The action of palytoxin on erythrocytes and resealed ghosts

Chhatwal, Gursharan S.; Hessler, H.-J.; Habermann, E.

doi:10.1007/bf00497672

Cited by 52 publications

(5 citation statements)

References 23 publications

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“…Though that fall in P o would contribute to an increase in K 0.5PTX (though <10-fold), the marked steroid-mediated reduction we observed in apparent affinity for PTX action (e.g., Figs. 10 and 11 , as previously reported: Chhatwal et al, 1983 ; Habermann, 1989 ) and the acceleration of PTX unbinding ( Fig. 11 C) both indicate that, when also bound to the Na/K pump, the steroids directly increase K dPTX .…”

Section: Discussionsupporting

confidence: 81%

“…In contrast, the affinity for PTX seemed only modestly enhanced by low-affinity nucleotide binding, regardless of whether the pumps were phosphorylated (K dPTX ∼300 pM with either 1 μM or 5 mM MgATP) or unphosphorylated (K dPTX ∼22 nM without nucleotides vs. ∼8 nM with 2 mM MgAMPPNP). Similarly, in work on red blood cell membranes, ATP, and to a lesser extent ADP, was shown to promote PTX-mediated cation flux ( Chhatwal et al, 1983 ), to enhance 125 I-PTX binding ( Böttinger et al, 1986 ), and to promote opening of channels in inside-out patches when PTX was in the pipette ( Kim et al, 1995 ).…”

Section: Discussionmentioning

confidence: 98%

“…Despite much work (for review see Habermann, 1989 ; Tosteson, 2000 ), a consistent picture of how palytoxin interacts with the Na/K pump has yet to emerge. This is undoubtedly due to the variety of methods, preparations, and Na/K pump isoforms examined, and the limited range of experimental conditions employed in each study ( Tosteson, 2000 ), as there were early indications that the effects of palytoxin could be influenced by, among other things, extracellular K ions ( Ahnert-Hilger et al, 1982 ; Ishida et al, 1983 ; Böttinger et al, 1986 ) or cytoplasmic nucleotides ( Chhatwal et al, 1983 ; Böttinger et al, 1986 ; Kim et al, 1995 ). Here we describe an extensive study of palytoxin interactions with native Na/K pumps in HEK293 cells (a human embryonic kidney cell line) and in guinea pig ventricular myocytes.…”

Section: Introductionmentioning

confidence: 99%

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Large Diameter of Palytoxin-induced Na/K Pump Channels and Modulation of Palytoxin Interaction by Na/K Pump Ligands

Artigas

Gadsby

2004

The Journal of General Physiology

104

114

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Palytoxin binds to Na/K pumps to generate nonselective cation channels whose pore likely comprises at least part of the pump's ion translocation pathway. We systematically analyzed palytoxin's interactions with native human Na/K pumps in outside-out patches from HEK293 cells over a broad range of ionic and nucleotide conditions, and with or without cardiotonic steroids. With 5 mM internal (pipette) [MgATP], palytoxin activated the conductance with an apparent affinity that was highest for Na+-containing (K+-free) external and internal solutions, lowest for K+-containing (Na+-free) external and internal solutions, and intermediate for the mixed external Na+/internal K+, and external K+/internal Na+ conditions; with Na+ solutions and MgATP, the mean dwell time of palytoxin on the Na/K pump was about one day. With Na+ solutions, the apparent affinity for palytoxin action was low after equilibration of patches with nucleotide-free pipette solution. That apparent affinity was increased in two phases as the equilibrating [MgATP] was raised over the submicromolar, and submillimolar, ranges, but was increased by pipette MgAMPPNP in a single phase, over the submillimolar range; the apparent affinity at saturating [MgAMPPNP] remained ∼30-fold lower than at saturating [MgATP]. After palytoxin washout, the conductance decay that reflects palytoxin unbinding was accelerated by cardiotonic steroid. When Na/K pumps were preincubated with cardiotonic steroid, subsequent activation of palytoxin-induced conductance was greatly slowed, even after washout of the cardiotonic steroid, but activation could still be accelerated by increasing palytoxin concentration. These results indicate that palytoxin and a cardiotonic steroid can simultaneously occupy the same Na/K pump, each destabilizing the other. The palytoxin-induced channels were permeable to several large organic cations, including N-methyl-d-glucamine+, suggesting that the narrowest section of the pore must be ∼7.5 Å wide. Enhanced understanding of palytoxin action now allows its use for examining the structures and mechanisms of the gates that occlude/deocclude transported ions during the normal Na/K pump cycle.

