Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: UNIPROT:P06889 (Mol)
 630,302 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Calcium exchange was measured in enzymatically isolated, cultured smooth muscle cells from the rabbit thoracic aorta. Cells were grown to confluence in monolayer on scintillator discs which were then inserted into a modified flow cell/spectrometer to measure 45Ca exchange in a continuous, on-line manner. This also allowed us to measure rapid movements of cellular Ca2+ without solubilizing the cells. Two components of Ca2+ exchange were detected: a La3+-displaceable, rapidly exchangeable fraction (t 1/2 = 15.6 s) and a slowly exchangeable fraction (t 1/2 = 34.5 min). The La3+ displaceable, rapidly exchangeable Ca2+ fraction represented 57 to 61% of the total exchangeable Ca2+. Multiple passaging of cells did not after the Ca2+ flux characteristics. Low Na+ perfusion increased smooth muscle cell Ca2+ flux by 6.71 mmol/kg dry weight. This increase was localized to both the rapidly and slowly exchangeable Ca2+ compartments. Ouabain, an inhibitor of the plasmalemmal Na+ -K+ pump, also increased net uptake of 45Ca. The results demonstrate that approximately 60% of exchangeable Ca2+ of smooth muscle cells is very rapidly exchangeable (t 1/2 less than 16 s). The results stress the importance of measuring 45Ca2+ movements in an on-line, continuous manner in order to ensure detection of the majority of total exchangeable Ca2+ in smooth muscle cells.
J Mol Cell Cardiol 1989 Sep
PMID:Calcium is rapidly exchangeable in cultured vascular smooth muscle cells from rabbit aorta. 281 Mar 78

Ouabain inhibits (IC50 congruent to 200 nM) the congruent to 100-fold adrenergic cyclic AMP stimulation of rat pineal arylalkylamine N-acetyltransferase (EC 2.3.1.87, serotonin N-acetyltransferase, NAT) activity in intact pineal glands. In the present study, ouabain binding sites in pineal membranes were characterized in detail and compared to sites in isolated pinealocytes, which mediate the inhibition of Na+,K+-ATPase, as indicated by 86Rb uptake and norepinephrine (NE) stimulation of NAT activity. High affinity ouabain-binding sites were identified in crude preparations of pineal membranes (Kd congruent to 14 nM; Bmax congruent to 4 pmol/mg of protein) and similar sites were also found in ovine and bovine pineal tissue. The ouabain Kd value for the rat pineal binding sites was similar to the estimated ouabain IC50 values for 86Rb uptake and the NE stimulation of NAT activity in intact rat pinealocytes. In addition, the relative orders of potency of four cardiac glycosides in displacing [3H]ouabain from high affinity binding sites and inhibiting both 86Rb uptake and NE stimulation of NAT activity were the same (acetyldigitoxin greater than ouabain greater than digitoxin greater than strophanthidin). The similarities in the characteristics of the high affinity [3H]ouabain-binding sites and the sites involved in the inhibition of 86Rb uptake and stimulation of NAT activity indicate that an alpha +-like Na+,K+-ATPase mediates the inhibitory effects of ouabain on the adrenergic induction of pineal NAT activity.
Mol Pharmacol 1987 Dec
PMID:Characterization of the alpha +-like Na+,K+-ATPase which mediates ouabain inhibition of adrenergic induction of N-acetyltransferase (EC 2.3.1.87) activity: studies with isolated pinealocytes. 282 93

