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)

Dispersed acini isolated by collagenase digestion of the rat submandibular gland were used to compare the effects of amiloride and furosemide on the uptake of the isotopic tracer 22Na and on the binding of [3H]quinuclidinyl benzylate ([3H]QNB). In mM concentrations, both inhibitors reduced 22Na uptake in resting cells 34 and 25-29%, respectively. Acetylcholine (1 microM) enhanced uptake 23% and this effect was reduced 45% by amiloride and 26% by furosemide. Amiloride inhibited the binding of [3H]QNB to crude membranes prepared from fresh submandibular glands in a dose-dependent fashion (IC50 = 8 x 10(-6) M). Furosemide (3 x 10(-8) to 10(-3) M) did not inhibit radioligand binding. Na influx into resting salivary acini thus appears to occur by both amiloride-sensitive and furosemide-sensitive transport systems. The similar inhibition by furosemide of unstimulated and stimulated uptake of 22Na suggests that acetylcholine does not significantly activate the cotransport system within the time frame (i.e., 2 min) of the experiments. Acetylcholine appears to activate an amiloride-sensitive Na/H antiport, but amiloride blocks cholinergic receptors and may thus affect Na transport by receptor blockade. Other actions of amiloride, such as its ability to penetrate into cells and to act as a weak base which alters intracellular pH, may also contribute to the inhibition of Na entry into salivary cells.
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PMID:Amiloride inhibits 22Na uptake and [3H]QNB binding in rat submandibular cells. 275 81

Acetylcholine receptors at the vertebrate neuromuscular junction are highly organized and metabolically very stable. We report here that digestion of adult rat skeletal muscle with collagenase disorganizes junctional receptors and increases their turnover rate.
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PMID:Collagenase digestion alters the organization and turnover of junctional acetylcholine receptors. 301 22

Vascular relaxation in rabbit aortic preparations induced by acetylcholine is endothelium-dependent. The nature of the endothelium-derived relaxing factor (EDRF) has not been ascertained because it is very labile (reported half-life 6-50 seconds). To obtain a stable source of EDRF, a system was developed in which the relaxing factor was continuously produced by freshly harvested porcine endothelial cells. Endothelial cells were collected from aortas by exposing the endothelial lining to collagenase 0.1%. Cells were washed and concentrated by repeated centrifugation to obtain a high cell count (7.2 X 10(6) cells/ml). Endothelium-deprived aortic strips from rabbits were incubated in these cells suspended in tissue culture medium and fetal calf serum. The strips were precontracted with histamine. Acetylcholine was added to induce EDRF release. Significant relaxation of endothelium-deprived aortic strips was observed. Superoxide dismutase, an enzyme known to protect EDRF against inactivation, caused further relaxation, which was inhibited by the addition of hemoglobin, an agent known to inhibit the relaxing action of EDRF. Even without the addition of acetylcholine, hemoglobin caused contraction of the denuded aortic strips in suspension of porcine endothelial cells, demonstrating spontaneous EDRF release. Hemoglobin had no effect in cell-free medium. Endothelial-cell-dependent relaxation occurred without attachment of endothelial cells to the endothelium-deprived aortic strips: when the cell suspension was replaced by cell-free medium, relaxation did not occur after acetylcholine. Scanning electron microscopy showed no attachment of endothelial cells to the subendothelial layer. It can be concluded that freshly harvested endothelial cells produce endothelium-derived relaxing factor with an without stimulation by acetylcholine.
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PMID:Release of endothelium-derived relaxing factor from freshly harvested porcine endothelial cells. 349 57

Choline acetyltransferase (ChAT) activity was measured in a fraction of endothelial cells obtained by collagenase digestion from previously isolated rat brain cortex capillaries. ChAT specific activity in the dissociated cells (0.264 nmol X mg protein-1 X min-1) was nearly 7 times higher than the activity measured in 'capillary-forming' cells (0.038 nmol X mg protein-1 X min-1). It is not known whether this increase was caused by ChAT enrichment of the former cellular fraction, and/or by a change in the specific activity of the enzyme during the isolation procedure, but the finding would be in favor of the intrinsic localization of cerebrovascular ChAT in endothelial cells. This would support the hypothesis of a role for locally synthesized ACh in the vasodilating response to noxious stimuli.
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PMID:Synthesis of acetylcholine by endothelial cells isolated from rat brain cortex capillaries. 360 47

