Marinobufagenin enhances cardiac contractility in mice with ouabain-sensitive alpha(1) Na+-K+-ATPase
ABSTRACT Endogenous Na(+) pump inhibitors are thought to play important (patho)physiological roles and occur in two different chemical forms in the mammalian circulation: cardenolides, such as ouabain, and bufadienolides, such as marinobufagenin (MBG). Although all alpha Na(+)-K(+)-ATPase isoforms (alpha(1-4)) are sensitive to ouabain in most species, in rats and mice the ubiquitously expressed alpha(1) Na(+)-K(+)-ATPase is resistant to ouabain. We have previously shown that selective modification of the putative ouabain binding site of either the alpha(1) or alpha(2) Na(+)-K(+)-ATPase subunit in mice substantially alters the cardiotonic influence of exogenously applied cardenolides. To determine whether the ouabain binding site also interacts with MBG and if this interaction plays a functional role, we evaluated cardiovascular function in alpha(1)-resistant/alpha(2)-resistant (alpha(1)(R/R)alpha(2)(R/R)), alpha(1)-sensitive/alpha(2)-resistant (alpha(1)(S/S)alpha(2)(R/R)), and alpha(1)-resistant/alpha(2)-sensitive mice (alpha(1)(R/R)alpha(2)(S/S), wild type). Cardiovascular indexes were evaluated in vivo by cardiac catheterization at baseline and during graded infusions of MBG. There were no differences in baseline measurements of targeted mice, indicating normal hemodynamics and cardiac function. MBG at 0.025, 0.05, and 0.1 nmol*min(-1)*g body wt(-1) significantly increased cardiac performance to a greater extent in alpha(1)(S/S)alpha(2)(R/R) compared with alpha(1)(R/R)alpha(2)(R/R) and wild-type mice. The increase in LVdP/dt(max) in alpha(1)(S/S)alpha(2)(R/R) mice was greater at higher concentrations of MBG compared with both alpha(1)(R/R)alpha(2)(R/R) and alpha(1)(R/R)alpha(2)(S/S) mice (P < 0.05). These results suggest that MBG interacts with the ouabain binding site of the alpha(1) Na(+)-K(+)-ATPase subunit and can thereby influence cardiac inotropy.
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- "Both are sensitive to ouabain in most species, including guinea pigs, however in rats and mice the ubiquitously expressed a1 subunit of Na + –K + -ATPase is resistant to ouabain. The transgenic " humanized " mice with R111Q and D122N substitutions in the a 1 isoform were used (Wansapura et al. 2009). In these mice, Na + –K + -ATPase a 1 was 2 orders of magnitude more sensitive to ouabain compared with the wild type mice. "
ABSTRACT: Natriuretic peptides and digitalis-like compounds serve as regulators of homeostasis, including control of volume expansion and blood pressure. The aim of the present study was to explore possible interactions between atrial natriuretic peptide (ANP) and ouabain in the heart. ANP (1 nmol/L) had no effect in papillary muscle preparations from guinea pigs. Ouabain (1 µmol/L) induced positive inotropic effect. The addition of ANP prior to ouabain resulted in a significant decrease in the ouabain-induced positive inotropic effect, manifested as an attenuated increase in twitch maximal upward force slope and resting muscular tension. In addition, ANP caused an increase in Na(+)-K(+)-ATPase activity in heart microsomal preparations. The effect of ouabain on Na(+)-K(+)-ATPase activity was shown in a biphasic manner. Ouabain (0.01-1 nmol/L) had a small but significant increase on pump activity, but higher doses of ouabain inhibited activity. ANP attenuated ouabain-induced Na(+)-K(+)-ATPase activity. Furthermore, ouabain (50 nmol/L) or ANP (10 nmol/L) alone induced Akt activation in cardiomyocytes. However, ANP blocked ouabain-induced Akt activation. These results point to the existence of interactions between ANP and ouabain on Na(+)-K(+)-ATPase signaling and function in the heart, which may be mediated by regulation of Na(+)-K(+)-ATPase activity and (or) signal transduction mechanisms.Canadian Journal of Physiology and Pharmacology 09/2012; 90(10):1386-93. DOI:10.1139/y2012-112 · 1.55 Impact Factor
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ABSTRACT: Mechanisms underlying the tissue-specific impact of cardiotonic steroids (CTS) on cell survival and death remain poorly understood. This study examines the role of Na(+),K(+)-ATPase alpha subunits in death of Madin-Darby canine kidney (MDCK) cells evoked by 24-h exposure to ouabain. MDCK cells expressing a variant of the alpha1 isoform, CTS-sensitive alpha1S, were stably transfected with a cDNA encoding CTS-resistant alpha1R-Na(+),K(+)-ATPase, whose expression was confirmed by RT-PCR. In mock-transfected and alpha1R-cells, maximal inhibition of (86)Rb influx was observed at 10 and 1000 muM ouabain, respectively, thus confirming high abundance of alpha1R-Na(+),K(+)-ATPase in these cells. Six-hour treatment of alpha1R-cells with 1000 muM ouabain led to the same elevation of the [Na(+)](i)/[K(+)](i) ratio that was detected in mock-transfected cells treated with 3 muM ouabain. However, in contrast to the massive death of mock-transfected cells exposed to 3 muM ouabain, alpha1R-cells survived after 24-h incubation with 1000 muM ouabain. Inversion of the [Na(+)](i)/[K(+)](i) ratio evoked by Na(+),K(+)-ATPase inhibition in K(+)-free medium did not affect survival of alpha1R-cells but increased their sensitivity to ouabain. Our results show that the alpha1R subunit rescues MDCK cells from the cytotoxic action of CTS independently of inhibition of Na(+),K(+)-ATPase-mediated Na(+) and K(+) fluxes and inversion of the [Na(+)](i)/[K(+)](i) ratio.Apoptosis 12/2009; 15(1):55-62. DOI:10.1007/s10495-009-0429-4 · 3.61 Impact Factor
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ABSTRACT: The cardiotonic steroid/ouabain-binding site of the α subunit of Na,K-ATPase is thought to play an important role in cardiovascular homeostasis. Previously, we demonstrated the cardiotonic steroid-binding site of the α2 Na,K-ATPase is involved in adrenocorticotropic hormone (ACTH)-induced hypertension by using gene-modified α2(R/R) mice in which the cardiotonic steroid-binding site is relatively resistant to ouabain compared to the ouabain-sensitive wild-type α2(S/S) mice. To further explore the importance of this site in the cardiovascular system, we investigated blood pressure regulation during pregnancy in mice with the α2(R/R) isoform. The systolic blood pressure (SBP) of the α2(S/S) and α2(R/R) mice was measured before and during pregnancy by tail-cuff. The expression of the α isoforms of Na, K-ATPase in various tissues and plasma endogenous ouabain contents were assessed prior to pregnancy as well as days 7 and 17 of gestation. The α2(S/S) mice showed a gradual decrease in the SBP during the first two trimesters, followed by an increase above the preconceptional level in the third trimester. However, the α2(R/R) mice exhibited a lower blood pressure in the third trimester. The cardiac expression of the α2 Na,K-ATPase in the α2(S/S) mice was significantly less than that of the α2(R/R) mice throughout the pregnancy. The plasma endogenous ouabain concentration significantly increased by twofold at day 17 of pregnancy in the α2(R/R) mice but not in the α2(S/S) mice. The cardiotonic steroid-binding site of the α2 Na,K-ATPase plays a role in maintaining normal SBP during pregnancy.American Journal of Hypertension 09/2010; 23(12):1279-85. DOI:10.1038/ajh.2010.195 · 3.40 Impact Factor