Two molecular forms of Na+,K+-ATPase in the ferret heart and developmental changes in digitalis sensitivity.
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Aging↗
Biomedical subjects
Y C Ng
Publications and source records attributed to Y C Ng.
Immune complexes and erythrocyte CR1 (complement receptor type 1): effect of CR1 numbers on binding and release reactions.
We performed experiments to investigate whether immune complexes opsonized with C3b and iC3b transferred from CR1 on one erythrocyte to CR1 on others, and studied the effect of variation in erythrocyte CR1 number on the transfer reaction. We used populations of cells of different blood groups to study this phenomenon which were separated by differential agglutination with monoclonal anti-group antibodies. The rate of transfer of immune complexes between erythrocytes was related to CR1 concentration of both donor and recipient cells; fastest transfer occurred from donor cells of low CR1 numbers to recipient cells of high CR1. These results were not explained by a difference in the binding constant of immune complexes to erythrocytes bearing different numbers of CR1. In the absence of factor I, complexes partitioned between erythrocytes according to their relative concentrations of CR1 with no release of complexes into solution. In serum, the proportion of complexes bound to donor and recipient erythrocytes was similarly related to their respective CR1 numbers with progressive release of complexes into solution. Erythrocyte CR1 may act as a dynamic buffering system which prevents immune complexes that have bound complement from fixing to vascular endothelium.
Antigen-Antibody Complex↗
Cardiodepressant actions of the orally active male contraceptive, gossypol, in guinea-pig heart muscle.
We have previously shown that gossypol has direct actions on isolated atrial muscle preparations. The possible mechanisms responsible for the transient positive and sustained negative inotropic effects of gossypol were examined under conditions that modify Ca2+ pools involved in contractile activation. In Langendorff preparations obtained from guinea-pig or rat heart, gossypol produced marked negative inotropic and arrhythmogenic effects but failed to produce a positive inotropic effect. Langendorff preparations were significantly more sensitive than atrial muscle preparations. In atrial muscle preparations, the negative inotropic effect of gossypol was not specific to utilization of superficial or intracellular Ca2+ pools; force-staircase phenomenon observed between 0.5 and 3 Hz, contractions elicited by slow action potentials in partially depolarized muscle, the inotropic effect of extracellular Ca2+ and potentiated post-rest contractions were all suppressed by gossypol to the similar extent. Low external Na+ concentrations abolished the positive inotropic effect of gossypol without affecting the negative inotropic effect. A low extracellular Ca2+ concentration enhanced the transient positive inotropic effect and delayed development of the negative inotropic effect. Simultaneous reduction of extracellular Na+ and Ca2+ concentrations abolished the positive inotropic effect and enhanced the negative inotropic effect. Gossypol inhibited ATP-dependent Ca2+ uptake by sarcolemmal vesicles obtained from dog heart. These results indicate that the actions of gossypol on cardiac muscle is not specific to utilization of either the superficial or intracellular Ca2+ pools involved in contractile activation.
Animals↗
Direct actions of gossypol on cardiac muscle.
Gossypol, an orally active male contraceptive, has been reported to produce cardiotoxicity. In the present study, direct actions of gossypol on the heart muscle were examined using atrial muscle preparations isolated from guinea-pig heart. Gossypol (20-30 microM) produced a transient positive chronotropic effect in spontaneously beating right atrial muscle preparations and a transient positive inotropic effect which reached a peak at about 5 min followed by a marked negative inotropic effect in paced left atrial muscle. These effects of gossypol were not affected by blocking concentrations of phentolamine, nadolol or propranolol, cimetidine and atropine. Gossypol inhibited Na+,K+-ATPase isolated from guinea-pig brain, Na+ pump activity in electrically stimulated left atrial muscle preparations of guinea-pig heart and Na+/Ca2+ exchange in Na+-loaded sarcolemmal vesicles obtained from dog heart. Inhibition of Na+/Ca2+ exchange required high concentrations of gossypol and was rapidly reversible whereas Na+,K+-ATPase inhibition was apparently irreversible. These results indicate that micromolar concentrations of gossypol have direct actions on cardiac muscle.
Animals↗
Paradoxical positive inotropic effect of K+ in the rat heart.
