Characterization of a new CCK antagonist, L364,718: in vitro and in vivo studies

1988 ◽  
Vol 255 (3) ◽  
pp. G261-G266 ◽  
Author(s):  
D. S. Louie ◽  
J. P. Liang ◽  
C. Owyang
Keyword(s):  
Pancreatic Juice ◽  
Competitive Inhibition ◽  
In Vivo Studies ◽  
Amylase Secretion ◽  
Receptor Antagonists ◽  
Amylase Release ◽  
Pancreatic Acini ◽  
Cck Antagonist

In this study we examined a novel, orally effective, nonpeptidal cholecystokinin (CCK) antagonist, 3S(-)-N-(2,3-dihydro-1-methyl-2-oxo-5-phenyl-1H-1,4-benzodiazepine-3-yl) -1H-indole-2-carboxamide (L364,718) on CCK-induced amylase release. We used isolated rat pancreatic acini and incubated them with CCK-8 with or without various CCK receptor antagonists. L364,718 (3-100 nM), proglumide (1-10 mM), and the proglumide derivative CR1409 (1-30 microM) each caused a progressive rightward shift in the CCK-8 dose-response curve without a change in maximal amylase secretion. Plotting the data according to the method of Schild indicated competitive inhibition. The Kis for L364,718, CR1409, and proglumide were 0.25 nM, 0.15 microM, and 0.5 mM, respectively. Thus L364,718 was 600-fold more potent than CR1409 and 2,000,000-fold more potent than proglumide in inhibiting CCK-8-induced amylase release. Inhibition of 125I-Bolton-Hunter-CCK-8 binding to acini by these receptor antagonists had a similar rank potency. L364,718 was tested against other pancreatic exocrine secretagogues and was effective against agonists that only act through the CCK receptor. In rats, diversion of pancreatic juice from the duodenum stimulated pancreatic amylase output and elevated plasma CCK levels. The stimulated amylase secretion was blocked by intravenous administration of L364,718. Furthermore, extracted plasma CCK from rats with diversion of pancreatic juice from the duodenum stimulated amylase release from pancreatic acini, and this stimulation was blocked by addition of L364,718.(ABSTRACT TRUNCATED AT 250 WORDS)

Comparative effects of CCK receptor antagonists on rat pancreatic secretion in vivo

1989 ◽  
Vol 256 (1) ◽  
pp. G150-G157 ◽  
Author(s):  
M. Niederau ◽  
C. Niederau ◽  
G. Strohmeyer ◽  
J. H. Grendell
Keyword(s):  
Pancreatic Secretion ◽  
Rank Order ◽  
Amylase Secretion ◽  
Receptor Antagonists ◽  
Pancreatic Acini ◽  
Cck Receptors ◽  
Competitive Kinetics ◽  
Cck Receptor Antagonists

The present experiments evaluate in vivo effects of recently described cholecystokinin (CCK) receptor antagonists on rat pancreatic secretion. Pancreaticobiliary secretion was studied after bile duct cannulation in anesthetized rats. After two basal 10-min fractions were selected, secretion was stimulated by intravenous caerulein (0.1-30.0 micrograms/kg) or secretin, and collected for seven further 10-min fractions. Peptide antagonists (CR 1409, CR 1392, and CR 1505) and nonpeptide antagonists (asperlicin and L364,718) were given intravenously 10 min before agonists. Increasing doses of antagonists gradually reduced secretion of protein and enzymes stimulated by submaximal and maximal doses of caerulein. The antagonists did not alter nonstimulated or secretin-stimulated secretion, indicating their specificity for the CCK receptor. Except for proglumide and asperlicin, all antagonists were able to abolish caerulein-stimulated pancreatic secretion, as evaluated by the mean integrated 1-h response to a near-maximal dose of caerulein. The caerulein dose-response curve was gradually shifted to the right by increasing doses of CR 1409, indicating competitive-like kinetics. Inhibition of secretion due to supramaximal doses of caerulein, however, could be reversed by doses of CR 1409 smaller than expected from extrapolating truely competitive kinetics from an in vitro situation to the in vivo situation. The rank order of potency of the compounds to antagonize caerulein-stimulated secretion in vivo agreed with their relative potencies to antagonize caerulein-stimulated amylase secretion from pancreatic acini in vitro as well as with their affinity to bind to peripheral CCK receptors in vitro.(ABSTRACT TRUNCATED AT 250 WORDS)

A new biological contribution of cyclo(His-Pro) to the peripheral inhibition of pancreatic secretion

