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)

The effects of different neurotransmitters were tested in vitro on a hypothalamic tissue, collagenase-digested isolated anterior pituitary and Leydig cell suspension system by measuring the testosterone production of the Leydig cells. Neurotransmitters were used in concentrations of 0.25, 1.0, 2.5, 5.0, and 10.0 micrograms/ml incubation medium. Dopamine in doses of 1.0, 2.5, and 5.0 micrograms/ml increased the hypothalamic tissue-induced pituitary-testis activation, while it had no direct effect on pituitary and Leydig cells. Noradrenaline in the concentration range 2.5--10.0 micrograms/ml decreased the luteinizing-hormone-releasing-hormone (LHRH) sensitivity of the pituitary cells. 5.0 and 10.0 micrograms/ml 5-hydroxytryptamine decreased the testosterone production and the hCG sensitivity of the isolated Leydig cells. Carbamylcholine and pilocarpine had no action on the in vitro system at the different levels studied.
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PMID:Study of the effects of neurotransmitters on the hypothalamus-pituitary-testis function in in vitro cell suspension system. 4 66

Single bovine adrenal medullary cells have been obtained by retrograde perfusion of adrenal medullae with a solution of 0.05% collagenase in Ca++-free Krebs Henseleit buffer. Chromaffin cells were obtained in high yield (5 X 10(6) cells/g medulla), and more than 95% of these were viable as shown by exclusion of trypan blue. The isolated cells were capable of respiring at a linear rate for a minimum of 120 min. Ultrastructural examination revealed that the cells were morphologically intact, and two distinct types of adrenal medullary cells were identified, on the basis of the morphology of their electron-dense vesicles, as (a) adrenaline-containing and (b) noradrenaline-containing cells. Biochemical analysis showed that the cells contained catecholamines and dopamine-beta-hydroxylase (DBH). The cells released catecholamines and DBH in response to acetylcholine (ACh), and this release was accompanied by changes in the vesicular and surface membranes observed at the ultrastructural level. The time-course of ACh-stimulated catecholamine and DBH release, and the dependence of this release on the concentration of ACh and extracellular Ca++ have been investigated. The isolated cells were pharmacologically sensitive to the action of the cholinergic blocking agents, atropine and hexamethonium.
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PMID:Functional and morphological characterization of isolated bovine adrenal medullary cells. 61 89

To determine whether specific hormonal responses were involved in the production of cryoprotectant (glucose) by liver of the freeze tolerant wood frog, Rana sylvatica, metabolically active hepatocytes were isolated in reasonable yields (mean 20.1 +/- 1.30% SEM, n = 29) by in situ liver perfusion with collagenase. Freshly isolated cells from autumn-collected frogs contained large amounts of glycogen (650 mumol glucosyl units/g packed cells) and produced glucose from this endogenous reserve at a rate of 10 mumol g-1 hr-1 at 0 degrees. Glucose output from cells was highly responsive to the addition of hormones; rates of glucose release increased 2.1-, 1.7-, and 1.7-fold with the addition of 10(-7) M bovine glucagon, 10(-7) M epinephrine, and 5 x 10(-6) M dibutyryl-cyclic AMP, respectively. Norepinephrine, 5-hydroxytryptamine, and bovine insulin were without effect at 0.1 microM/l. Hormone stimulation of glucose release was correlated with an increase in both the total activity and the percentage a of glycogen phosphorylase in hepatocytes. However, none of the hormones tested affected the kinetic properties of hepatocyte pyruvate kinase, suggesting the absence of covalent modification control of the enzyme. The data indicate that the freezing-stimulated production of large quantities of glucose as a cryoprotectant by R. sylvatica liver does not involve qualitative differences in the hormonal control of liver glycogenolysis, compared with other lower vertebrates. However, quantitative differences were seen, such as the much greater phosphorylase activity, 4.38 +/- 0.33 mumol min-1 g-1 packed cells, in freshly isolated R. sylvatica hepatocytes compared with 0.36 +/- 0.06 mumol min-1 g-1 in Rana pipiens hepatocytes.
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PMID:Hormonal effects on glycogen metabolism in isolated hepatocytes of a freeze-tolerant frog. 162 97