1. We measured changes in resting membrane potential (Em) and Na-K pump activity, assayed by ouabain-sensitive 86Rb uptake, in response to carbamylcholine (CCh) and its continued presence in single rat skeletal myotubes in culture. 2. CCh caused immediate depolarization from control Em (-80 to -85 mV) to near 0 followed by repolarization of varying degrees depending on the age of the culture and temperature of the recording medium; repolarization of Em was most apparent by culture age 8-9 days in vitro (DIV), Em reaching values as high as -60 mV by 5-10 min after peak depolarization at 37 degrees C. 3. Input resistance, which decreased during CCh depolarization, increased only slightly during the initial phase of repolarization and then remained essentially unchanged during the major component of membrane repolarization in the presence of CCh. 4. Ouabain, given before CCh, prevented repolarization of Em and, when given after repolarization had begun, reversed it and caused Em to return to about -7 mV. 5. Na-K pump activity was decreased in myotubes in which Em did not repolarize or did so only slightly, and was increased by over 40-50% in myotubes whose Em repolarized by 40-60 mV, even though CCh was still present in the medium. Inhibition of pump activity in non repolarizing myotubes was related to Na influx, inhibition being reversed to stimulation when CCh was administered to myotubes in Na-free medium. 6. Repeated (three or four times) or prolonged (up to 60-min) administration of CCh to myotubes in which repolarization was hardly expressed (age 6-7 DIV) caused increases both in the amount of repolarization and in 86Rb uptake, both being related to the number or duration of CCh exposures. 7. We conclude that repolarization of Em following CCh-induced depolarization of cultured rat skeletal myotubes depends to a large extent on an increase in activity of the electrogenic Na-K pump.
Cell Mol Neurobiol 1988 Dec
PMID:Effects of carbamylcholine on membrane potential and Na-K pump activity of cultured rat skeletal myotubes. 285 60

Calcium (Ca2+) exchanges were studied in dog thyroid slices incubated in vitro. With 45Ca2+-prelabeled slices, carbamylcholine 10(-7)-10(-5) M (Cchol) induced an important transitory spike efflux, inhibited by procaine and atropine while the stimulated efflux obtained with high concentrations of TSH (10 mU/ml) was progressive and sustained over time. The effects observed with both agents did not require extracellular Ca2+ and were insensitive to verapamil 10(-6)-10(-4) M. Neither dibutyryl (Bu2)-cAMP, nor any agent raising intracellular cAMP (prostaglandin E2, choleratoxin, inhibitors of phosphodiesterases with low concentrations of TSH) were able to reproduce the action of TSH 10 mU/ml, forskolin 10(-5) M being the only exception. Replacement of sodium by choline (+ atropine) in the incubation medium decreased the basal efflux and inhibited the TSH effect. Ouabain 10(-3) M also abolished the TSH-induced Ca2+ efflux, while having no influence on carbamylcholine action. TSH 10 mU/ml and 1 mU/ml, Bu2-cAMP 10(-3) M, choleratoxin and prostaglandin E2 with inhibitors of phosphodiesterase decreased the total 45Ca2+ uptake of the slices, while no effect of Cchol could be detected on this parameter. The results obtained suggest that (1) Cchol and TSH stimulate 45Ca2+ efflux from dog thyroid slices with different kinetics, by mobilization of intracellular Ca2+ stores; (2) this effect of TSH is not mediated by cAMP; (3) independently TSH at low concentrations (1 mU/ml), through cAMP, decreased 45Ca2+ uptake; this suggests that increased 45Ca2+ efflux and decreased uptake result from different mechanisms, as has been described for iodide exchange in FRTL-5 cells.
Mol Cell Endocrinol 1987 May
PMID:Regulation of calcium fluxes in the thyroid. 303 26

Myocardial ischaemia induces cytosolic acidification, which promotes cardiac damage, dysfunction or arrhythmia. In this study, we investigated the effect of ouabain on the intracellular pH (pHi) in cultured mouse ventricular cells, using 2',7'-bis(carboxyethyl)-5,6-carboxyfluorescein (BCECF). The average resting pHi in myocytes was 7.19. After myocytes were acid-loaded with NH4Cl, the pHi recovered from acidosis to the resting level within a few minutes via amiloride-sensitive Na+/H+ exchange. Ouabain inhibited this pHi recovery dose-dependently with half-maximal inhibition at 3 X 10(-5) M, but did not suppress the ionophore monensin-induced pHi elevation. The inhibition of the pHi recovery from acidosis by ouabain is possibly caused by an inhibition of amiloride-sensitive Na+/H+ exchange, which is secondary to a suppression of Na+ efflux through (Na+, K+) pump. Above results demonstrate the possibility that digitalis promotes intracellular acidosis or inhibits the pHi recovery from acidosis in ischaemic myocardium.
J Mol Cell Cardiol 1988 Jan
PMID:Ouabain inhibits intracellular pH recovery from acidosis in cultured mouse heart cells. 336 75