1997 ◽  
Vol 273 (6) ◽  
pp. E1127-E1132 ◽  
Author(s):  
Pascal Fragner ◽  
Olivier Presset ◽  
Nicole Bernad ◽  
Jean Martinez ◽  
Claude Roze ◽  
...  
Keyword(s):  
Pancreatic Secretion ◽  
Biological Activities ◽  
Systemic Circulation ◽  
Inhibitory Effect ◽  
Pancreatic Acinar Cells ◽  
Exocrine Pancreatic Secretion ◽  
Amylase Release ◽  
Pancreatic Acini

The tripeptide pyro-Glu-His-Pro-NH2[thyrotropin-releasing hormone (TRH)] was isolated from the hypothalamus as a thyrotropin-releasing factor. It has a broad spectrum of central nervous system-mediated actions, including the stimulation of exocrine pancreatic secretion. TRH is also synthesized in the endocrine pancreas and found in the systemic circulation. Enzymatic degradation of TRH in vivo produces other bioactive peptides such as cyclo(His-Pro). Because of the short half-life of TRH and the stability of cyclo(His-Pro) in vivo, we postulated that at least part of the peripheral TRH effects on the exocrine pancreatic secretion may be attributed to cyclo(His-Pro), which has been shown to have other biological activities. This study determines in parallel the peripheral effects of TRH and cyclo(His-Pro) as well as the putative contribution of other TRH-related peptides on exocrine pancreatic secretion in rats. TRH and its metabolite cyclo(His-Pro) dose dependently inhibited 2-deoxy-d-glucose (2-DG)-stimulated pancreatic secretion. TRH and all the related peptides tested had no effect on the basal and cholecystokinin-stimulated amylase release from pancreatic acinar cells in vitro. These data indicate that cyclo(His-Pro) mimics the peripheral inhibitory effect of TRH on 2-DG-stimulated exocrine pancreatic secretion. This effect is not detected on isolated pancreatic acini. Our findings provide a new biological contribution for cyclo(His-Pro) with potential experimental and clinical applications.

Amylase secretion by isolated pancreatic acini after acute cholecystokinin treatment in vivo

1984 ◽  
Vol 246 (4) ◽  
pp. G419-G425 ◽  
Author(s):  
M. Otsuki ◽  
Y. Okabayashi ◽  
A. Ohki ◽  
S. R. Hootman ◽  
S. Baba ◽  
...  
Keyword(s):  
Binding Sites ◽  
Amylase Secretion ◽  
Response Curves ◽  
Amylase Release ◽  
Pancreatic Acini ◽  
Cholecystokinin Receptor ◽  
Isolated Pancreatic Acini ◽  
And Control ◽  
Quinuclidinyl Benzilate

A single dose of synthetic cholecystokinin octapeptide (CCK8, 5 micrograms/kg) in a depot carrier was injected subcutaneously into rats 2 and 14 h before the removal of the pancreas and the preparation of isolated pancreatic acini. CCK8 treatment induced no significant change in body weight or total amount of pancreatic DNA, but pancreatic weight, total pancreatic protein and amylase, and the concentration of amylase and total protein relative to DNA were significantly decreased. In acini prepared from CCK8-pretreated rats, responsiveness to maximal and supramaximal concentrations of CCK8 was significantly increased, irrespective of whether the amount of amylase released was expressed relative to DNA or calculated as a percentage of the acinar content. The dose-response curves for CCK8 were similarly shaped in both CCK8-pretreated and control rats but shifted threefold toward higher concentrations of CCK8 2 or 14 h after CCK8 treatment. Specific 125I-CCK binding was significantly increased only for high-affinity binding sites. Although these observations suggest that alterations in pancreatic amylase release could be due to changes at the cholecystokinin receptor, the secretory responsiveness to maximal and supramaximal concentrations of carbachol was also increased without any change in the sensitivity. Moreover, in contrast to the cholecystokinin receptor, there was no change in the number of muscarinic receptors or in their affinity for either agonists or antagonists measured with [3H]quinuclidinyl benzilate.(ABSTRACT TRUNCATED AT 250 WORDS)

Proglumide analogues: potent cholecystokinin receptor antagonists

1985 ◽  
Vol 249 (2) ◽  
pp. G214-G220 ◽  
Author(s):  
R. T. Jensen ◽  
R. B. Murphy ◽  
M. Trampota ◽  
L. H. Schneider ◽  
S. W. Jones ◽  
...  
Keyword(s):  
Cell Function ◽  
Fold Increase ◽  
Close Correlation ◽  
Hydroxyl Group ◽  
Amylase Secretion ◽  
Receptor Antagonists ◽  
Inhibitory Potency ◽  
Amylase Release ◽  
Pancreatic Acini ◽  
Cholecystokinin Receptor

