Release of gastric inhibitory polypeptide from cultured canine endocrine cells

1994 ◽  
Vol 267 (4) ◽  
pp. E489-E496 ◽  
Author(s):  
T. J. Kieffer ◽  
A. M. Buchan ◽  
H. Barker ◽  
J. C. Brown ◽  
R. A. Pederson
Keyword(s):  
Endocrine Cells ◽  
Gastric Inhibitory Polypeptide ◽  
Adherent Cells ◽  
Inhibitory Peptide ◽  
Cell Content ◽  
Gastrin Releasing Peptide ◽  
A 23187 ◽  
Basal Release ◽  
Cyclase Activator ◽  
Adenylate Cyclase Activator

Canine intestinal duodenal and jejunal epithelial cell preparations enriched for endocrine cells were obtained by sequential collagenase digestion and centrifugal elutriation and maintained in culture for a 40-h period. Adherent cells contained a total cell content (TCC) of 11.5 +/- 2.5 ng (mean +/- SE) immunoreactive gastric inhibitory peptide (IRGIP)/well and 1.4 +/- 0.2 ng immunoreactive somatostatin (IRS)/well. Release experiments were performed by incubation of the cells with various stimuli over a 2-h period. Basal release of IRGIP in 5 mM glucose-5 mM K+ was 2.7 +/- 0.4% TCC. Incubation with concentrations of K+ > 20 mM or glucose > 15 mM significantly increased IRGIP release, as did the addition of a somatostatin immunoneutralizing antibody to the basal media. The addition of the Ca2+ ionophore, A-23187 (10 microM), or the adenylate cyclase activator, forskolin (100 microM), resulted in an IRGIP output greater than four times basal. Porcine gastrin-releasing peptide (GRP), at 1-100 nM, significantly stimulated IRGIP release in a concentration-dependent fashion. IRS release was increased significantly by 55 mM K+, 20 mM glucose, 10 microM A-23187, 100 nM GRP, or 100 microM forskolin.

Gastric inhibitory polypeptide release from a tumor-derived cell line

1995 ◽  
Vol 269 (2) ◽  
pp. E316-E322 ◽  
Author(s):  
T. J. Kieffer ◽  
Z. Huang ◽  
C. H. McIntosh ◽  
A. M. Buchan ◽  
J. C. Brown ◽  
...  
Keyword(s):  
Cell Line ◽  
High Performance ◽  
Neutralizing Antibody ◽  
Gastric Inhibitory Polypeptide ◽  
Total Cell ◽  
Controlled Environment ◽  
Intestinal Tumors ◽  
Cell Content ◽  
Basal Release ◽  
A Cell

A cell line derived from intestinal tumors of transgenic mice (STC-1) was subcloned to produce a stable line with approximately 30% immunoreactive gastric inhibitory polypeptide (irGIP)-containing cells (STC 6-14). High-performance liquid chromatography (HPLC) of STC 6-14 extracts indicated that the tumor cell-derived irGIP had the same retention time as synthetic porcine GIP-(1-42) (pGIP). Approximately 30% of the cells also contained immunoreactive somatostatin (irSS), which eluted as a single peak on HPLC, corresponding with SS-(1-14). On average, each well of extracted cells (5.0 x 10(5) cultured 4 days) contained 33.3 +/- 1.4 ng irGIP and 18.4 +/- 1.5 ng irSS. Basal release of irGIP in the presence of 5 mM glucose was 733 +/- 58 pg.ml cells-1.2h-1 (2.20 +/- 0.17% of total cell content; TCC) and doubled at 20 mM glucose (4.20 +/- 0.42% TCC). The response to glucose was augmented by addition of a SS neutralizing antibody (SOMA-10) and suppressed by 10 nM SS. Basal release of irSS in 5 mM glucose was 377 +/- 35 pg.ml cells-1.2h-1 (2.05 +/- 0.19% TCC) and was increased by glucose (> or = 15 mM) and the addition of pGIP (> or = 1 nM). The STC 6-14 cell line represents a model to study the synthesis, storage, and release of GIP and SS in a controlled environment.