The ability of alpha 1a- and alpha 1b-adrenergic receptor subtypes to stimulate [3H]inositol phosphate [( 3H]InsP) formation was examined in collagenase-dispersed hepatocytes and renal cells. alpha 1-Adrenergic receptor binding sites were labeled with 125I-BE 2254, and the proportion of alpha 1a and alpha 1b subtypes was determined with chloroethylclonidine (CEC) and WB 4101. Hepatocytes contained only alpha 1b-adrenergic receptors, whereas renal cells had approximately equal proportions of both subtypes. Pretreatment of renal cells with CEC selectively inactivated the alpha 1b subtype, leaving a homogeneous population of alpha 1a receptors. Norepinephrine stimulated [3H]InsP accumulation to a similar extent in both hepatocytes and renal cells. Pretreatment with CEC inactivated this response completely in hepatocytes but only partially in renal cells. WB 4101 was 1000-fold more potent in inhibiting the [3H]InsP response in renal cells than hepatocytes; however, some of this difference was due to rapid metabolism of WB 4101 by hepatocytes. After correction for metabolism, WB 4101 was still 11-fold more potent in inhibiting norepinephrine-stimulated [3H]InsP formation in hepatocytes (alpha 1b) than in CEC-pretreated renal cells (alpha 1a). These results demonstrate that both alpha 1a- and alpha 1b-adrenergic receptor subtypes activate formation of [3H]InsP, although the molecular mechanisms by which these responses occur remain to be determined.
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PMID:Alpha 1-adrenergic receptor subtypes and formation of inositol phosphates in dispersed hepatocytes and renal cells. 216 16

Using hepatocytes isolated by collagenase perfusion, we studied the accumulation of 3H-noradrenaline. Cells incubated during 15 min in the presence of 0.4 mumol/l 3H-noradrenaline (without inhibition of noradrenaline metabolism) accumulated 8.32 +/- 1.77 pmol/10(6) cells (n = 3). The accumulation of 3H-noradrenaline in isolated parenchymal liver cells was sensitive to 10 mumol/l cocaine (inhibition 36.6 +/- 7.9%, n = 3) and 1 mumol/l desipramine (inhibition 27.2 +/- 6.9, n = 3). Accumulation of 3H-noradrenaline was temperature and sodium dependent (inhibition 33.2 +/- 9.4%, n = 9, when Na+ was replaced by Tris+) and was influenced by the inhibition of the membrane Na(+)-K(+)-adenosine triphosphatase (Na(+)-K(+)-ATPase) by 150 mumol/l ouabain (34.7 +/- 6.9% inhibition, n = 3). Accumulation of 3H-noradrenaline in the hepatocytes was not affected by the presence of uptake2 inhibitors, normetanephrine (30 mumol/l) and corticosterone (30 mumol/l), but was reduced by 30 mumol/l isoprenaline (76.3 +/- 5.0% inhibition, n = 6). Thus, the system that takes up and accumulates noradrenaline in the isolated rat liver cells possesses some characteristics of both, uptake1 and uptake2 systems and appears to be different from other extraneuronal cocaine-sensitive systems, such as the one reported for pulmonary endothelial cells.
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PMID:Accumulation of 3H-(+/-)-noradrenaline by isolated rat liver cells. 216 96

Separation of viable adrenaline-containing from noradrenaline-containing chromaffin cells in large amounts has been achieved. The procedure involves collagenase digestion of bovine adrenomedullary tissue, isolation of cells through gentle filtration, separation of chromaffin from nonchromaffin cells on discontinuous gradients of the radiopaque contrast Renografin, and separation of adrenaline-enriched from noradrenaline-enriched fractions after centrifugation on self-generated Percoll gradients. Collection of 1-ml Percoll fractions gave two clear-cut catecholamine peaks. The denser peak was enriched in adrenaline and phenylethanolamine-N-methyltransferase (PNMT), suggesting that over 90% of cells were adrenergic. The lighter peak was preferentially enriched in noradrenaline but not in PNMT. With this information, we could collect by gentle aspiration two main fraction layers of larger volumes; one at the bottom of the Percoll gradient, which contained essentially adrenaline-storing cells and the other at the top of the gradient, enriched in noradrenaline cells. Those cells could be maintained viable for at least 1 week in primary monolayer cultures, as shown by neutral red staining and trypan blue exclusion. This method will allow the identification of chemical components, receptors, or ionic channels present in one specific type of cell, to determine their relevance to the regulation of the differential secretion of specific materials present in one but not in the other cell type and to ascertain whether the released materials from one cell type affect the functions of the other.
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PMID:Separation and culture of living adrenaline- and noradrenaline-containing cells from bovine adrenal medullae. 233 81

