Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:3.4.24.3 (collagenase)
18,340 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Effect of cholinergic stimulation on the release of digestive enzymes from isolated rat pancreatic acini prepared by collagenase digestion was investigated. The release of enzymes (amylase, chymotrypsinogen, lipase) increased linearly with the time of incubation in the absence of secretagogues. Carbachol, a muscarinic agonist, induced a remarkable increase in the release of these enzymes. This carbachol-stimulated amylase release showed a biphasic curve, and its maximal response was observed at 10(-5) M. The release patterns of chymotrypsinogen and lipase were similar to that of amylase. This carbachol-stimulated amylase release was inhibited by atropine in a dose-dependent manner, with an ED50 value of 1.17 X 10(-8) M. Other cholinergic agonists such as methacholine also stimulated the release of these enzymes, and these increases in the release were inhibited by atropine. Scatchard analysis for [3H]quinuclidinyl benzilate ([3H]QNB) binding to isolated pancreatic acini revealed that the binding site of [3H]QNB was a single component with a Kd value of 0.09 nM and a Bmax value of 89.3 fmol/mg protein, respectively. The effect of other cholinergic antagonists, pirenzepine and AF-DX 116 (11-[[2-[(diethylamino) methyl]1-piperidinyl]acetyl]-5,11-dihydro- 6H-pyrido[2,3-b][1,4]-benzodiazepine-6-one), on pancreatic acini was also investigated. Based on these results, it has been concluded that isolated rat pancreatic acini have muscarinic receptors and are useful for analyzing the mechanism of pancreatic enzyme secretion.
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PMID:Release of digestive enzymes from isolated rat pancreatic acini following muscarinic stimulation: a comparative study with enzyme release and receptor binding. 171 8

We undertook the present studies to explore the mechanisms by which carbachol inhibits the release of somatostatin-like immunoreactivity (SLI) from D cells. D cells were isolated from canine fundic mucosa by collagenase/EDTA dispersion followed by counterflow elutriation. Carbachol inhibited the release of SLI induced by forskolin, dibutyryl 3':5' cyclic adenosine monophosphate (cAMP), pentagastrin (PG), and 12-0-tetradecanoyl-phorbol-13-acetate in a fashion that could be prevented by pertussis toxin (PT) pretreatment of the D cells. Pertussis toxin also prevented the carbachol-induced inhibition of forskolin-stimulated cAMP generation and PG-stimulated [Ca2+]i mobilization. These data indicate that pertussis toxin sensitive inhibitory guanine nucleotide binding proteins mediate many of carbachol's inhibitory actions on D cells.
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PMID:Mechanisms for muscarinic inhibition of somatostatin release from canine fundic D cells. 197 5

Stimulation-induced changes in Cl- content and O2 consumption of collagenase-isolated rat parotid acini were measured. In less than 10 s, carbachol caused a net Cl- efflux, corresponding to approximately 50% of the Cl- content, and increased the O2 uptake by 100%. The increase was inhibitable by ouabain and was dependent on the presence of extracellular Ca2+. Furosemide reduced the unstimulated 36Cl- uptake and prevented the reuptake of Cl- after carbachol-induced release. This suggests that a cotransport system is operating in both the unstimulated and stimulated states. Furthermore, furosemide inhibited the stimulated ouabain-sensitive O2 uptake. Raising intracellular Ca2+ by the calcium ionophore A23187 evoked the same pattern of Cl- loss and O2 uptake as carbachol. Our results are compatible with the following hypothesis. Carbachol raises intracellular Ca2+, causing an increased Cl- permeability of the luminal membrane, resulting in a net Cl- efflux. A subsequently enhanced influx of Cl- and Na+ via a furosemide-sensitive cotransport system increases intracellular Na+. This stimulates the Na+-K+-ATPase and thereby the O2 consumption.
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PMID:Effects of Ca2+ and furosemide on Cl- transport and O2 uptake in rat parotid acini. 242 56