Increase in protein and tubulin mRNA synthesis in frog sensory neurons treated with the adenylate cyclase activator, forskolin

1990 ◽  
Vol 1 (3,4) ◽  
pp. 225-232
Author(s):  
Richard C. Carlsen ◽  
Marino De Leon ◽  
Wolfram Tetzlaff ◽  
Irma M. Parhad ◽  
Mark A. Bisby
Keyword(s):  
Adenylate Cyclase ◽  
Sensory Neurons ◽  
Mrna Synthesis ◽  
Tubulin Mrna ◽  
Cyclase Activator ◽  
Adenylate Cyclase Activator

scholarly journals β3-Adrenoceptor agonists inhibit purinergic receptor-mediated contractions of the murine detrusor

2019 ◽  
Vol 317 (1) ◽  
pp. C131-C142 ◽  
Author(s):  
Zhihui Fong ◽  
Caoimhín S. Griffin ◽  
Mark A. Hollywood ◽  
Keith D. Thornbury ◽  
Gerard P. Sergeant
Keyword(s):  
Purinergic Receptor ◽  
Isometric Tension ◽  
P2x Receptor ◽  
Rt Pcr ◽  
Patch Clamp Technique ◽  
Real Time Quantitative Pcr ◽  
Cell Current ◽  
Adrenoceptor Agonists ◽  
Cyclase Activator ◽  
Adenylate Cyclase Activator

β3-Adrenoceptor (β3-AR) agonists are used to treat overactive bladder syndrome; however, their mechanism of action has not been determined. The aims of this study were to compare the effects of β3-AR agonists on cholinergic versus purinergic receptor-mediated contractions of the detrusor and to examine the mechanisms underlying inhibition of the purinergic responses by β3-AR agonists. Isometric tension recordings were made from strips of murine detrusor and whole cell current recordings were made from freshly isolated detrusor myocytes using the patch-clamp technique. Transcriptional expression of exchange protein directly activated by cAMP (EPAC) subtypes in detrusor strips was assessed using RT-PCR and real-time quantitative PCR. The β3-AR agonists BRL37344 and CL316243 (100 nM) inhibited cholinergic nerve-mediated contractions of the detrusor by 19 and 23%, respectively, but did not reduce contractions induced by the cholinergic agonist carbachol (300 nM). In contrast, BRL37344 and CL316243 inhibited purinergic nerve-mediated responses by 55 and 56%, respectively, and decreased the amplitude of contractions induced by the P2X receptor agonist α,β-methylene ATP by 40 and 45%, respectively. The adenylate cyclase activator forskolin inhibited purinergic responses, and these effects were mimicked by a combination of the PKA activator N6-monobutyryl-cAMP and the EPAC activator 8-pCPT-2′- O-methyl-cAMP-AM (007-AM). Application of ATP (1 μM) evoked reproducible P2X currents in isolated detrusor myocytes voltage-clamped at −60 mV. These responses were reduced in amplitude in the presence of BRL37344 and also by 007-AM. This study demonstrates that β3-AR agonists reduce postjunctional purinergic responses in the detrusor via a pathway involving activation of the cAMP effector EPAC.

176. INHIBITION OF PORCINE OOCYTE NUCLEAR MATURATION IN VITRO USING A PHOSPHODIESTERASE INHIBITOR AND AN ADENYLATE CYCLASE ACTIVATOR

2010 ◽  
Vol 22 (9) ◽  
pp. 94
Author(s):  
M. Bertoldo ◽  
T. Sellens ◽  
C. G. Grupen
Keyword(s):  
Adenylate Cyclase ◽  
Phosphodiesterase Inhibitor ◽  
Phosphodiesterase Inhibitors ◽  
Extended Period ◽  
Nuclear Maturation ◽  
Porcine Oocyte ◽  
Defined Culture ◽  
Cyclase Activator ◽  
Adenylate Cyclase Activator

Asynchronous nuclear and cytoplasmic maturation is thought to contribute to poor embryo production in vitro. Nuclear arrest is mediated by cAMP and can be maintained within the oocyte using non-specific phosphodiesterase inhibitors (3-isobutyl-1-methylxanthine ; IBMX) and the adenylate cyclase activator forskolin (FSK) (1). The aim of this study was to investigate the effect of IBMX and FSK supplementation on porcine oocyte nuclear maturation during COC recovery and IVM using a defined culture system. In all experiments, cAMP modulators were added to Hepes-buffered media held in collection tubes. COCs recovered from 3–5 mm diameter follicles of prepubertal ovaries were cultured in basic maturation media in the absence of FSH. Nuclear maturation was assessed using orcein dye. In Experiment 1, IVM media was supplemented with 0, 50 or 500 µM IBMX. In Experiment 2, IVM media was supplemented with 0, 5, 10, 50 and 100µM FSK. In Experiment 3, IVM medium was supplemented with combinations of IBMX and FSK to give the treatments; control, 50IBMX/50FSK, 50IBMX/100FSK, 500IBMX/50FSK and 500IBMX/100FSK. Nuclear maturation was assessed at 0, 2, 4 and 18 h after the onset of IVM. At 18 h of culture, there were no differences in the proportion of oocytes supplemented with 0, 50 or 500 µM IBMX reaching MII. Incubation with 10, 50 or 100 µM FSK resulted in 8-16% of oocytes at MII at 18 h compared to the other groups (25–29%; P < 0.001). The combinations of IBMX and FSK resulted in greater proportions (86–98%) of oocytes remaining at the GV stage at 18 h compared to the control (16%; P < 0.001). There were no differences in the proportion of oocytes remaining at the GV stage at the earlier time points (P > 0.05). The results demonstrate that these cAMP modulators, in combination, are highly effective in maintaining porcine oocyte meiotic arrest in vitro for an extended period. (1) Albuz FK et al., Proceedings of the 25th Annual Meeting of ESHRE, Amsterdam, The Netherlands, 2009.