In cardiac muscle isolated from most mammalian species, an elevation of extracellular K+ concentration decreases developed tension. This is explained from stimulation of the Na pump. In rat myocardium, however, developed tension increased when K+ concentration was raised from 3.5 to 9.5 mM, seemingly inconsistent with the above explanation. Activation of isolated Na+,K+-adenosinetriphosphatase by K+ was not different in rat and guinea pig heart. The K+-induced increase in developed tension observed in left atrial muscle preparations obtained from rat heart was not blocked by phentolamine and propranolol or by reserpine pretreatment but attenuated by stimulation at higher frequency, incubation at a lower temperature, or by veratridine, all of which have been shown to increase Na+ loading of myocardial cells. K+ slightly prolonged action potential duration and partially depolarized resting membrane potential; however, K+-induced changes in electrophysiological parameters or possible inactivation of early outward current observed in myocytes isolated from rat heart are unlikely to be the primary cause of the positive inotropic effect. Caffeine or Cd2+ eliminated the inotropic effect of K+ but ryanodine was ineffective. An increase in extracellular K+ from 1 to 3.5 mM decreased developed tension. These results indicate that K+ has dual effects on developed tension in rat myocardium: a negative inotropic effect resulting from Na pump stimulation and a positive inotropic effect. The latter effect is due to a process that is either unique or more strongly expressed in the rat myocardium and masks the former effect in the range of physiological K+ concentrations.
Action Potentials↗
Two classes of ouabain binding sites in ferret heart and two forms of Na+-K+-ATPase.
In partially purified Na+-K+-adenosinetriphosphatase (ATPase) obtained from ferret heart, ouabain produced a monophasic inhibition curve; however, the curve spanned over 5 logarithmic units, indicating the presence of more than one classes of enzyme. [3H]ouabain binding studies revealed high-and low-affinity binding sites in approximately equal abundance, with apparent dissociation constants of 10 and 230 nM, respectively. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis of phosphoenzyme formed from [gamma-32P]ATP showed two distinct K+-sensitive bands of approximately 100,000 molecular weight. Phosphoenzyme formation from the high-molecular-weight alpha(+) form was selectively inhibited by N-ethylmaleimide. Ouabain caused a 50% inhibition of phosphorylation of the alpha(+) form at 40 nM and the lower-molecular-weight alpha form at 300 nM. In papillary muscle preparations, 1-30 nM ouabain produced a modest positive inotropic effect that reached an apparent plateau at 30 nM. Further increases in ouabain concentrations, however, produced additional and prominent inotropic effects at 0.1-10 microM. These results indicate for the first time in cardiac muscle that the high- and low-affinity ouabain binding sites are associated with the alpha(+) and alpha forms of the Na+-K+-ATPase, respectively, and that binding of ouabain to either of these sites causes enzyme inhibition and the positive inotropic effect.
Animals↗
Erythrocytes transfused into patients with SLE and haemolytic anaemia lose complement receptor type 1 from their cell surface.
Erythrocyte complement receptor type 1 (CR1) shows a numerical deficiency in patients with SLE and with haemolytic anaemias. This receptor is a cofactor for the enzymatic degradation of C3b and is believed to play a role in the transport of immune complexes from the circulation to the reticulo-endothelial system. Erythrocyte CR1 was enumerated on cells which had been transfused into patients with SLE and haemolytic anaemias. In three patients with active disease up to 60% of CR1 was lost from erythrocytes during 5 days after blood transfusion and up to 410 molecules of C3 were deposited on these cells. These are the first in vivo data showing that CR1 may be lost from circulating erythrocytes. This acquired deficiency of CR1 on erythrocytes may have pathological consequences in patients with SLE.
Anemia, Hemolytic, Autoimmune↗
Relative abundance of two molecular forms of Na+,K+-ATPase in the ferret heart: developmental changes and associated alterations of digitalis sensitivity.
We have previously reported that adult ferret hearts have high affinity and low affinity ouabain-binding sites in approximately equal abundance corresponding, respectively, to alpha(+) and alpha subunits of Na+,K+-ATPase. In the present study, possible changes in relative abundance of the two molecular forms of Na+,K+-ATPase during postnatal development were examined. In 5-day-old or 7- to 10-day-old ferret heart, there was a predominance of the alpha form of Na+,K+-ATPase, the alpha(+)/alpha ratio being approximately 1:7 and 1:4 at 5 and 7-10 days, respectively, in contrast to a 1:1 ratio observed previously in adult heart. Corresponding to the reduced number of high affinity ouabain-binding sites associated with the alpha(+) form of Na+,K+-ATPase, ouabain sensitivity of the Na+,K+-ATPase isolated from newborn ferret heart was lower than that of adult ferret heart. Furthermore, the low dose positive inotropic effect, which was apparent in the adult ferret heart, could not be observed in the newborn ferret heart. The present finding may represent a plausible explanation for low digitalis sensitivity of neonatal heart observed in several mammalian species.
Age Factors↗
A lysine-binding protein in SLE sera inhibits the binding of immune complexes to normal erythrocyte CR1 (complement receptor type 1).