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Section: Discussionsupporting

confidence: 81%

Section: Discussionmentioning

confidence: 98%

Section: Introductionmentioning

confidence: 99%

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Large Diameter of Palytoxin-induced Na/K Pump Channels and Modulation of Palytoxin Interaction by Na/K Pump Ligands

Artigas

Gadsby

2004

The Journal of General Physiology

104

114

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“…Although the mechanism of action of the Na/K pump is not completely understood, in recent years, the Albers-Post model has gained increasing support as a description of the Na/K pump in terms of alternating gates [1,2]. Evidence in favor of this model has come from experiments in which palytoxin is used to disturb ionic transport across the membrane via the appearance of nonselective monovalent cation channels [3][4][5][6][7][8][9]. In this work we extend the Albers-Post model to include the effect of palytoxin on the Na/K pump, in efforts to understand the experimental results of [7].…”

Section: Introductionmentioning

confidence: 99%

Effect of palytoxin on the sodium–potassium pump: model and simulation

2008

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We propose a reaction model for the palytoxin-sodium-potassium (PTX-Na(+)/K(+)) pump complex. The model, which is similar to the Albers-Post model for Na(+)/K(+)-ATPase, is used to elucidate the effect of PTX on Na(+)/K(+)-ATPase during the enzyme interactions with Na(+) and/or K(+) ions. Conformational substates and reactions for the pump are incorporated into the Albers-Post model to represent enzymes with or without bound PTX. A mathematical model based on the reaction scheme is used in simulations modeling experimental studies of PTX-induced ionic currents. Our simulations suggest that (i) extracellular Na(+) as well as K(+) promotes PTX-induced channel blockage; (ii) extracellular K(+) accelerates PTX unbinding; and (iii) K(+) occlusion in the PTX-pump complex is essential for describing the PTX-induced current dynamics.

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“…The PTX-induced depolarization has been shown to depend on extracellular Na ions. It also causes K + release from erythrocytes [15,16] and smooth muscles [17, 1 XI. Recently it has been reported that PTX does not increase the permeability of artificial lipid membranes [18, 191.…”

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confidence: 99%

Interaction of palytoxin and cardiac glycosides on erythrocyte membrane and (Na+ + K+) ATPase

1985

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Palytoxin (PTX), at extremely low concentrations (0.01 -1 nM), caused K f release from rabbit erythrocytes.Among the various chemical compounds tested, cardiac glycosides potently inhibited the PTX-induced K + release.The order of inhibitory potency (ICs0) was cymarin (0.42 pM) > convallatoxin (0.9 pM) > ouabain (2.3 pM) > digitoxin (88 pM) > digoxin (90 pM). Their corresponding aglycones, even at 10 pM, did not inhibit the K' release, but competitively antagonized the inhibitory effect of the glycosides. All these cardiotonic steroids inhibited the activity of (Na' + K+)-ATPase prepared from hog cerebral cortex in narrow concentration ranges (IC50 = 0.1 5 -2.4 pM), suggesting that the inhibition of K + release is not related to their inhibitory potency on the (Na' + K+)-ATPase activity, and the sugar moiety of cardiac glycosides is involved in the inhibition. On the other hand PTX, at higher concentrations (> 0.1 pM), inhibited the (Na' + K+)-ATPase activity. However, this inhibitory effect of PTX was not antagonized by ouabain. It is suggested that, compared with ouabain, PTX has additional binding site(s) on the (Na' + K+)-ATPase.Palytoxin (PTX; C129H223N3054; M , = 2677), isolated from marine coelenterates of some zoanthid species (genus Palythoa), is the most potent animal toxin known to date [l]. Its stereochemistry has recently been determined [2, 31. It consists of a long partially unsaturated aliphatic chain, interspaced with five sugar moieties.In the nanomolar concentration range PTX depolarizes cardiac [4-81, smooth [9] and skeletal muscles [4, 101 and nerve tissues [ l l -141. The PTX-induced depolarization has been shown to depend on extracellular Na ions. It also causes K + release from erythrocytes [15,16] and smooth muscles [17, 1 XI. Recently it has been reported that PTX does not increase the permeability of artificial lipid membranes [18, 191. This suggests that some membrane proteins seem to be required for the PTX action.In the present experiments we tested the effects of various chemical compounds on the effects of PTX in erythrocytes. Among the compounds tested it was found that cardiac glycosides potently inhibited the K + release. In this paper we report the effects of several types of cardiac glycosides and their aglycones on the actions of PTX. A part of this work has been reported [20]. MATERIALS AND METHODSErythrocytes from heparinized (30 IU/ml) blood from rabbits, rats and humans were washed three times in 5 vol. physiological salt solution (136.9 mM NaCl, 1.0 mM CaC12, 1.0 mM MgC12, 5.5 mM glucose, 10 mM Hepes, pH 7.3) by centrifugation at 600 -1200 x g for 5 min. Buffy coat wasCorrespondence to H. Ozaki, Department of Veterinary Pharmacology, Faculty of Agriculture, University of Tokyo, Yayoi 1-1-1.Bunkyo-ku, Tokyo-shi, Tokyo-to, Japan 11 3Abbreviations. PTX, palytoxin isolated from Pulyfhoa tuherculosa; ICs0, concentration causing 50% inhibition. removed by aspiration. Packed cells were resuspended in physiological salt solution (approximately 100000 cells/ml) and stored at ...

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The action of palytoxin on erythrocytes and resealed ghosts

Cited by 52 publications

References 23 publications

Large Diameter of Palytoxin-induced Na/K Pump Channels and Modulation of Palytoxin Interaction by Na/K Pump Ligands

Large Diameter of Palytoxin-induced Na/K Pump Channels and Modulation of Palytoxin Interaction by Na/K Pump Ligands

Effect of palytoxin on the sodium–potassium pump: model and simulation

Interaction of palytoxin and cardiac glycosides on erythrocyte membrane and (Na+ + K+) ATPase

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