Gastrin-releasing peptide directly releases pepsinogen from guinea pig chief cells

1990 ◽  
Vol 259 (5) ◽  
pp. G760-G766 ◽  
Author(s):  
S. Fiorucci ◽  
K. E. McArthur
Keyword(s):  
Substance P ◽  
Guinea Pig ◽  
Receptor Antagonist ◽  
Fold Increase ◽  
Chief Cells ◽  
Gastrin Releasing Peptide ◽  
Basal Release ◽  
Substance P Receptor ◽  
Bombesin Receptor ◽  
Muscarinic Agents

Gastrin-releasing peptide (GRP) and bombesin can stimulate pepsinogen release by both gastrin-dependent and -independent mechanisms. Using isolated guinea pig gastric chief cells, we determined that GRP can act directly on the guinea pig chief cell to cause pepsinogen release. GRP and bombesin stimulated a 2.5- to 3-fold increase in pepsinogen release above basal release. Substance P also stimulated a small but significant increase in pepsinogen release. No gastrin immunoreactivity was detected in the supernatants of cells stimulated with up to 1 microM GRP or bombesin or 1 mM carbachol. GRP-stimulated pepsinogen release was completely inhibited by GRP/bombesin receptor agonists as well as substance P receptor antagonist but not by antagonists to receptors for gastrin, the octapeptide of cholecystokinin (CCK-8), secretin, vasoactive intestinal peptide (VIP), or muscarinic agents. Substance P-stimulated pepsinogen release was completely inhibited by substance P receptor antagonist but not by GRP/bombesin receptor antagonists. An additive effect on pepsinogen release was seen when GRP was combined with maximally effective concentrations of adenosine 3',5'-cyclic monophosphate (cAMP)-mediated agents (VIP, secretin, 8-BrcAMP) but not with calcium-mediated agents (carbachol, CCK-8, gastrin). These results indicate that GRP can directly stimulate pepsinogen release from guinea pig chief cells by a specific GRP receptor that mobilizes intracellular calcium.

Conditions to optimise the developmental competence of immature equine oocytes

2020 ◽  
Vol 32 (11) ◽  
pp. 1012
Author(s):  
Elizabeth S. Metcalf ◽  
Keith R. Masterson ◽  
David Battaglia ◽  
Jeremy G. Thompson ◽  
Robert Foss ◽  
...  
Keyword(s):  
Adenylate Cyclase ◽  
Culture Media ◽  
Phosphodiesterase Inhibitor ◽  
Developmental Competence ◽  
Sodium Ascorbate ◽  
Blastocyst Formation ◽  
Developmental Potential ◽  
Holding Temperature ◽  
Cyclase Activator ◽  
Adenylate Cyclase Activator

Optimising the developmental potential of immature equine oocytes and invitro-produced (IVP) embryos was explored through modifications of established media and holding temperature. In Experiment 1, delaying spontaneous resumption of meiosis through the process of simulated physiological oocyte maturation with the addition of the adenylate cyclase activator forskolin (50µM) and the phosphodiesterase inhibitor 3-isobutyl-1-methylxanthine (100µM) to overnight holding medium before maturation improved blastocyst production (P&lt;0.05). In Experiment 2, the blastocyst production rate was increased significantly when cumulin (100ng mL−1) was added to the overnight holding or culture media (P&lt;0.05). In Experiment 3, immature oocytes held overnight at 16°C before maturation had improved developmental competence than those held at 20°C and 5°C (P&lt;0.05). There was no difference between maturation rates, but blastocyst formation per cleaved oocyte was significantly greater in oocytes held overnight at 16°C than at 20°C or 5°C. Furthermore, blastocyst formation per recovered oocyte and per fertilised oocyte was greater when oocytes were held before maturation at 16°C than at 5°C (P&lt;0.05). In Experiment 4, the addition of sodium ascorbate (AC; 50µg mL−1) to the maturation and/or culture media of oocytes and IVP embryos did not improve blastocyst production, but did appear to lower cleavage rates compared with oocytes and embryos cultured without AC.

