological concentrations based on hypophyseal portal blood (OT, 10 nM; AVP, 0.5 nM; CRF, 0.1 nM). OT released ACTH at concentrations as low as 1 nM. Moreover, a dose- response relationship up to 10 microM was revealed. Combinations of a constant amount of CRF (0.1 nM) with increasing concentrations of OT exerted a synergistic effect on ACTH release. In contrast, OT given in various concentrations in combination with AVP (0.5 nM) produced an additive effect on ACTH release. To study the mechanism of action of OT on ACTH secretion, cytosolic free calcium levels in single pituitary cells exposed to OT or AVP were measured using the calcium-sensitive fluorescent indicator Fura-2. Corticotrophs among mixed adenohypophyseal cell types in the primary cultures were identified by immunocytochemistry. More than 500 cells were individually stimulated with OT or AVP. Basal cytosolic free calcium levels ranged between 80- 130 nM free calcium. The addition of 100 nM OT or 1 microM AVP increased the cytosolic free calcium concentration within 3 sec to values ranging from 500-800 nM. An increase in intracellular calcium ranging from 200-500 nM due to OT could still be observed after extracellular calcium depletion. Taken together, our data demonstrate that physiological concentrations of OT stimulate ACTH secretion, independent of the other ACTH secretagogues, by mobilizing calcium mainly from intracellular stores

Physiological concentrations of bile acids down-regulate agonist induced secretion in colonic epithelial cells

In patients with bile acid malabsorption, high concentrations of bile acids enter the colon and stimulate Cl− and fluid secretion, thereby causing diarrhoea. However, deoxycholic acid (DCA), the predominant colonic bile acid, is normally present at lower concentrations where its role in regulating transport is unclear. Thus, the current study set out to investigate the effects of physiologically relevant DCA concentrations on colonic epithelial secretory function. Cl− secretion was measured as changes in short-circuit current across voltage-clamped T84 cell monolayers. At high concentrations (0.5–1 mM), DCA acutely stimulated Cl− secretion but this effect was associated with cell injury, as evidenced by decreased transepithelial resistance (TER) and increased lactate dehydrogenase (LDH) release. In contrast, chronic (24 hrs) exposure to lower DCA concentrations (10–200 μM) inhibited responses to Ca2+ and cAMP-dependent secretagogues without altering TER, LDH release, or secretagogue-induced increases in intracellular second messengers. Other bile acids – taurodeoxycholic acid, chenodeoxycholic acid and cholic acid – had similar antisecretory effects. DCA (50 μM) rapidly stimulated phosphorylation of the epidermal growth factor receptor (EGFr) and both ERK and p38 MAPKs (mitogen-activated protein kinases). The EGFr inhibitor, AG1478, and the protein synthesis inhibitor, cycloheximide, reversed the antisecretory effects of DCA, while the MAPK inhibitors, PD98059 and SB203580, did not. In summary, our studies suggest that, in contrast to its acute prosecretory effects at pathophysiological concentrations, lower, physiologically relevant, levels of DCA chronically down-regulate colonic epithelial secretory function. On the basis of these data, we propose a novel role for bile acids as physiological regulators of colonic secretory capacity

[6]-Gingerol, from Zingiber officinale, potentiates GLP-1 mediated glucose-stimulated insulin secretion pathway in pancreatic β-cells and increases RAB8/RAB10-regulated membrane presentation of GLUT4 transporters in skeletal muscle to improve hyperglycemia in Leprdb/db type 2 diabetic mice

This article is distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated.Abstract Background [6]-Gingerol, a major component of Zingiber officinale, was previously reported to ameliorate hyperglycemia in type 2 diabetic mice. Endocrine signaling is involved in insulin secretion and is perturbed in db/db Type-2 diabetic mice. [6]-Gingerol was reported to restore the disrupted endocrine signaling in rodents. In this current study on Leprdb/db diabetic mice, we investigated the involvement of endocrine pathway in the insulin secretagogue activity of [6]-Gingerol and the mechanism(s) through which [6]-Gingerol ameliorates hyperglycemia. Methods Leprdb/db type 2 diabetic mice were orally administered a daily dose of [6]-Gingerol (200 mg/kg) for 28 days. We measured the plasma levels of different endocrine hormones in fasting and fed conditions. GLP-1 levels were modulated using pharmacological approaches, and cAMP/PKA pathway for insulin secretion was assessed by qRT-PCR and ELISA in isolated pancreatic islets. Total skeletal muscle and its membrane fractions were used to measure glycogen synthase 1 level and Glut4 expression and protein levels. Results 4-weeks treatment of [6]-Gingerol dramatically increased glucose-stimulated insulin secretion and improved glucose tolerance. Plasma GLP-1 was found to be significantly elevated in the treated mice. Pharmacological intervention of GLP-1 levels regulated the effect of [6]-Gingerol on insulin secretion. Mechanistically, [6]-Gingerol treatment upregulated and activated cAMP, PKA, and CREB in the pancreatic islets, which are critical components of GLP-1-mediated insulin secretion pathway. [6]-Gingerol upregulated both Rab27a GTPase and its effector protein Slp4-a expression in isolated islets, which regulates the exocytosis of insulin-containing dense-core granules. [6]-Gingerol treatment improved skeletal glycogen storage by increased glycogen synthase 1 activity. Additionally, GLUT4 transporters were highly abundant in the membrane of the skeletal myocytes, which could be explained by the increased expression of Rab8 and Rab10 GTPases that are responsible for GLUT4 vesicle fusion to the membrane. Conclusions Collectively, our study reports that GLP-1 mediates the insulinotropic activity of [6]-Gingerol, and [6]-Gingerol treatment facilitates glucose disposal in skeletal muscles through increased activity of glycogen synthase 1 and enhanced cell surface presentation of GLUT4 transporters