Free parasites were isolated from Plasmodium chabaudi-infected rat erythrocytes by N2-cavitation and purified on Percoll gradients. The membrane potential of the free parasites determined from the transmembrane distribution of the lipophilic cation, tetraphenylphosphonium, was -93 +/- 10 mV for late stage parasites and -90 +/- 3 mV for ring forms. Studies with intact infected erythrocytes demonstrated that the membrane potential of ring forms was much smaller compared to late trophozoites and schizonts and thus the present findings with free parasites suggest that host cell cytoplasmic factors may determine the magnitude of the parasite membrane potential. Both extracellular pH and [Na+] were found to modify the membrane potential of free parasites. Electrogenic protonophores, the H+-ATPase inhibitor dicyclohexylcarbodiimide and orthovanadate collapsed the potential of free parasites. Ouabain (or its membrane permeant derivative, strophanthidin), and oligomycin were without effect. These inhibitor studies suggest that an electrogenic H+-ATPase similar to that found in yeast generates in part the membrane potential of malaria parasites. Using weak acid distribution or a pH sensitive fluorescent dye, it was demonstrated that free parasites maintain an alkaline intracellular pH at extracellular pH greater than 6.5. The pH gradient was partially collapsed by orthovanadate or dicyclohexylcarbodiimide and by substitution of Na+ for K+ in the suspending buffer. The H+-ATPase and K+:H+ exchange may therefore both contribute to regulation of intracellular pH in Plasmodium.
Mol Biochem Parasitol 1986 Oct
PMID:Membrane potential of erythrocytic stages of Plasmodium chabaudi free of the host cell membrane. 377 36

A new method is described for making direct measurements of compartmentalized subcellular stores of calcium in the isolated perfused dog heart. Cellular calcium was immobilized by freezing the myocardium in diastole with an extremely cold fluorocarbon fluid (-125 degrees C). All subsequent procedures were conducted under conditions which prevented ionic diffusion, either at temperatures well below the freezing point of water or in the absence of water. Sarcolemmal and mitochondrial enriched fractions were segregated from dessicated, homogenized, myocardial tissue by ultracentrifugation utilizing density-gradients composed of blends of silicone and halocarbon on fluids within which physiological salts are insoluble. The total calcium content of these isolated fractions were then determined by atomic absorption spectrophotometry. Ouabain and epinephrine were subsequently used to alter the contractility of the perfused hearts and such contractile alterations were then related to changes noted in the calcium activity of the isolated subcellular fractions. In this study the calcium levels of the enriched mitochondrial fractions were elevated by both ouabain and epinephrine, while the calcium levels of the enriched sarcolemmal fractions were elevated only by ouabain. The advantage of this segregative procedure is that it prevents artifactual intercompartmental calcium rearrangement and preserves calcium levels to those initially fixed in situ at the time of freezing.
J Mol Cell Cardiol 1985 Mar
PMID:Nonpolar density gradient ultracentrifugation in the direct determination of myocardial subcellular calcium. 383 24

