Inhibition of Na-K pump current in guinea pig ventricular myocytes by dihydroouabain occurs at high- and low-affinity sites.

Abstract

Binding of cardiac glycosides to the Na + ,K + -dependent ATPase has been shown to occur at both high-and low-affinity sites. However, recent reports suggest that glycoside-induced inhibition of electrogenic Na-K pump current occurs with simple first-order binding kinetics at relatively low-affinity sites. This implies that high-affinity binding sites have little to do with Na-K pump inhibition during exposure to cardiac glycosides. To better understand the role of the high-affinity site, we investigated the concentration dependence of I p^ inhibition by dihydroouabain (DHO) in guinea pig ventricular myocytes through use of wide-pore patch pipettes to "fix" internal Na + activity at -3 0 mM and to voltage clamp at -4 0 mV (T=34° C). DHO was found to have no effect on membrane conductance at a holding potential of -4 0 mV. Holding current was monitored and the difference between steady-state holding current before and during external exposure to nine concentrations (range, 0.01-1,000 /iM) of DHO was measured and normalized to cellular membrane capacitance. The concentration dependence of the inhibition of Na-K pump current was biphasic and well fitted to a two-binding site model with inhibitory K D values of 0.05 fiM and 64.5 fiM. This is consistent with previously reported 3 H-ouabain binding studies in guinea pig myocardium. These findings indicate that the electrogenic properties of the Na-K pump can be inhibited by glyeoside binding to both highand low-affinity sites. {Circulation Research 1989;64:1063-1069)

Dependence of Na+ pump current on external monovalent cations and membrane potential in rabbit cardiac Purkinje cells.

“…3B) indicates that no other current than Ip is affected by a change in the external K+ concentration or by application of the cardiac steroid under our experimental conditions. Corresponding observations were reported from measurements on guinea-pig ventricular myocytes (Gadsby et al 1985;Mogul, Rasmussen, Singer & Ten Eick, 1989;, squid giant axons (Rakowski, Gadsby & De Weer, 183 F. V. BIELEN AND OTHERS 1989) and Xenopus laevis oocytes (Lafaire & Schwarz, 1986;Schweigert, Lafaire & Schwarz, 1988).…”

beta-Adrenergic modulation of the inwardly rectifying potassium channel in isolated human ventricular myocytes. Alteration in channel response to beta-adrenergic stimulation in failing human hearts.

“…To eliminate sodium ('Na) and L-type calcium current ('Ca), tetrodotoxin (TTX, 5 MM) and Co2" (2 mM) were added to the external solution. Dihydroouabain (DHO, 10 kM) was added to the external solution to block the Na+/K' pump current (40). Na+/K' pump is known to be enhanced by fi-adrenergic stimulation (1,41,42).…”

Digitalis: new actions for an old drug

“…The dog heart has mostly ␣ 1 -and ␣ 3 -isoforms, whereas ␣ 1 -isoform predominates in sheep and guinea pigs (120), despite the fact that all three species are glycoside sensitive. It should be noted, however, that there is evidence that both a glycoside-sensitive and -insensitive isoform exist in the guinea pig that may correlate with the ␣ 2 -and ␣ 1 -isoforms found in rats, respectively (37,38,87). The fact that these isoforms are conserved across species has improved our understanding of their individual role in ion regulation in cardiac cells but has made it difficult to understand how different species might have different sensitivities to glycoside even though they share similar ␣-isoform distribution.…”