Proglumide [N-(benzoyl)-L-glutamic acid-1-di-n-propylamide] is a specific cholecystokinin receptor antagonist. In the present study we synthesized various analogues of proglumide and used pancreatic acini from guinea pig pancreas to examine the abilities of these analogues to function as cholecystokinin receptor antagonists. Each analogue inhibited cholecystokinin octapeptide-stimulated amylase secretion but did not stimulate amylase secretion when present alone. There was a close correlation between the ability of a particular analogue to inhibit the action of cholecystokinin on acinar cell function and its ability to inhibit binding of 125I-cholecystokinin. Structure-function studies demonstrated that neither the dipropylamide nor the benzoyl moieties are essential for inhibiting the action of cholecystokinin but that both groups are important in determining the inhibitory potency. Replacing the dipropylamide group with a hydroxyl group caused a 13-fold decrease in potency. Replacing the benzoyl moiety by an acetyl group caused a 30- to 40-fold decrease in inhibitory potency, whereas replacing the benzoyl moiety by a p-chlorophenoxyacetyl or phenoxyacetyl moiety caused a 75-fold increase in potency. Replacing both the dipropylamide moiety with a hydroxyl group and the benzoyl moiety with a phenoxyacetyl group resulted in a 5-fold decrease in inhibitory potency. Inhibition of cholecystokinin-stimulated amylase release by both the phenoxyacetyl and p-chlorophenoxyacetyl analogues was competitive in nature, fully reversible, and specific for those secretagogues that interact with the cholecystokinin receptor.(ABSTRACT TRUNCATED AT 250 WORDS)

Action of pancreatic polypeptide on rat pancreatic secretion: in vivo and in vitro

1985 ◽  
Vol 249 (4) ◽  
pp. G489-G495 ◽  
Author(s):  
D. S. Louie ◽  
J. A. Williams ◽  
C. Owyang
Keyword(s):  
Protein Secretion ◽  
Pancreatic Polypeptide ◽  
Pancreatic Secretion ◽  
Specific Binding ◽  
Pancreatic Enzyme ◽  
Enzyme Secretion ◽  
Amylase Release ◽  
Pancreatic Acini

The biological activity of bovine pancreatic polypeptide (BPP) on rat exocrine pancreatic secretion was compared in vivo and in vitro. In anesthetized rats prepared with a bile-pancreatic duct cannula, BPP inhibited cholecystokinin (CCK)-stimulated (10 IDU . kg-1 X h-1) protein secretion in a dose-related manner (P less than 0.001). CCK, from 5-20 IDU . kg-1 X h-1, did not alter the degree of inhibition by BPP at 40 micrograms . kg-1 X h-1, suggesting a nonsurmountable inhibition. Analogues of BPP, including rat pancreatic polypeptide, neuropeptide Y, peptide YY, and the C-terminal hexapeptide of PP, also inhibited CCK-stimulated protein secretion. To determine whether BPP acts directly on acinar cells to suppress enzyme secretion, in vitro studies were performed. BPP and its analogues did not suppress octapeptide of CCK (CCK-8)-stimulated amylase release from either isolated rat pancreatic acini or preparations of pancreatic lobules. Specific binding of 125I-BPP to pancreatic acini was also not observed. From our data we conclude that BPP acts to inhibit pancreatic enzyme secretion in the rat in a noncompetitive manner. Absence of an effect by BPP or its analogues in vitro coupled with an absence of 125I-BPP binding to acini suggest that the inhibitory action of PP on exocrine pancreatic function is mediated by indirect mechanisms.

Effects of synthetic cyclic AMP analogs on amylase exocytosis from rat pancreatic acini

1994 ◽  
Vol 72 (10) ◽  
pp. 1138-1147 ◽  
Author(s):  
Claus Schäfer ◽  
Hanna Steffen ◽  
Hartmut Printz ◽  
Burkhard Göke
Keyword(s):  
Protein Kinases ◽  
Vasoactive Intestinal Polypeptide ◽  
Inhibitory Effect ◽  
Amylase Secretion ◽  
Amylase Release ◽  
Pancreatic Acini ◽  
Dependent Protein ◽  
Maximal Induction ◽  
Intestinal Polypeptide