Pseudopregnant rabbit mammary glands in organ culture were used to investigate prolactin (PRL) receptor turnover. Chloroquine (100 microM) results in an increase in prolactin receptor levels (15.7 +/- 1.2% to 35.9 +/- 3.5% specific binding), whereas cycloheximide (1 microgram/ml) induces a rapid decline (to 6.4 +/- 1.2%) suggesting a rapid synthesis and degradation of the receptor molecule. Inhibitors of cellular transcription have little effect on receptor levels. Neither actinomycin D nor dichlororibofuranosylbenzimidazole (DRB) diminish PRL receptor levels whereas total protein synthesis is almost completely inhibited, and chloroquine increases the binding even in the presence of transcriptional inhibitors. These results imply that receptor synthesis continues and that the mRNA for the receptor protein is particularly stable. Ouabain (3 micrometers), which blocks the ATP-dependent Na+/K+ pump, provokes a greater than 60% reduction in PRL receptor levels without modifying total protein synthesis. Dinitrophenol (DNP, 1 mM), an oxidative uncoupler, has little effect on receptor levels, possibly due to a blockage of both synthesis and degradation. Prolactin is capable of inducing a 60% down-regulation of its own receptor, and this phenomenon appears to be energy-dependent because it is partially inhibited by DNP. This process seems to involve an increased rate of receptor degradation. These studies suggest that, at any one time, the level of PRL receptors in a target cell is the result of a dynamic equilibrium between receptor synthesis and degradation and that the most frequent modulations occur at the level of translation and lysosomal degradation. In conclusion, in mammary glands of the pseudopregnant rabbit, the prolactin receptor molecule appears to have a short half-life; the mRNA for this protein, however, is relatively stable.
Mol Cell Endocrinol 1982 Feb
PMID:Prolactin receptor turnover in explants of pseudopregnant rabbit mammary gland. 627 49

Potassium-stimulated release of many hormones requires the presence of extracellular calcium. At variance with this mechanism, potassium-evoked parathyroid hormone (PTH) release from perifused dispersed bovine parathyroid cells also occurred in calcium-free medium containing 1 mM EGTA. Tetraethylammonium, which presumably suppresses the efflux of potassium from parathyroid cells, also stimulated PTH secretion. Removal of sodium did not suppress potassium-stimulated PTH secretion, but inhibited the release of the hormone evoked by calcium removal and by isoproterenol. Ouabain, on the other hand, suppressed the release of PTH evoked by calcium removal and by potassium, but not by isoproterenol. Unlike high potassium and removal of calcium, isoproterenol caused a parallel increase in cAMP release. In conclusion, PTH secretion is reversibly stimulated by potassium in the absence of extracellular calcium. Our findings suggest that potassium stimulates PTH secretion by a unique mechanism the nature of which remains to be elucidated.
Mol Cell Endocrinol 1983 Sep
PMID:Potassium stimulates parathyroid hormone release in the absence of extracellular calcium. 631 51

The effect of ouabain on the secretion of catecholamines from isolated bovine adrenal medullary cells was investigated. Ouabain enhances the basal rate of secretion approximately 2-fold, with half-maximal stimulation occurring at a glycoside concentration of around 5 X 10(-7) M. Parallel measurements of the release of dopamine beta-hydroxylase (EC 1.14.17.1) (an enzyme associated with chromaffin granules) and lactate dehydrogenase (EC 1.1.1.27) (which is confined to the cytosolic compartment) suggest that this increase in secretion occurs as a result of an enhanced rate of exocytosis rather than by any other route. The stimulatory effect of ouabain is dependent on extracellular sodium but is maintained in the nominal absence of calcium and is unaffected by changes in the major external anion. Neither tetrodotoxin nor phenoxybenzamine alters the response to glycoside treatment, but the calcium channel blocker methoxyverapamil reduces the catecholamine secretion evoked by ouabain in a dose-dependent fashion. This study serves to characterize the secretory action of ouabain in isolated chromaffin cells and to provide a foundation for the ion flux studies reported in the following paper.
Mol Pharmacol 1983 May
PMID:Ionic and metabolic requirements for stimulation of secretion by ouabain in bovine adrenal medullary cells. 640 91


<< Previous 1 2 3 4 5 6 7 8 9 10 Next >>