Evaluation and Diagnostic Potential of Serum Ghrelin in Feline Hypersomatotropism and Diabetes Mellitus

BACKGROUND: Ghrelin is a growth hormone secretagogue. It is a potent regulator of energy homeostasis. Ghrelin concentration is down‐regulated in humans with hypersomatotropism (HS) and increases after successful treatment. Additionally, ghrelin secretion seems impaired in human diabetes mellitus (DM). HYPOTHESIS: Serum ghrelin concentration is down‐regulated in cats with HS‐induced DM (HSDM) compared to healthy control cats or cats with DM unrelated to HS and increases after radiotherapy. ANIMALS: Cats with DM (n = 20) and with HSDM (n = 32), 13 of which underwent radiotherapy (RT‐group); age‐matched controls (n = 20). METHODS: Retrospective cross‐sectional study. Analytical performance of a serum total ghrelin ELISA was assessed and validated for use in cats. Differences in serum ghrelin, fructosamine, IGF‐1 and insulin were evaluated. RESULTS: Ghrelin was significantly higher (P < .001) in control cats (mean ± SD: 12.9 ± 6.8 ng/mL) compared to HSDM‐ (7.9 ± 3.3 ng/mL) and DM‐cats (6.7 ± 2.3 ng/mL), although not different between the HSDM‐ and DM‐cats. After RT ghrelin increased significantly (P = .003) in HSDM‐cats undergoing RT (from 6.6 ± 1.9 ng/mL to 9.0 ± 2.2 ng/mL) and the after RT ghrelin concentrations of HSDM cats were no longer significantly different from the serum ghrelin concentration of control cats. Serum IGF‐1 did not significantly change in HSDM‐cats after RT, despite significant decreases in fructosamine and insulin dose. CONCLUSION AND CLINICAL IMPORTANCE: Ghrelin appears suppressed in cats with DM and HSDM, although increases after RT in HSDM, suggesting possible presence of a direct or indirect negative feedback system between growth hormone and ghrelin. Serum ghrelin might therefore represent a marker of treatment effect

Quinolones modulate ghrelin receptor signaling: potential for a novel small molecule scaffold in the treatment of cachexia

Cachexia is a metabolic wasting disorder characterized by progressive weight loss, muscle atrophy, fatigue, weakness, and appetite loss. Cachexia is associated with almost all major chronic illnesses including cancer, heart failure, obstructive pulmonary disease, and kidney disease and significantly impedes treatment outcome and therapy tolerance, reducing physical function and increasing mortality. Current cachexia treatments are limited and new pharmacological strategies are needed. Agonists for the growth hormone secretagogue (GHS-R1a), or ghrelin receptor, prospectively regulate the central regulation of appetite and growth hormone secretion, and therefore have tremendous potential as cachexia therapeutics. Non-peptide GHS-R1a agonists are of particular interest, especially given the high gastrointestinal degradation of peptide-based structures, including that of the endogenous ligand, ghrelin, which has a half-life of only 30 min. However, few compounds have been reported in the literature as non-peptide GHS-R1a agonists. In this paper, we investigate the in vitro potential of quinolone compounds to modulate the GHS-R1a in both transfected human cells and mouse hypothalamic cells. These chemically synthesized compounds demonstrate a promising potential as GHS-R1a agonists, shown by an increased intracellular calcium influx. Further studies are now warranted to substantiate and exploit the potential of these novel quinolone-based compounds as orexigenic therapeutics in conditions of cachexia and other metabolic and eating disorders.Irish Research Council for Science and Technology (IRCSET)Science Foundation Ireland (SFI/12/IP/1315)Science Foundation Ireland (SFI/12/RC/2275)Science Foundation Ireland (SFI/12/RC/2273)Universidad de Sevill

Ghrelin Decreases Firing Activity of Gonadotropin-Releasing Hormone (GnRH) Neurons in an Estrous Cycle and Endocannabinoid Signaling Dependent Manner.