Smooth muscle cells were isolated from estrogenized rat myometrium by collagenase digestion. Electron microscopic examination and measurement of cell lengths by image-splitting micrometry were carried out after fixation with acrolein. Mean lengths of cells before and after isolation were 81.7 and 66.9 micron, respectively. Responses of cells were compared with contractions of isolated strips recorded isometrically. Effects of carbachol and KCl were examined in 2 mM Ca, 2 mM Ca + 4 mM EGTA, and 2 mM Ca + 10(-8) M nitrendipine solution. Carbachol and KCl produced concentration-dependent shortening of isolated cells maximal at 30 s after addition. The concentrations of carbachol required to produce shortenings were about 100-fold less than those required to produce isometric contractions; but no major difference was observed in the concentration dependence of cell shortening and isometric contraction produced by potassium-induced depolarization. In 2 mM Ca solution, there was a phasic response, followed by a tonic response such that more than 50% of maximum cell shortening was maintained for 10 min. However, in 2 mM Ca + 4 mM EGTA or 10(-8) M nitrendipine, the tonic contraction was abolished and cells rapidly relaxed after 30 s. If carbachol was added to cells after varying times in the EGTA-containing solution, the ability to initiate a contraction declined exponentially with a half-time of 160 s. Effects of depolarization by KCl were examined in 2 mM Ca plus nitrendipine and 2 mM Ca + 4 mM EGTA solution. Shortening occurred in 2 mM Ca solution by depolarization but not if nitrendipine was added. Though shortening was not observed in 2 mM Ca + 4 mM EGTA solution by KCl, subsequent addition of carbachol induced shortening. These results suggested that there was an intracellular Ca store site from which Ca was released by carbachol and which was not affected by depolarization in the absence of external Ca. No evidence was obtained that the contraction persists in Ca2+-free medium in isolated cells, which is in agreement with previous findings in small muscle strips in which only a similar transient response was obtained.
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PMID:Calcium and contractions of isolated smooth muscle cells from rat myometrium. 244 28

The neuropeptide galanin is present in intrapancreatic nerve fibers and is known to affect the secretion of the islet hormones. Its most potent effect is thereby the inhibition of insulin secretion. In the present study, we investigated whether galanin influences amylase secretion from isolated rat pancreatic acini. Acini were isolated by the collagenase digestion technique and incubated for 45 min in a Krebs-Henseleit medium with or without addition of the cholinergic agonist carbachol or the C-terminal octapeptide of cholecystokinin (CCK-8) in the presence or absence of galanin. Carbachol, at its optimal concentration (10(-5) M), stimulated amylase secretion to 11.8 +/- 0.5% of total amylase content compared to 4.3 +/- 0.3% in controls (p less than 0.001). Galanin, (10(-8)-10(-9) M), reduced the carbachol-induced amylase secretion to 10.4 +/- 0.3% (p less than 0.01). Galanin at concentration levels below 10(-10) M had no significant effect. At 10(-8) M, CCK-8 stimulated amylase secretion to 9.7 +/- 0.6% compared with 5.2 +/- 0.3% in controls (p less than 0.01). Galanin (10(-7) M) reduced this stimulation to 8.0 +/- 0.4% (p less than 0.05). Galanin did not affect basal amylase secretion. It is concluded that the intrapancreatic neuropeptide galanin weakly inhibits carbachol- and CCK-8-induced amylase secretion from isolated rat pancreatic acini. Thus, galanin has the capability to directly affect not only endocrine but also exocrine pancreatic secretion although its effect of inhibiting amylase secretion seems weak.
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PMID:Galanin inhibits amylase secretion from isolated rat pancreatic acini. 246 Aug 54

