This application proposes to continue our studies of the Na,K-ATPase and is directed toward several ongoing goals. One of these is to test whether the cardiac glycoside binding site of the alpha2 or alpha3 isoforms of this enzyme, which is highly conserved among species, plays a biological role. Such studies will provide insight as to whether endogenous cardiac glycosides such as endogenous ouabain observed by many laboratories is physiologically significant. Another objective is to define the role of the alpha2 isoform in heart and vascular smooth muscle by analyzing tissue-specific alpha2 isoform knockout animals.
The specific aims of our grant are (1) To determine whether the cardiac glycoside binding site of the Na,K-ATPase has biological significance and whether endogenous cardiac glycosides play a physiological role. To accomplish this, genetically engineered mice expressing either ouabain-resistant alpha2 or alpha3 isoforms will be analyzed under conditions known to increase the levels of endogenous cardiac glycosides such as ACTH-induced hypertension. If the ouabain-resistant mice respond differently from wild type animals, this will suggest a physiological role for endogenous ouabain. However, if mice with ouabain resistant alpha2 or alpha3 isoforms respond similarly to wild type animals, this will detract from the hypothesis that endogenous cardiac glycosides have physiological significance. (2) To define the specific role of the alpha2 isoform using tissuespecific knockouts. We have already developed mice where the alpha2 isoform gene is flanked by loxP sites and we have mated these to animals carrying Cre-recombinase under the control of a cardiac-specific promoter. We will now mate the alpha2 isoform loxP animals with those expressing Cre-recombinase under the control of a smooth muscle promoter. These matings will provide animals where the alpha2 isoform is missing only in heart or smooth muscle and this will allow us to study the specific role of this isoform in cardiovascular function.

Agency
National Institute of Health (NIH)
Institute
National Heart, Lung, and Blood Institute (NHLBI)
Type
Research Project (R01)
Project #
3R01HL066062-08S1
Application #
7822942
Study Section
Cardiac Contractility, Hypertrophy, and Failure Study Section (CCHF)
Program Officer
Wang, Lan-Hsiang
Project Start
2009-06-01
Project End
2010-10-31
Budget Start
2009-06-01
Budget End
2010-10-31
Support Year
8
Fiscal Year
2009
Total Cost
$67,510
Indirect Cost
Name
University of Cincinnati
Department
Genetics
Type
Schools of Medicine
DUNS #
041064767
City
Cincinnati
State
OH
Country
United States
Zip Code
45221
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Wansapura, Arshani N; Lasko, Valerie M; Lingrel, Jerry B et al. (2011) Mice expressing ouabain-sensitive ?1-Na,K-ATPase have increased susceptibility to pressure overload-induced cardiac hypertrophy. Am J Physiol Heart Circ Physiol 300:H347-55
Lingrel, Jerry B (2010) The physiological significance of the cardiotonic steroid/ouabain-binding site of the Na,K-ATPase. Annu Rev Physiol 72:395-412
Oshiro, Naomi; Dostanic-Larson, Iva; Neumann, Jon C et al. (2010) The ouabain-binding site of the ?2 isoform of Na,K-ATPase plays a role in blood pressure regulation during pregnancy. Am J Hypertens 23:1279-85
Hou, Xiaohong; Theriault, Steven F; Dostanic-Larson, Iva et al. (2009) Enhanced pressor response to increased CSF sodium concentration and to central ANG I in heterozygous alpha2 Na+ -K+ -ATPase knockout mice. Am J Physiol Regul Integr Comp Physiol 296:R1427-38
Blaustein, Mordecai P; Zhang, Jin; Chen, Ling et al. (2009) The pump, the exchanger, and endogenous ouabain: signaling mechanisms that link salt retention to hypertension. Hypertension 53:291-8
Radzyukevich, T L; Lingrel, J B; Heiny, J A (2009) The cardiac glycoside binding site on the Na,K-ATPase alpha2 isoform plays a role in the dynamic regulation of active transport in skeletal muscle. Proc Natl Acad Sci U S A 106:2565-70
Wansapura, Arshani N; Lasko, Valerie; Xie, Zijian et al. (2009) Marinobufagenin enhances cardiac contractility in mice with ouabain-sensitive alpha1 Na+-K+-ATPase. Am J Physiol Heart Circ Physiol 296:H1833-9
Lorenz, John N; Loreaux, Elizabeth L; Dostanic-Larson, Iva et al. (2008) ACTH-induced hypertension is dependent on the ouabain-binding site of the alpha2-Na+-K+-ATPase subunit. Am J Physiol Heart Circ Physiol 295:H273-80
Loreaux, Elizabeth L; Kaul, Baksho; Lorenz, John N et al. (2008) Ouabain-Sensitive alpha1 Na,K-ATPase enhances natriuretic response to saline load. J Am Soc Nephrol 19:1947-54

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