In vitro modulation of porcine Leydig cell steroidogenesis by phorbol-12-myristate-13-acetate and 1,2-dioctanoylglycerol

1990 ◽  
Vol 122 (1) ◽  
pp. 101-106 ◽  
Author(s):  
Joseph T. French ◽  
Thomas H. Welsh
Keyword(s):  
Protein Kinase C ◽  
Protein Kinase ◽  
Leydig Cell ◽  
Leydig Cells ◽  
Primary Cultures ◽  
Kinase C ◽  
Protein Kinase C Activators ◽  
Cyclase Activator ◽  
Adenylate Cyclase Activator

Abstract Serum-free primary cultures of neonatal (1-day-old) porcine Leydig cells were used to study the effects of phorbol-12-myristate-13-acetate and 1,2-dioctanoylglycerol on testosterone and pregnenolone production. Phorbol-12-myristate-13-acetate alone from 0.001-10 μmol/l stimulated testosterone and pregnenolone production, whereas 1,2-dioctanoylglycerol alone had no effect on steroid production, relative to control. Phorbol12-myristate-13-acetate and 1,2-dioctanoylglycerol each inhibited pLH-stimulated testosterone and pregnenolone production. To further clarify the influence of these protein kinase C activators on steroidogenesis, cultured Leydig cells were treated with either phorbol-12-myristate-13-acetate or 1,2-dioctanoylglycerol plus forskolin (an adenylate cyclase activator). Both phorbol-12-myristate-13-acetate and 1,2-dioctanoylglycerol inhibited forskolin-stimulated testosterone production. Phorbol-12-myristate-13-acetate had no effect on forskolin-stimulated pregnenolone production and only the highest concentration of 1,2-dioctanoylglycerol (100 μmol/l) inhibited forskolin-stimulated production of pregnenolone. These data demonstrate that porcine Leydig cell steroidogenesis can be modulated by interactions of the protein kinase C and protein kinase A second messenger systems.

Localization of bombesin and GRP (gastrin releasing peptide) sequences in gut nerves or endocrine cells

1982 ◽  
Vol 76 (4) ◽  
pp. 457-467 ◽  
Author(s):  
R. Buffa ◽  
I. Solovieva ◽  
R. Fiocca ◽  
S. Giorgino ◽  
G. Rindi ◽  
...  
Keyword(s):  
Endocrine Cells ◽  
Gastrin Releasing Peptide

Direct measurement of nitric oxide release from vascular endothelial cells

1996 ◽  
Vol 81 (2) ◽  
pp. 774-779 ◽  
Author(s):  
J. P. Guo ◽  
T. Murohara ◽  
M. Buerke ◽  
R. Scalia ◽  
A. M. Lefer
Keyword(s):  
Nitric Oxide ◽  
Endothelial Cells ◽  
Methyl Ester ◽  
Vascular Endothelial Cells ◽  
Calmodulin Antagonist ◽  
Nitric Oxide Release ◽  
A 23187 ◽  
No Release ◽  
Basal Release ◽  
Isolated Rat

A nitric oxide (NO)-selective electrode was used to directly measure NO release from isolated rat aortic endothelium and cultured rat aortic endothelial cells (RAECs). Basal release of NO was significantly attenuated by a NO synthase inhibitor NG-nitro-L-arginine methyl ester (1 mM) to 42 +/- 14 pmol/1 x 10(5) cells (P < 0.01). The basal release of NO was also significantly inhibited by a calmodulin antagonist W-7 at 15 microM (P < 0.01). L-Arginine (1 mM), significantly stimulated NO release (P < 0.05 vs. control basal release). Stimulation of cultured RAECs with two endothelium-dependent vasodilators, acetylcholine (100 nM) and A-23187 (1 microM), significantly increased NO release [574 +/- 112 pmol/1 x 10(5) cells (n = 5) and 658 +/- 119 pmol/1 x 10(5) cells (n = 5) in acetylcholine- and A-23187-stimulated RAECs, respectively]. Basal release of NO was also detectable in isolated rat aortic rings with intact endothelium. NO release was significantly attenuated by NG-nitro-L-arginine methyl ester and augmented by human superoxide dismutase. These data indicate the physiological usefulness of the amperometric measurement of NO employing a NO-specific electrode in biological systems.

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