title: Ouabain-digoxin antagonism in rat arteries and neurones.
abstract: 'Classic' cardiotonic steroids (CTSs) such as digoxin and ouabain selectively inhibit Na+, K+ -ATPase (the Na+ pump) and, via Na+ / Ca2+ exchange (NCX), exert cardiotonic and vasotonic effects. CTS action is more complex than previously thought: prolonged subcutaneous administration of ouabain, but not digoxin, induces hypertension, and digoxin antagonizes ouabain's hypertensinogenic effect. We studied the acute interactions between CTSs in two indirect assays of Na+ pump function: myogenic tone (MT) in isolated, pressurized rat mesenteric small arteries, and Ca2+ signalling in primary cultured rat hippocampal neurones. The 'classic' CTSs (0.3-10 nm) behaved as 'agonists': all increased MT70 (MT at 70 mmHg) and augmented glutamate-evoked Ca2+ (Fura-2) signals. We then tested one CTS in the presence of another. Most CTSs could be divided into ouabain-like (ouabagenin, dihydroouabain (DHO), strophanthidin) or digoxin-like CTS (digoxigenin, digitoxin, bufalin). Within each group, the CTSs were synergistic, but ouabain-like and digoxin-like CTSs antagonized one another in both assays: For example, the ouabain-evoked (3 nm) increases in MT70 and neuronal Ca2+ signals were both greatly attenuated by the addition of 10 nm digoxin or 10 nm bufalin, and vice versa. Rostafuroxin (PST2238), a digoxigenin derivative that displaces 3H-ouabain from Na+, K+ -ATPase, and attenuates some forms of hypertension, antagonized the effects of ouabain, but not digoxin. SEA0400, a Na+ / Ca2+ exchanger (NCX) blocker, antagonized the effects of both ouabain and digoxin. CTSs bind to the α subunit of pump αβ protomers. Analysis of potential models suggests that, in vivo, Na+ pumps function as tetraprotomers ((αβ)4) in which the binding of a single CTS to one protomer blocks all pumping activity. The paradoxical ability of digoxin-like CTSs to reactivate the ouabain-inhibited complex can be explained by de-oligomerization of the tetrameric state. The interactions between these common CTSs may be of considerable therapeutic relevance.
authors.affiliation: Department of Physiology, University of Maryland School of Medicine, 655 W. Baltimore Street, Baltimore, MD 21201, USA.  mblaustein@som.umaryland.edu or jhamlyn@umaryland.edu.
authors.forename: Hong
authors.initials: H
authors.lastname: Song
authors.forename: Eiji
authors.initials: E
authors.lastname: Karashima
authors.forename: John M
authors.initials: JM
authors.lastname: Hamlyn
authors.forename: Mordecai P
authors.initials: MP
authors.lastname: Blaustein
mesh_terms: D000818:Animals; D001783:Blood Flow Velocity / Q000187:drug effects / Q000502:physiology; D020013:Calcium Signaling / Q000187:drug effects / Q000502:physiology*; D002316:Cardiotonic Agents / Q000008:administration & dosage; D002478:Cells, Cultured; D004077:Digoxin / Q000008:administration & dosage*; D004305:Dose-Response Relationship, Drug; D004347:Drug Interactions / Q000502:physiology; D006624:Hippocampus / Q000166:cytology / Q000187:drug effects / Q000502:physiology*; D066298:In Vitro Techniques; D008297:Male; D008638:Mesenteric Arteries / Q000187:drug effects / Q000502:physiology*; D009474:Neurons / Q000166:cytology / Q000187:drug effects / Q000502:physiology*; D010042:Ouabain / Q000008:administration & dosage*; D051381:Rats; D017207:Rats, Sprague-Dawley; D014655:Vascular Resistance / Q000187:drug effects / Q000502:physiology; D014661:Vasoconstriction / Q000187:drug effects / Q000502:physiology*
chemical_list: D002316:Cardiotonic Agents; D010042:Ouabain; D004077:Digoxin