Ouabain augments Ca(2+) transients in arterial smooth muscle without raising cytosolic Na(+).
Ouabain and other cardiotonic steroids (CTS) inhibit Na(+) pumps and are widely believed to exert their cardiovascular effects by raising the cytosolic Na(+) concentration ([Na(+)](cyt)) and Ca(2+). This view has not been rigorously reexamined despite evidence that low-dose CTS may act without elevating [Na(+)](cyt); also, it does not explain the presence of multiple, functionally distinct isoforms of the Na(+) pump in many cells. We investigated the effects of Na(+) pump inhibition on [Na(+)](cyt) (with Na(+) binding benzofuran isophthalate) and Ca(2+) transients (with fura 2) in primary cultured arterial myocytes. Low concentrations of ouabain (3-100 nM) or human ouabain-like compound or reduced extracellular K(+) augmented hormone-evoked mobilization of stored Ca(2+) but did not increase bulk [Na(+)](cyt). Augmentation depended directly on external Na(+), but not external Ca(2+), and was inhibited by 10 mM Mg(2+) or 10 microM La(3+). Evoked Ca(2+) transients in pressurized small resistance arteries were also augmented by nanomolar ouabain and inhibited by Mg(2+). These results suggest that Na(+) enters a tiny cytosolic space between the plasmalemma (PL) and the adjacent sarcoplasmic reticulum (SR) via an Mg(2+)- and La(3+)-blockable mechanism that is activated by SR store depletion. The Na(+) and Ca(2+) concentrations within this space may be controlled by clusters of high ouabain affinity (alpha3) Na(+) pumps and Na/Ca exchangers located in PL microdomains overlying the SR. Inhibition of the alpha3 pumps by low-dose ouabain should raise the local concentrations of Na(+) and Ca(2+) and augment hormone-evoked release of Ca(2+) from SR stores. Thus the clustering of small numbers of specific PL ion transporters adjacent to the SR can regulate global Ca(2+) signaling. This mechanism may affect vascular tone and blood flow and may also influence Ca(2+) signaling in many other types of cells.
Arnon
A
A
Department of Physiology, University of Maryland School of Medicine, Baltimore, Maryland 21201, USA.
Hamlyn
J M
JM
Blaustein
M P
MP
D000818:Animals; D001127:Arginine Vasopressin / Q000494:pharmacology; D001685:Biological Factors / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism*; D020013:Calcium Signaling / Q000187:drug effects / Q000502:physiology*; D002298:Cardenolides; D002316:Cardiotonic Agents / Q000494:pharmacology; D002478:Cells, Cultured; D003600:Cytosol / Q000378:metabolism; D004077:Digoxin; D006801:Humans; D007700:Kinetics; D008638:Mesenteric Arteries / Q000166:cytology / Q000187:drug effects / Q000502:physiology*; D008955:Models, Cardiovascular; D009131:Muscle, Smooth, Vascular / Q000166:cytology / Q000187:drug effects / Q000502:physiology*; D010042:Ouabain / Q000494:pharmacology*; D010656:Phenylephrine / Q000494:pharmacology; D011188:Potassium / Q000494:pharmacology; D051381:Rats; D012503:Saponins; D012701:Serotonin / Q000494:pharmacology; D012964:Sodium / Q000378:metabolism*; D013779:Tetrodotoxin / Q000494:pharmacology
D001685:Biological Factors; D002298:Cardenolides; D002316:Cardiotonic Agents; D012503:Saponins; C034504:digoxin-like factors; D001127:Arginine Vasopressin; D010656:Phenylephrine; D012701:Serotonin; D013779:Tetrodotoxin; D010042:Ouabain; D004077:Digoxin; D012964:Sodium; D011188:Potassium; D002118:Calcium