Aggregates of collagenase-dissociated neonatal rat heart cells have been tested for several membrane properties and shown to be comparable with cells from the intact heart. Action potentials, recorded from driven aggregates, are fully suppressed by tetrodotoxin (TTX). Under Mn2+, the plateau phase of the action potential disappears and no more mechanical activity can be detected. In aggregates, therefore, apparently both the fast sodium inward current and a slow inward current, which is at least partly carried by Ca2+ ions, contribute to the action potential. Pacemaker activity in spontaneously active aggregates is enhanced by adrenaline and slowed down by acetylcholine. Adrenaline also increases the plateau phase amplitude of the action potential and thereby the rate of repolarization. Acetylcholine shortens the action potential duration and increases the resting membrane potential. The electrical coupling between the cells in the aggregates is so tight that the aggregate seems to behave passively, like a single cell. It is concluded that aggregates of collagenase-dissociated neonatal rat heart cells may be used to study active electrical properties using the voltage clamp technique.
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PMID:Membrane properties of aggregate of collagenase-dissociated rat heart cells. 624 36

1. The clusters composed of 10-20 cells of the rabbit sinoatrial node were prepared by coronary perfusion of collagenase and their response to ionophoretic application of ACh was recorded. The hyperpolarizing response showed a sigmoidal onset without any clear latency when the ACh pipette was positioned close to the cell surface. 2. Equations describing the kinetics and amplitude of the ACh-induced K current were determined based on the reported experimental results obtained by relaxation and noise analysis. 3. The kinetics of the ACh-induced K current simulated well the time course of the ACh response and also the effect on the spontaneous action potential when data on the kinetics was incorporated into the S-A node pacemaker mathematical model.
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PMID:Ionic mechanism underlying muscarinic acetylcholine response in the rabbit sinoatrial node. 628 57

1. Neurotransmitter-receptors in the membrane of Xenopus oocytes have been studied using electrophysiological techniques. Neurotransmitters and related agents were applied while recording either membrane potential or membrane current. The majority of ovarian oocytes used were at stages IV and V.2. Three types of oocytes were examined: inner ovarian epithelium covered (e.c.) oocytes; epithelium manually removed (e.r.) oocytes; and collagenase treated (c.t.) ooctyes.3. Ovarian oocytes are sensitive to some cholinergic and catecholaminergic agents. Responses to serotonin were seldom observed and when present were much weaker than responses to other agents. No responses were observed to the amino acids: aspartate, glutamate, gamma-aminobutyric acid, and glycine; or to octopamine and histamine.4. Acetylcholine (ACh) usually depolarized the membrane, in a dose-dependent manner, with threshold concentrations as low as 10(-9)m. The ACh-potential was due to an increase in Cl permeability and had a reversal potential around - 19 mV. The intracellular Cl ion activity, measured with a Cl-ion sensitive micro-electrode, was about 65 mm and the estimated Cl-ion equilibrium potential, E(Cl), agreed with the reversal potential of the ACh-potential.5. Curare (10(-4)m), tetrodotoxin (10(-6)m), or alpha-bungarotoxin (10(-6) g/ml.) did not block the response to 10(-6)m-ACh; whereas atropine (10(-7)m) blocked it. No response to nicotinic agents (e.g. nicotine, 1,1-dimethyl-4-phenylpiperazinium) was observed. These results suggest that the ACh receptors in the oocyte membrane are muscarinic in nature.6. The apparent latency of the ACh potential, examined by ionophoretic application of ACh to e.r. oocytes and c.t. oocytes, ranged from 0.5 sec to over 20 sec. Intracellular injection of ACh was without effect.7. Responses to catecholamines were observed mostly in e.c. oocytes; while in e.r. and c.t. oocytes they were rare and of very small amplitudes.8. The usual response to both dopamine and (-)-epinephrine was a transient hyperpolarization manifested by an initial increase in K-permeability followed by a decrease. The latency of these responses ranged from 10 sec to over 30 sec and their reversal potential was nearly - 100 mV, which coincided with E(K).9. Oocytes responded to the beta-adrenergic receptor agonist, isoproterenol, as well as (-)-epinephrine. Pre-treatment with the beta-adrenergic receptor blocker, propranolol, abolished the response to both (-)-epinephrine and (-)-isoproterenol. The dopamine potential was also reduced considerably. Both the alpha-adrenergic receptor agonist, phenylephrine, and the alpha-adrenergic receptor blocker, phentolamine, were without effect.10. Maturation of the oocytes, induced in vivo by gonadotropin or in vitro by progesterone, led to loss of responsiveness to both cholinergic and catecholaminergic agents.
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PMID:Cholinergic and catecholaminergic receptors in the Xenopus oocyte membrane. 713 11