The binding of 125I-labelled immune complexes (IC) to normal human erythrocyte CR1 (complement receptor type 1) by sera from patients with SLE was found to be significantly decreased compared to normal sera. In 13/29 patients, there was an inhibitor which decreased the binding of opsonized IC in normal sera to normal erythrocytes. It was found in each of the nine patients who had clinically active disease. The inhibitor was shown to be a globulin that was labile at 56 degrees C and bound to lysine; low concentrations of tranexamic acid and of lysine abolished the effects of the inhibitor which suggests that it possesses lysine-binding sites: these may block the CR1-binding site on IC opsonized with complement. This inhibitor may decrease the efficiency of IC carriage by erythrocyte CR1.
Antigen-Antibody Complex↗
High affinity and low affinity ouabain binding sites in the rat heart.
Ventricular muscle of rat heart has two classes of receptors which are responsible for the positive inotropic effect of ouabain. Low affinity receptors are apparently related to Na+, K+-ATPase. To determine if high affinity receptors are also sarcolemmal Na+, K+-ATPase of muscle cells, their characteristics were examined. Binding of [3H]ouabain to the high affinity binding site required ATP in the presence of Mg2+ and Na+, was stimulated by Na+ in the presence of Mg2+ and ATP, and was inhibited by K+. Digoxin, digitoxin and cassaine all inhibited [3H]ouabain binding to the high affinity site. Cassaine was about an order of magnitude less potent than the glycosides. These results indicate similarities in high affinity ouabain binding sites in ventricular muscle of rat heart and Na+, K+-ATPase obtained from other sources. Destruction of sympathetic nerve terminals with 6-hydroxydopamine failed to affect the high affinity ouabain binding sites indicating that high affinity sites do not represent the Na+, K+-ATPase in sympathetic nerve terminals. Labeling of Na+, K+-ATPase from [gamma-32P]ATP indicates that high affinity ouabain binding sites account for 25% of the total enzyme molecules present in ventricular muscle of rat heart.
Animals↗
The role of complement and its receptor in the elimination of immune complexes.
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Animals↗
Amiloride: effects on myocardial force of contraction, sodium pump and Na+/Ca2+ exchange.
The direct effects of amiloride on myocardial contractility were examined in electrically stimulated left atrial muscle of guinea-pig heart. Amiloride (0.3 to 1.5 mM) produced a positive inotropic effect which, at higher concentrations, was followed by a decline in developed tension. These effects were not accompanied by contracture or arrhythmia and were not affected by a combination of phentolamine, nadolol, cimetidine, tripelennamine and atropine. The above concentrations of amiloride prolonged the action potential duration during the development of the positive inotropic effect; however, no further change in the action potential duration was observed during the decline in developed tension caused by high concentrations of amiloride. Myocardial membrane Na,K-ATPase, ouabain-sensitive 86Rb+ uptake and Na+-dependent Ca2+ efflux from sarcolemmal membrane vesicles were all inhibited by amiloride. The positive inotropic effect of the agent is reduced and the negative inotropic action is enhanced in low Na+ solutions, i.e., under conditions likely to favor Ca2+ influx via Na+/Ca2+ exchange. These results suggest that amiloride, under the present conditions, has a complex interaction with cardiac muscle fibers. Amiloride may produce its inotropic effects in guinea-pig atrial muscle by several mechanisms including sodium pump inhibition, Na+/Ca2+ exchange inhibition, prolongation of the action potential duration, and/or actions such as Na+/H+ exchange inhibition which were not directly addressed in this study.
Action Potentials↗
The aetiology and pathogenesis of major systemic vasculitides.
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Animals↗
Effects of inotropic agents on isolated guinea pig heart under conditions that modify calcium pools involved in contractile activation.
Positive inotropic effects of strophanthidin were compared with those of isoproterenol, BAY K 8644, grayanotoxin, veratridine, and monensin in electrically stimulated left atrial muscle preparations of guinea pig heart under conditions in which the calcium pool, playing a primary role in contractile activation, was altered. In concentrations that caused similar degrees of increase in developed tension under 1 Hz stimulation, grayanotoxin and strophanthidin caused a relatively large increase in potentiated postrest contraction compared with that caused by isoproterenol, whereas the effect of BAY K 8644 on the postrest contraction was the smallest. The effect of high concentrations of grayanotoxin or strophanthidin, however, resembled that of isoproterenol. The sensitivity of the isolated heart muscle to these agents was compared under conditions in which utilization of various calcium pools contributing to contractile activation was suppressed. Mn2+, which reduces contribution of very superficial Ca2+, reduced sensitivity of heart muscle to the positive inotropic effect of isoproterenol and enhanced the inotropic effect of monensin or veratridine. Verapamil, nifedipine, diltiazem, or ryanodine did not have marked effects on the positive inotropic action of Ca2+, monensin, veratridine, or strophanthidin. These results suggest that the positive inotropic actions of veratridine, grayanotoxin, and strophanthidin share a common mechanism and that low concentrations of strophanthidin may increase loading of Ca2+ pool, which plays an important role in potentiated postrest contraction.