Diastereomers of adenosine 3′,5′-phosphorothioate activate cAMP-dependent protein kinases (cAMP-PK) in vitro. We found that these compounds are highly selective tools to monitor cAMP-dependent PKA activation and its effect on amylase exocytosis from pancreatic acini. In permeabilized rat acinar cells, (Sp)-cAMPS dose-dependently stimulated amylase secretion, while (Rp)-cAMPS inhibited (Sp)-cAMPS-induced amylase release. In intact rat acini, 8-Br-(Sp)-cAMPS reduced the secretory responses to secretin, vasoactive intestinal polypeptide (VIP), 8-Br-cAMP, and 8-Br-(Sp)-cAMPS, but not to cerulein. Another derivative, dibutyryl-(Rp)-cAMPS, induced a small inhibitory effect against 8-Br-(Sp)-cAMPS and VIP, which was overlapped by an unspecific stimulatory effect on amylase exocytosis induced by the degradation product butyrate. Furthermore, (Sp)-5,6-dichloro-1-β-D-ribofuranosylbenzimidazole-3′,5′-monophosphorothioate ((Sp)-5,6-DCl-cBIMPS), a specific cAMP-PK activator, induced a maximal induction of cAMP-PK activity, but its stimulation of amylase secretion was less than that by secretin. (Sp)-5,6-DCl-cBIMPS regulated the phosphorylation of several proteins, which were also affected by secretin. However, secretin had additional effects. Its action was most likely mediated by a dual effect on the cAMP and the calcium pathway. Our results indicate that the cAMP-dependent pathway is involved in amylase exocytosis from rat pancreatic acini.Key words: secretin, vasoactive intestinal polypeptide, (Rp)-cAMPS, (Sp)-cAMPS, 8-Br-(Rp)-cAMPS, 8-Br-(Sp)-cAMPS, dibutyryl-(Rp)-cAMPS, (Sp)-5,6-DCl-cBIMPS, cAMP-dependent protein kinases, pancreatic acini, amylase secretion.

Amylase secretion by isolated pancreatic acini after chronic cholecystokinin treatment in vivo

1983 ◽  
Vol 244 (6) ◽  
pp. G683-G688 ◽  
Author(s):  
M. Otsuki ◽  
J. A. Williams
Keyword(s):  
Amylase Secretion ◽  
Response Curves ◽  
Amylase Release ◽  
Pancreatic Acini ◽  
Subcutaneous Injections ◽  
Cck Receptors ◽  
Wet Weight ◽  
Initial Content ◽  
And Control

Rats were given subcutaneous injections of synthetic cholecystokinin octapeptide (CCK8, 5 micrograms/kg) in a depot carrier twice daily for 7–14 days. The pancreatic wet weight increased by 20.6 and 30.9% in the rats treated with CCK8 for 7 and 14 days, respectively. The increase in pancreatic weight was associated with an increase in the amount of protein per DNA, indicating hypertrophy of the acinar cells, and with an increase in the total amount of pancreatic DNA. Moreover, CCK administration also increased the amylase content per DNA. In acini prepared from CCK8-treated rats, responsiveness to CCK8 was increased when amylase release was expressed relative to DNA but was decreased when calculated as the percentage of the initial content in the acini. The dose-response curves for CCK8 were similarly shaped in both CCK8-treated and control rats, but they were shifted 3- to 10-fold toward higher concentrations of CCK8 after 7 and 14 days of CCK8 treatment. There were no major changes in the affinity and capacity of CCK receptors determined by studying the binding of radioiodinated CCK, suggesting that alterations in pancreatic amylase release were due to changes at a postreceptor loci. In support of this hypothesis, the secretory response to carbachol, known to act on a different receptor but by a common intracellular mechanism, was altered in a manner identical to the response to CCK8. Thus chronic stimulation with CCK sufficient to induce pancreatic hypertrophy does not greatly alter CCK receptors and induces only moderate postreceptor desensitization.

scholarly journals A Cytosolic Splice Variant of Cab45 Interacts with Munc18b and Impacts on Amylase Secretion by Pancreatic Acini

2007 ◽  
Vol 18 (7) ◽  
pp. 2473-2480 ◽  
Author(s):  
Patrick P.L. Lam ◽  
Kati Hyvärinen ◽  
Maria Kauppi ◽  
Laura Cosen-Binker ◽  
Saara Laitinen ◽  
...  
Keyword(s):  
Splice Variant ◽  
Amylase Secretion ◽  
Dependent Manner ◽  
Vitro Assay ◽  
Amylase Release ◽  
Pancreatic Acini ◽  
Protein Factor ◽  
Ef Hand ◽  
Syntaxin 3