Acetylcholine, acetylthiocholine, carbachol, suberyldicholine, propionylcholine, succinylcholine, methylfurmethide and F 2268 were tested on motor nerve ending currents recorded with an extracellular microelectrode. The isolated and transversally cut cutaneous pectoris muscle of frog Rana ridibunda was used. Only acetylcholine and acetylthiocholine affected the spike waveforms in a concentration-dependent manner. Lower concentrations (1-6 x 10(-4) M) prolonged the inward Na+ current and increased the outward K+ current at the proximal and central parts of the nerve terminal. Most remote parts of the terminal were not affected. At 7 x 10(-4) M and higher, both drugs further prolonged the Na+ current and eliminated the K+ component of the spike. The potentiating effect of acetylcholine and acetylthiocholine on the K+ phase of nerve terminal current disappeared after treatment with tetraethylammonium and 4-aminopyridine. The effect also disappeared when synaptic cholinesterase was inhibited by the anticholinesterases or by treatment with collagenase. Reactivation of cholinesterase by dipyroxime restored the facilitating effect of acetylcholine. Choline and slight acidification to pH 6.8 did not mimic the acetylcholine action on the terminal currents. Facilitation of the K+ current by acetylcholine was not calcium-dependent. The results indicate that lower acetylcholine concentrations inhibit the delayed rectifier only, whereas 7 x 10(-4) M and higher concentrations of acetylcholine depress all outward currents of the terminal.
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PMID:The effect of acetylcholine and related drugs on currents at the frog motor nerve terminal. 782 42

It is shown that ADP and DNP does not intensify the respiration rate in hepatocytes of rats obtained by means of trypsin or EDTA. The same cells obtained using collagenase, phosphorylate added ADP and increase the respiration rate after DNP addition. Acetylcholine added to the cell suspension in a dose of 5 x 4 x 10(-8) M) increases the efficiency of oxidative phosphorylation. The scheme of neurotransmitter regulation of intensity of respiration and efficiency of oxidative phosphorylation on the cell level is suggested.
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PMID:[Acetylcholine and effectiveness of oxidative phosphorylation in isolated rat hepatocytes]. 816 Mar 6

In order to choose the best procedure to inactive the endothelium from vascular beds perfused in vitro, we compared four methods: perfusion with sodium deoxycholate 0.3% for 30 sec; 3-[(3-cholamidopropyl)dimethylammonio]-1-propane sulfonate 0.3% (CHAPS) for 2.5 min; collagenase 0.2% for 15 min, and distilled water for 10 min, using the mesenteric arterial bed (MAB) of the rat. The effectiveness of the treatments used to inactivate the endothelium was assessed functionally by using acetylcholine and sodium nitroprusside and histologically using light microscopy. Phenylephrine was used to test the contractile properties of the preparations after each treatment. After collagenase, distilled water, and CHAPS treatment, a potentiated response to phenylephrine was observed, whereas sodium deoxycholate treatment did not modify phenylephrine-induced responses. Acetylcholine-induced responses were reduced by collagenase (60% reduction), CHAPS (30% reduction), and distilled water (52% reduction) treatment, and sodium deoxycholate completely abolished acetylcholine-induced responses. Except after collagenase treatment, smooth muscle relaxant responses were not altered. Medial smooth muscle cells displayed an unchanged morphology, appearing similar to those in control mesenteric arterial beds, except for collagenase and distilled water. Despite the fact that sodium deoxycholate treatment completely abolished acetylcholine-induced response, endothelial cells were still found. No treatment totally removed endothelial cells. In conclusion, we suggest that sodium deoxycholate treatment is the best procedure to inactivate endothelial cells from vascular beds perfused in vitro since it completely abolished endothelium-dependent relaxation and did not interfere with smooth muscle vasodilating and contracting properties.
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PMID:Endothelium inactivation in in vitro perfused vascular beds. Comparison of methods. 836 29


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