Recently, we have demonstrated that carbachol, a cholinergic agonist, stimulates the hydrolysis of phosphoinositides (PI) in the inner medullary (IM) slices from the rabbit kidney. In order to localize the effects of carbachol in the IM, we measured PI hydrolysis in IM collecting duct (CD) cells which form approximately 50% of the IM and play an important role in determining the final composition of the urine. The IMCD cells were prepared from IM slices of the rabbit kidney by treatment with collagenase followed by addition of water to lyse the cells other than IMCD cells. To measure PI hydrolysis, the IMCD cells were incubated with [3H]inositol for its incorporation into PI before measurement of inositol phosphates (IP) released and accumulated in the presence of LiCl which prevents the dephosphorylation of IP. Carbachol (1 mM) produced greater than 16-fold increase in the release of IP (from 1.53 +/- 1.34% in control to 26.26 +/- 4.59% in drug-treated) in the isolated IMCD cells. The effect was concentration-dependent with an EC50 (50% maximum effective concentration) of 4 microM carbachol. Carbachol-stimulated PI hydrolysis was blocked completely by 1 microM atropine, a muscarinic antagonist, and not by 1 microM hexamethonium, a nicotinic antagonist. The nicotinic agonist, 1,1-dimethyl-4-phenylpiperazinium iodide (1 mM), had no significant effect on PI hydrolysis in the IMCD cells. We conclude that the stimulation of PI hydrolysis by cholinergic agents in the IMCD cells occurs through their interaction with muscarinic receptors and this process may play a role in the diuretic and natriuretic effects of these agents.
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PMID:Cholinergic stimulation of phosphoinositide hydrolysis in renal medullary collecting duct cells. 253 9

A superfusion technique was developed as a model system for the study of stimulus-secretion coupling in collagenase-dispersed rat pancreatic acinar cells. Cells (10(7)) were combined with a slurry of Biogel P-4 beads and the mixture was decanted into a plastic column (1.5 cm X 8.5 cm) and perfused with Krebs-Ringer. Amylase activity was determined in sequentially collected effusate fractions and used to estimate the secretory rate. Carbachol, carbachol plus dibutyryl cyclic AMP, cholecystokinin-pancreozymin, and the ionophore A-23187 all stimulated a rapid increase in the rate of secretion. Cell integrity was unaffected by these stimulants as evidenced microscopically and by the lack of lactate dehydrogenase activity in the effusates. Enzymes secreted in response to secretagogues were collected, concentrated, and isoelectrofocused on polyacrylamide gels. A film detection technique was developed to localize amylase activity. The model system has the following advantages: (1) secreted proteolytic products are removed from the vicinity of cells, thereby preventing direct cellular damage and hydrolysis of peptide agonist; (2) the need to add trypsin inhibitors is eliminated and only a minimal addition of albumin (0.001%) is required, thus allowing the separation and distortion-free analysis of secreted proteins; (3) the perfusion conditions can be changed rapidly without disturbing the cells. The model described is therefore well suited to the study of both molecular and kinetic events involved in the enzyme secretory phenomenon in exocrine pancreas.
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PMID:A model system for the study of stimulus - enzyme secretion coupling in rat pancreatic acinar cells. 616 55

The role of calcium gating in cholinergic stimulation of the function of parietal cells was studied using cells isolated from canine fundic mucosa by treatment with collagenase and EDTA and enriched by velocity separation in an elutriator rotor. Monitoring the accumulation of [14C[ aminopyrine as an index of parietal cell response, stimulation by carbachol, but not by histamine, was highly dependent upon the concentration of extracellular calcium. Incubation of parietal cells in 0-.1 mM calcium, rather than the usual 1.8 mM concentration, reduced the response to 100 microM carbachol by 92 +/- 2%, whereas histamine stimulation was impaired by 28 +/- 5%. A similar reduction in extracellular calcium suppressed the response to gastrin (100 nM) by 67 +/- 7%. The impairment of cholinergic stimulation found at low extracellular calcium concentrations was rapidly reversed with the readdition of calcium. Lanthanum, which blocks calcium movement across membranes, caused a similar pattern of effects on secretagogue stimulation of aminopyrine accumulation, with 100 microM lanthanum suppressing carbachol stimulation by 83 +/- 2%. This concentration of lanthanum suppressed gastrin stimulation by 40 +/- 7% and histamine stimulation by only 12 +/- 9%. Carbachol, but not histamine nor gastrin, stimulated 45Ca++ uptake. The magnitude of carbachol-stimulated calcium uptake correlated with the parietal cell content of the fractions examined (r = 0.88), and was dose responsive over carbachol concentrations from 1 microM to 1 mM. Atropine (100 nM) caused surmountable inhibition, and these effects of carbachol and atropine on calcium uptake correlated with their effects on oxygen consumption (r = 0.93) and [14C]-aminopyrine accumulation (r = 0.90). Cells preloaded with 45Ca++ lost cellular calcium in a time-dependent fashion; however, this rate of egress was not accelerated by treatment with histamine, gastrin, or carbachol, thus failing to implicate mobilization of intracellular calcium as primary mechanism for activation of parietal cell function. These data indicate a close link between stimulation of parietal cell function and enhancement of calcium influx by cholinergic agents.
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PMID:Extracellular calcium and cholinergic stimulation of isolated canine parietal cells. 725 63

