Binding of (/sup 3/H)ouabain to myocytes isolated from guinea-pig heart

Conference · Wed Mar 05 04:00:00 EST 1986 · Fed. Proc., Fed. Am. Soc. Exp. Biol.; (United States)

OSTI ID:7024951

Stemmer, P; Akera, T; Brody, T M

Accurate estimation of the specific binding of (/sup 3/H)ouabain in intact tissues is difficult. Toxic actions of non-labeled ouabain at high concentrations is necessary to prevent specific binding of (/sup 3/H)ouabain to Na,K-ATPase caused a loss of cell viability and non-specific binding to be underestimated in these preparations. Dissociation of (/sup 3/H)ouabain from guinea-pig myocytes over 180 min occurred in two phases. The fast-releasing component, which had a half-life of approximately 8 min, accounted for greater than 85% of all (/sup 3/H)ouabain bound to myocytes following a 60-min incubation in the presence of 50 nM (/sup 3/H)ouabain. (/sup 3/H)ouabain bound to the slow releasing site increased with incubation time and with increasing (/sup 3/H)ouabain concentration. Analysis of (/sup 3/H)ouabain binding to the fast releasing site yielded non-linear Scatchard plots. Non-linearity appears to result from reduced (/sup 3/H)ouabain binding due to low intracellular Na/sup +/ concentration, especially at low (/sup 3/H)ouabain concentrations. Addition of 2 ..mu..M monesin, a Na/sup +/ ionophore, promoted binding of (/sup 3/H)ouabain in a manner that Scatchard plots became linear. These results indicate that the fast releasing component of (/sup 3/H)ouabain binding to guinea-pig myocytes represents a single binding site and that at high concentrations (/sup 3/H)ouabain promotes its binding to this site by elevating intracellular Na/sup +/.

• Research Organization: Michigan State Univ., East Lansing
• OSTI ID: 7024951
• Report Number(s): CONF-8604222-
• Conference Information: Journal Name: Fed. Proc., Fed. Am. Soc. Exp. Biol.; (United States) Journal Volume: 45:4
• Country of Publication: United States
• Language: English

Related Subjects

550201* -- Biochemistry-- Tracer Techniques
59 BASIC BIOLOGICAL SCIENCES
ACID ANHYDRASES
ALKALI METAL COMPOUNDS
ANIMAL CELLS
ANIMALS
ATP-ASE
BIOCHEMICAL REACTION KINETICS
BIOLOGICAL HALF-LIFE
BODY
CARBOHYDRATES
CARDIAC GLYCOSIDES
CARDIOTONICS
CARDIOVASCULAR AGENTS
CARDIOVASCULAR SYSTEM
DRUGS
ENZYMES
GLYCOSIDES
GUINEA PIGS
HEART
HYDROLASES
ISOTOPE APPLICATIONS
KINETICS
LABELLED COMPOUNDS
MAMMALS
MEMBRANE PROTEINS
ORGANIC COMPOUNDS
ORGANS
OUABAIN
PHOSPHOHYDROLASES
PROTEINS
REACTION KINETICS
RECEPTORS
RODENTS
SODIUM COMPOUNDS
STROPHANTHINS
TRACER TECHNIQUES
TRITIUM COMPOUNDS
VERTEBRATES
VIABILITY