Action of ouabain on rat heart: comparison with its effect on guinea‐pig heart

Abstract

The inotropic dose‐response curve of ouabain in rat cardiac ventricular strips exceeded a concentration range of two decades (1 × 10−7 M to 3 × 10−5 M) displaying an intermediate plateau phase. In guinea‐pig ventricular strips the inotropic ouabain concentrations spanned only one decade (1 × 10−7 M‐1 × 10−6 M). Ouabain‐intoxication in guinea‐pig ventricular strips occurring at 3 × 10−6 M consisted of arrhythmia and contracture, while in rat ventricular strips at the toxic concentration of 1 × 10−4 M only a progressive increase in diastolic tension was observed. By means of atomic absorption spectroscopy the ouabain‐induced loss of cellular potassium and gain of sodium in rat ventricular strips was detected only at concentrations of ouabain higher than 10−4 M. Ouabain reduced the activity of Na/K‐ATPase prepared from rat and guinea‐pig cardiac ventricles to half of its maximum at 6.5 × 10−5 M in rat and 1.0 × 10−6 M in guinea‐pig, rat heart Na/K‐ATPase thus being about 60 fold less sensitive towards ouabain. Specific [3H]‐ouabain binding to membrane suspensions prepared from rat and guinea‐pig ventricles was characterized by a similar affinity in rat (KD = 4 × 10−8 M) and guinea‐pig (KD = 13 × 10−8 M). The number of ouabain binding sites in rat membranes was only about 10% of the number found in guinea‐pig membranes. In rat the presence of additional ouabain‐binding with low affinity and high capacity seemed possible, but could not be verified for methodological reasons. In the light of the biochemical results and binding data, the wider range of ouabain concentration exerting a positive inotropic effect in the rat may be attributed to the existence in the latter of two populations of receptors with different affinities for ouabain and different capacities. In contrast, in the guinea‐pig, there is a single population. Nevertheless it is probable that all the receptors in both species are part of the Na/K‐ATPase complex and mediate a positive inotropic effect after ouabain‐binding in an identical manner.

Two functional Na+/K+‐ATPase isoforms in the left ventricle of guinea pig heart

“…One functional enzyme form may exist according to the following different approaches : the electrophysiological properties of isolated myocytes [4], the Na+/K+-ATPase activity [5,6], the [3H]ouabain-binding sites on microsomal preparations with low Na+/K +-ATPase activity [7], gel electrophoresis mobility and immunoblotting with specific antibodies [8, 91, and the force of isometric contraction of ventricular strips induced by ouabain [5]. The highly controversial aspect of these data is reflected by the scattering of the IC50 values: the apparent sensitivity of the enzyme (or site) to ouabain has been estimated as 0.130 pM 171, 2.2 pM [6] and 46 pM [4].…”

Sodium load and high affinity ouabain binding in rat and guinea‐pig cardiac tissue

“…Reduction of the stimulation frequency to 0.3 Hz, which induced a positive inotropic effect in rat and a negative inotropic effect in guinea-pig, diminished°- (Herzig & Mohr, 1984 (Forssmann & Girardier, 1970) leading to a potassium-induced reduction of ouabain affinity; or a decrease in the number of high affinity binding sites could occur at a high beat frequency, if high-and low-affinity binding sites were interconvertible (Mansier & Lelievre, 1982;Grupp et al, 1984). In any case, the absence of a frequency-induced elevation of [3H]-ouabain binding matches the observation that ouabain is less effective at high beat frequencies with regard to the positive inotropism in rats (Arletti & Bazzani, 1982).…”

“…Strips were prepared from rat and guinea-pig right ventricles as described previously (Herzig & Mohr, 1984). The preparations were equilibrated for 1 h in an organ bath containing 500 ml of a modified Tyrode solution (in mM: NaCl 136.8, KCI 5.4, CaC12 1.8, NaHCO3 11.9, MgCl2 1.05, NaH2PO4 0.21, glucose 5.5) gassed with carbogen (95% 02; 5% C02) at a stimulation frequency of 1.5 Hz (rectangular pulse, duration 5 ms, intensity about 30% over threshold), and maintained at a constant temperature of 32°C.…”

“…The existence of two populations of ouabain binding sites in rat heart ventricular muscle, both related to positive inotropy, has been shown in recent studies (Erdmann et al, 1980: Adams et al, 1982Finet et al, 1983;Noel & Godfraind, 1984;Herzig & Mohr, 1984). The low-affinity, high-capacity ouabain binding sites are Na/K-ATPase molecules.…”

“…The low-affinity, high-capacity ouabain binding sites are Na/K-ATPase molecules. It has been suggested that the high-affinity, low-capacity binding sites could be either (a) Na/K-ATPase molecules involved in Na/ K-transport, (b) inactive isozymes, or (c) completely different membrane proteins (Adams et al, 1982;Noel & Godfraind, 1984;Herzig & Mohr, 1984). Noel & Godfraind (1984) found that high-affinity ouabain binding corresponded to an inhibition of ATPaseactivity in rat heart microsomal preparations.…”