Rat submandibular gland was dissociated by enzymatic digestion with collagenase and hyaluronidase, followed by mild mechanical shearing and filtration through a nylon mesh. The dissociated cell populations contained predominantly groups of acinar cells which maintained their acinar arrangement. The morphological and functional viability of the cells was confirmed by electron microscopic examination and a normal secretory response to beta-adrenergic or cholinergic stimulation was observed. Both isoproterenol (IPR) and carbachol caused the fusion of secretory granules into large vacuoles which were also continuous with the lumen, and into which the secretory product was released. Secretion was assessed quantitatively from the incorporation of 14C-glucosamine into the acinar cells and its subsequent release into the culture medium as labelled glycoprotein. IPR stimulated secretion to 125% of untreated controls in the concentration range t x 10(-5) to 5x 10(-7) M and to 110% of controls at 5 x 10(-8) M, after 40 min incubation. Carbachol stimulated secretion to 131% of controls at 5 x 10(-5) M and to 115% at 5 x 10(-6) M but had no effect at 5 x 10(-7) or 5 x 10(-8) M. The secretory response was blocked by the respective beta-adrenergic and cholinergic antagonists, propranolol and atropine. These findings show that dissociated rat submandibular acinar cells provide a useful in vitro model for the study of mucus synthesis and secretion.
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PMID:The secretory response in dissociated acini from the rat submandibular gland. 737 55

Equine oxyntic mucosal cells were obtained by sequential exposure to pronase and collagenase. Acid production by parietal cells was assessed by uptake of [14C]aminopyrine (AP), a weak base that accumulates in intracellular acidic spaces. Incubation for various times revealed a maximal AP uptake at 10 minutes for histamine and carbachol. Similar secretagogue responses were observed for parietal cells from the mucosal cell preparation or after enrichment by elutriation. Histamine and isobutyl-methylxanthine (IBMX) stimulated AP uptake with a dose-dependent response and maximal effective concentration of 100 microM. Carbachol, 1 to 100 microM, and pentagastrin (PG), 1 to 1,000 nM, were ineffective stimulants of AP uptake. The AP uptake values for 100 microM IBMX, 1 microM carbachol, or 100 nM PG were 77 +/- 6%, 50 +/- 3.2%, and 40 +/- 4.5%, respectively, of that observed with maximal stimulation by 100 microM histamine (mean +/- SEM, n = 4 to 14). Uptake of AP by nonstimulated control cells was 36 +/- 3.6% of maximal histamine stimulation. The AP accumulations during control conditions and after stimulation with 100 microM histamine and IBMX, 1 microM carbachol, or 100 nM PG were 1.18 +/- 0.39, 2.81 +/- 0.85, 1.93 +/- 0.48, 1.44 +/- 0.36, and 1.23 +/- 0.33 pmol of AP/10(5) parietal cells, respectively. Individual histamine dose-response curves were shifted to the right by increasing ranitidine and cimetidine concentrations (0.1 to 50 microM). These results indicate that isolated equine parietal cells are maximally stimulated by histamine and minimally stimulated by carbachol and PG.
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PMID:Secretagogue-induced [14C]aminopyrine uptake in isolated equine parietal cells. 751 58


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