3-Pyridinecarboxylic acid, 1,4-dihydro-2,6-dimethy↗
Effect of nephritic factor on C3 and on the terminal pathway of complement in vivo and in vitro.
Plasma samples from patients with nephritic factor (NeF) were examined for their C3 converting activity. C3, C3dg, C5 and the fluid phase terminal complement complex (TCC) were quantified. All patients had evidence of C3 activation with low plasma C3 and high C3dg. Some patients had normal C5 and normal TCC levels, and thus no evidence of terminal pathway activation in vivo; others, with slower C3 conversion in vitro, had low C5 levels with TCC either elevated or in the upper normal range, suggesting in vivo activation of the terminal pathway. These observations were confirmed by in vitro experiments using purified NeFs. It is concluded that considerable activation of C3 may occur in vivo without a simultaneous activation of the terminal pathway, and that NeF is heterogeneous with regard to its ability to activate complement.
Adult↗
C3 nephritic factor (C3NeF): dissociation of cell-bound and fluid phase stabilization of alternative pathway C3 convertase.
Fluid phase C3 conversion by C3 nephritic factor (C3NeF) is usually easily detectable and forms the basis of the C3NeF screening test. We have described a group of patients with membranoproliferative glomerulonephritis or partial lipodystrophy and hypocomplementaemia who have an unusual C3NeF which stabilizes cell-bound C3 convertase of the alternative pathway (C3bBb) but causes such weak fluid phase C3 conversion that a C3NeF screening test is negative. These patients have low concentrations of C5 in serum.
Adolescent↗
Ascorbic acid: an endogenous inhibitor of isolated Na+,K+-ATPase.
During attempts to isolate and identify an endogenous ligand for the glycoside binding sites on Na+,K+-ATPase, bovine adrenal glands were found to contain a potent inhibitor of isolated Na+,K+-ATPase. The inhibitory principle was extracted from adrenal cortex, following homogenization in NaHCO3 solution and separation on a Sephadex G-10 column. The active principle was recovered from a fraction which eluted from the column after the 3H2O peak. The extract inhibited isolated Na+,K+-ATPase and the specific [3H]ouabain binding reaction. Sensitivity of the enzyme to the inhibitory action of the extract was species and tissue dependent; however, the pattern and the magnitude of the sensitivity were different from those of the digitalis glycosides. Moreover, the inhibitory principle failed to inhibit sodium pump activity, estimated from ouabain inhibitable 86Rb+ uptake by guinea pig brain slices. The activity of the extract to inhibit isolated Na+,K+-ATPase was stable under acidic condition but was lost rapidly at neutral pH, and could be eliminated by EDTA. In an acidic medium, the inhibitory principle had an absorption maximum at 244 nm which shifted to 264 nm and decayed rapidly at neutral pH. By using mass spectrometry, the principle was identified to be ascorbic acid, which has been shown previously to inhibit isolated Na+,K+-ATPase under appropriate conditions. Because ascorbic acid was incapable of inhibiting the sodium pump in intact cells, this inhibitor of the isolated enzyme does not appear to be the endogenous ligand which regulates sodium pump activity in vivo.
Adrenal Glands↗
Effects of K+ on the interaction between cardiac glycosides and Na,K-ATPase.
Inhibition of Na,K-ATPase by cardiac glycosides is at least partially antagonized by K+. The kinetics of the antagonism, however, appear complicated because K+ is capable of reducing both association and dissociation rate constants for the glycoside-enzyme interaction. In order to better understand the effect of K+, inhibition of partially purified Na,K-ATPase obtained from rat brain, guinea-pig heart and rat heart by ouabain, digoxin, digoxigenin, dihydrodigoxin and cassaine were compared in the presence of 1, 3 or 10 mM K+. Higher concentrations of K+ caused a parallel shift to the right in the concentration-inhibition curves for these compounds. For ouabain or digoxin, the extent of the shift was minimal with rat brain enzyme, intermediate with guinea-pig heart enzyme and more substantial with rat heart enzyme. For digoxigenin, dihydrodigoxin or cassaine, the extent of the shift was substantial in all enzyme preparations. These results could not be explained from either the affinity of the enzyme for the compound or its lipid solubility alone. The concentrations of these compounds required to cause a 50 percent inhibition of enzyme activity were markedly different with rat brain enzyme, but relatively similar with rat heart enzyme. The effects of K+, which depend on the source of the enzyme and chemical structures of the compounds, have to be considered in studies on comparative effects of various compounds on Na,K-ATPase, [3H]ouabain binding, sodium pumping and the force of myocardial contraction.
Abietanes↗