The orexigenic peptide, ghrelin is known to influence function of GnRH neurons, however, the direct effects of the hormone upon these neurons have not been explored, yet. The present study was undertaken to reveal expression of growth hormone secretagogue receptor (GHS-R) in GnRH neurons and elucidate the mechanisms of ghrelin actions upon them. Ca(2+)-imaging revealed a ghrelin-triggered increase of the Ca(2+)-content in GT1-7 neurons kept in a steroid-free medium, which was abolished by GHS-R-antagonist JMV2959 (10µM) suggesting direct action of ghrelin. Estradiol (1nM) eliminated the ghrelin-evoked rise of Ca(2+)-content, indicating the estradiol dependency of the process. Expression of GHS-R mRNA was then confirmed in GnRH-GFP neurons of transgenic mice by single cell RT-PCR. Firing rate and burst frequency of GnRH-GFP neurons were lower in metestrous than proestrous mice. Ghrelin (40nM-4μM) administration resulted in a decreased firing rate and burst frequency of GnRH neurons in metestrous, but not in proestrous mice. Ghrelin also decreased the firing rate of GnRH neurons in males. The ghrelin-evoked alterations of the firing parameters were prevented by JMV2959, supporting the receptor-specific actions of ghrelin on GnRH neurons. In metestrous mice, ghrelin decreased the frequency of GABAergic mPSCs in GnRH neurons. Effects of ghrelin were abolished by the cannabinoid receptor type-1 (CB1) antagonist AM251 (1µM) and the intracellularly applied DAG-lipase inhibitor THL (10µM), indicating the involvement of retrograde endocannabinoid signaling. These findings demonstrate that ghrelin exerts direct regulatory effects on GnRH neurons via GHS-R, and modulates the firing of GnRH neurons in an ovarian-cycle and endocannabinoid dependent manner

Glucocorticoids rapidly inhibit oxytocin-stimulated adrenocorticotropin release from rat anterior pituitary cells, without modifying intracellular calcium transients

Glucocorticoid hormones suppress the secretion of ACTH evoked by secretagogues such as CRF and arginine vasopressin. In this study, we investigated the effects of glucocorticoids on ACTH release induced by oxytocin (OT) and on intracellular free calcium ion levels in corticotropes prepared from the adenohypophyses of female Wistar rats. Pulsatile additions of physiological concentration of OT (10 nM) to superfused anterior pituitary cells caused pulsatile ACTH release about 4-fold above basal secretion with similar peak amounts of ACTH during subsequent OT pulses. Exposure of the cells to corticosterone (100 nM) or to a selective glucocorticoid receptor agonist RU 28362 (100 nM) for 30 min suppressed OT-stimulated but not basal ACTH release by approximately 60%. Inhibition gradually disappeared during subsequent pulses of OT in the absence of corticosterone. Pretreatment with the selective antagonist RU 38486 (1 microM) completely blocked the inhibitory effect of corticosterone on OT-induced ACTH secretion. Changes in free cytosolic calcium levels in single cultured pituitary cells were measured using the calcium indicator Fura-2. OT caused calcium transients in corticotropes, which were identified by immunocytochemistry. They responded in a similar manner to a second OT stimulus when preincubated for 30 min with corticosterone (1 microM) or with RU 28362 (1 microM). Our data indicate that glucocorticoids, via glucocorticoid receptors, rapidly inhibit OT-stimulated ACTH secretion by corticotropes without affecting intracellular calcium transients due to OT. Therefore, we conclude that rapid inhibition of ACTH release by glucocorticoids interferes with cellular signal transduction beyond the step of calcium mobilization

Ghrelin axis genes, peptides and receptors : recent findings and future challenges

The ghrelin axis consists of the gene products of the ghrelin gene (GHRL), and their receptors, including the classical ghrelin receptor GHSR. While it is well-known that the ghrelin gene encodes the 28 amino acid ghrelin peptide hormone, it is now also clear that the locus encodes a range of other bioactive molecules, including novel peptides and non-coding RNAs. For many of these molecules, the physiological functions and cognate receptor(s) remain to be determined. Emerging research techniques, including proteogenomics, are likely to reveal further ghrelin axis-derived molecules. Studies of the role of ghrelin axis genes, peptides and receptors, therefore, promises to be a fruitful area of basic and clinical research in years to come