We identified in a yeast two-hybrid screen the EF-hand Ca2+-binding protein Cab45 as an interaction partner of Munc18b. Although the full-length Cab45 resides in Golgi lumen, we characterize a cytosolic splice variant, Cab45b, expressed in pancreatic acini. Cab45b is shown to bind45Ca2+, and, of its three EF-hand motifs, EF-hand 2 is demonstrated to be crucial for the ion binding. Cab45b is shown to interact with Munc18b in an in vitro assay, and this interaction is enhanced in the presence of Ca2+. In this assay, Cab45b also binds the Munc18a isoform in a Ca2+-dependent manner. The endogenous Cab45b in rat acini coimmunoprecipitates with Munc18b, syntaxin 2, and syntaxin 3, soluble N-ethylmaleimide-sensitive factor attachment protein receptors with key roles in the Ca2+-triggered zymogen secretion. Furthermore, we show that Munc18b bound to syntaxin 3 recruits Cab45b onto the plasma membrane. Importantly, antibodies against Cab45b are shown to inhibit in a specific and dose-dependent manner the Ca2+-induced amylase release from streptolysin-O–permeabilized acini. The present study identifies Cab45b as a novel protein factor involved in the exocytosis of zymogens by pancreatic acini.

Effects of chloroquine and methylamine on lysosomal enzyme secretion by rat pancreas

1992 ◽  
Vol 262 (3) ◽  
pp. G439-G444
Author(s):  
T. Hirano ◽  
A. Saluja ◽  
P. Ramarao ◽  
M. M. Lerch ◽  
M. L. Steer
Keyword(s):  
Lysosomal Enzyme ◽  
Cathepsin B ◽  
Secretory Pathway ◽  
Amylase Secretion ◽  
Zymogen Granule ◽  
Lysosomal Hydrolases ◽  
Pancreatic Acini ◽  
Lysosomotropic Agents

In vivo pancreatic secretion of the lysosomal hydrolase cathepsin B was found to be increased by infusion of the secretagogue caerulein. The basal as well as caerulein-stimulated in vivo rate of cathepsin B was further increased by infusion of either chloroquine or methylamine while neither the basal nor the secretagogue-stimulated rates of amylase secretion were altered by the lysosomotropic agents. These observations indicate that neutralization of the acidic prelysosomal compartment by administration of lysosomotropic agents results in lysosomal enzyme entry, by default, into the regulated secretory pathway. In vitro stimulation of pancreatic acini with caerulein was also found to stimulate cathepsin B secretion. That in vitro rate of cathepsin B secretion stimulated by caerulein was not increased in acini prepared from animals infused with caerulein, chloroquine, or methylamine, but the in vitro rate of cathepsin B secretion stimulated by caerulein was increased in acini prepared from animals infused with caerulein plus either chloroquine or methylamine. Under these conditions, redistribution of cathepsin B from the lysosome-enriched to the zymogen granule-enriched subcellular fraction was noted, and lysosomal enzyme-containing organelles became increasingly fragile. These observations indicate that in vivo secretagogue stimulation increases the degree of diversion of lysosomal hydrolases into the regulated secretory compartment when the prelysosomal compartment has been neutralized with lysosomotropic agents.

Possible site of action of CGRP antagonists in migraine

Cephalalgia ◽  
2011 ◽  
Vol 31 (6) ◽  
pp. 748-750 ◽  
Author(s):  
Peer Tfelt-Hansen ◽  
Jes Olesen
Keyword(s):  
In Vivo Studies ◽  
Cgrp Receptor ◽  
Receptor Antagonists ◽  
Calcitonin Gene ◽  
Site Of Action ◽  
Cgrp Receptor Antagonists ◽  
The Central Nervous System ◽  
High Doses

Background: The calcitonin gene-related peptide (CGRP) receptor antagonists olcegepant and telcagepant are very potent drugs. Both are effective in migraine but in doses much higher than would be predicted from receptor binding and other in vitro results. This could perhaps suggest an effect of CGRP antagonists behind the blood-brain barrier (BBB), i.e. in the central nervous system (CNS). Methods: Comparison of doses needed for CGRP blocking effect in vitro with dose needed in vivo in man and monkeys. Discussion of these doses in relation to doses needed for anti-migraine activity. Results: In vivo studies in monkeys and man showed that high doses compared to doses needed in vitro are needed to block capsaicin-induced in skin blood flow, a CGRP-mediated reaction. These doses are close to those needed for anti-migraine activity. Conclusion: The apparently high doses of CGRP receptor antagonists, olcegepant and telcagepant needed for anti-migraine effect are not so high after all. They do not allow a conclusion as to whether CGRP antagonists act on peripheral sites or central sites in migraine.

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