Individual cells were isolated from the sino-atrial node area of the rabbit heart using an enzyme medium containing collagenase and elastase. After enzymatic treatment the cells were placed in normal Tyrode solution, where beating resumed in a fraction of them. Isolated cells were studied in the whole cell configuration. Action potentials as well as membrane currents under voltage-clamp conditions were similar to those in multicellular preparations. Pulses to voltages more negative than about -50 mV caused activation of the hyperpolarizing-activated current, if. Investigation of the properties of this current was carried out under conditions that limited the influence of other current systems during voltage clamp. The if current activation range usually extended approximately from -50 to -100 mV, but varied from cell to cell. In several cases, pulsing to the region of -40 mV elicited a sizeable if. Both current activation and deactivation during voltage steps had S-shaped time courses. A high variability was however observed in the sigmoidal behaviour of if kinetics. Plots of the fully-activated current-voltage (I-V) relation in different extracellular Na and K concentrations showed that both ions carry the current if. While changes in the external Na concentration caused the current I-V relation to undergo simple shifts along the voltage axis, changes in extracellular K concentration were also associated with changes in its slope. Again, a large variability was observed in the increase of I-V slope on raising the external K concentration. The current if was strongly depressed by Cs, and the block induced by 5 mM-Cs was markedly voltage dependent. Adrenaline (1-5 microM) and noradrenaline (1 microM) increased the current if around the half-activation voltage range and accelerated its activation at more negative voltages. Often, however, drug application failed to elicit any modification of if. Current run-down was observed in nearly all cells, although at a highly variable rate. It was accelerated by raising the extracellular K concentration but did not show a marked use dependence. Both the if activation curve and the fully activated I-V relation were affected by run-down, the former being shifted to more negative values along the voltage axis and the latter being depressed with no apparent change of the if reversal potential.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:Properties of the hyperpolarizing-activated current (if) in cells isolated from the rabbit sino-atrial node. 243 47

To study the properties of vascular smooth muscle in hypertension without the influence of the nerves and endothelium, a procedure was developed to isolate single smooth muscle cells from tail arteries of spontaneously hypertensive rats (SHR) and Wistar-Kyoto (WKY) normotensive control rats. Perfusion of intact arteries with a solution of papain and collagenase produced dense populations of viable cells (more than 10(4) cells/ml) that remained relaxed in the presence of physiological levels of calcium. Contractile responses of smooth muscle cells from the SHR were significantly more sensitive to noradrenaline, potassium depolarization, and the calcium channel agonist Bay K 8644 compared with those from WKY rats. Enhanced sensitivity to calcium in the SHR was also observed on readdition of calcium to cells preincubated in noradrenaline or KCl in a calcium-free medium. These results provide evidence for alterations in the properties of vascular smooth muscle in the SHR at the single cell level.
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PMID:Isolation and characterization of single vascular smooth muscle cells from spontaneously hypertensive rats. 247 94

The purpose of this study was to investigate the possible role of leucocytes in the pathogenesis of reperfusion-induced vasospasm of ischaemic mesenteric arteries. Scanning electron microscopy of the rat superior mesenteric artery (SMA) after 30 min of ischaemia in vivo revealed adherence of leucocytes to the vessel wall. Isolated SMA preparations were perfused with Krebs-Henseleit buffer containing noradrenaline. Infusion of homologous leucocytes resuspended in perfusate (3 x 10(6) cells/ml) into these preconstricted preparations caused a fall in resistance of 29 +/- 2%. Removal of the endothelium by collagenase treatment abolished this response. Indeed, leucocyte infusion caused an increase in resistance of 39 +/- 8% under these circumstances. Following 30 min of normothermic ischaemia, leucocyte infusion caused a transient vasodilatation of 31 +/- 4% followed by an increase of 38 +/- 11% in the perfusion resistance of isolated SMA preparations. In each case, a similar response was obtained to infusion of the cell-free supernatant. These results suggest that leucocyte activation occurs in vivo during reperfusion of the SMA after as little as 30 min of ischaemia, and that activated leucocytes can release humoral vasoactive factors which evoke an endothelium-dependent vasodilator response in normal vessels but a predominantly vasoconstrictor response following brief intervals of ischaemia.
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PMID:Leucocyte-induced endothelium-dependent vasodilatation and post-ischaemic vasospasm in the isolated rat superior mesenteric artery. 281 30

The effects of collagenase on the membrane response to catecholamines were studied with intracellular microelectrodes in the guinea-pig taenia caeci and main pulmonary artery. In the taenia, the inhibitory actions of adrenaline mediated through alpha- and beta-adrenoceptors were both nearly abolished by two 5-min treatments with 0.005% collagenase. In the pulmonary artery, the depolarizing action of noradrenaline mediated through beta-adrenoceptors was markedly reduced by three 5-min treatments with 0.01% collagenase.
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PMID:Collagenase eliminates the electrical responses of smooth muscle to catecholamines. 282 42


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