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Molecular Mechanisms Linking Salt to Hypertension

Physiological role of the ₁- and ₂-isoforms of the Na⁺-K⁺-ATPase and biological significance of their cardiac glycoside binding site

¹Departments of Molecular Genetics, Biochemistry and Microbiology, and ²Molecular and Cellular Physiology, College of Medicine, University of Cincinnati, Cincinnati, Ohio; ³University of Ottawa Heart Institute, Hypertension Unit, Ottawa; and ⁴Departments of Medicine and Biochemistry, Microbiology, and Immunology, University of Ottawa, Ottawa, Ontario, Canada

	ABSTRACT

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An interesting feature of Na⁺-K⁺-ATPase is that it contains four isoforms of the catalytic -subunit, each with a tissue-specific distribution. Our laboratory has used gene targeting to define the functional role of the ₁- and ₂-isoforms. While knockout mice demonstrated the importance of the ₁- and ₂-isoforms for survival, the knockin mice, in which each isoform can be individually inhibited by ouabain and its function determined, demonstrated that both isoforms are regulators of cardiac muscle contractility. Another intriguing aspect of the Na⁺-K⁺-ATPase is that it contains a binding site for cardiac glycosides, such as digoxin. Conservation of this site suggests that it may have an in vivo role and that a natural ligand must exist to interact with this site. In fact, cardiac glycoside-like compounds have been observed in mammals. Our recent study demonstrates that the cardiac glycoside binding site of the Na⁺-K⁺-ATPase plays a role in the regulation of blood pressure and that it mediates both ouabain-induced and ACTH-induced hypertension in mice. Whereas chronic administration of ouabain or ACTH caused hypertension in wild-type mice, it had no effect on blood pressure in mice with a ouabain-resistant ₂-isoform of Na⁺-K⁺-ATPase. Interestingly, animals with the ouabain-sensitive ₁-isoform and a ouabain-resistant ₂-isoform develop ACTH-induced hypertension to a greater extent than wild-type animals. Taken together, these results demonstrate that the cardiac glycoside binding of the Na⁺-K⁺-ATPase has a physiological role and suggests a function for a naturally occurring ligand that is stimulated by administration of ACTH.

ouabain; adrenocorticotropic hormone; blood pressure regulation; cardiotonic steroids

The Na⁺-K⁺-ATPase is a member of the P-type family of ATPases and is closely related to the Ca²⁺-ATPase family and the H⁺-K⁺-ATPase (31, 36). Na⁺-K⁺-ATPase is composed of two major subunits, the catalytic - and glycosylated -subunit. However, other proteins, such as members of the FXYD family of proteins, interact with this enzyme in some tissues, such as heart, kidney, and brain (20, 49, 50, 52). The association of these proteins modulates cation binding affinity of the Na⁺-K⁺-ATPase (1, 6, 11, 57). Three isoforms of the -subunit exist: ₁, ₂, and ₃. The -subunit has four isoforms, ₁, ₂, ₃, and ₄. While the ₁-isoform is expressed ubiquitously, the ₂-isoform is present largely in skeletal muscle, heart, brain, adipocytes, vascular smooth muscle, and eye, as well as a number of other tissues. The ₃-isoform is found almost exclusively in neurons and ovaries, but also occurs in white blood cells and heart of some species, such as humans (46, 48). The ₄-isoform is expressed in sperm and is specifically synthesized at the spermatagonia stage, where it is required for sperm motility (47, 54). It is reasonable to believe that the tissue-specific distribution of the -isoforms indicates that each isoform exhibits a particular function associated with the tissue in which it is expressed.

Our initial studies directed toward determining the role of the ₁- and ₂-isoforms used animals in which the genes coding for these isoforms were knocked out. Animals lacking the expression of the ₁-isoform died during embryogenesis. Specifically, embryos failed to develop beyond the blastocyst stage (2, 28). This is expected as the ₁-isoform is ubiquitously expressed and is required for multiple biological functions. Animals lacking the ₂-isoform gene are born but die immediately following birth (28). Subsequent studies by both our laboratory (41) and Ikeda et al. (27) demonstrated that ₂-isoform knockout mice have a defect in the breathing center of the brain, causing a failure to breathe and, thereby, death from asphyxia.

Role of the ₁- and ₂-Isoforms in Cardiac Contraction: Gene Knockout Studies

In contrast to animals lacking both copies of the ₁- or ₂-isoform, animals lacking only one copy of either gene survive and appear normal. The ₁^+/– animals develop normally with no observable phenotype (28). The functional role of the ₁- and the ₂-isoforms in cardiac and skeletal muscle have been studied. In both cases, the particular isoform is reduced by 40% compared with wild-type, and both genotypes exhibit altered cardiac and skeletal muscle contractile functions (24, 28).

Whereas mice lacking one copy of the ₂-isoform gene exhibit an increase in the force of cardiac and skeletal muscle contraction, animals lacking one copy of the ₁-isoform have decreased contraction in both cardiac and skeletal muscle (24, 28). These different phenotypes were unexpected and suggest that the ₁- and ₂-isoforms play different roles in the regulation of muscle contractility. The increase in contractile force of the ₂-isoform heterozygous knockout mice fit with the proposed model that inhibition of Na⁺-K⁺-ATPase activity results in an increase in intracellular Na⁺ levels, which, in turn, causes an increase in Ca²⁺ through the reverse mode of the Na/Ca exchanger. However, the hypocontractility in the ₁-isoform heterozygous knockout mice is opposite of the proposed model. Subsequent studies showed that the hypocontractility in ₁^+/– hearts is not the result of calcium overload, which could have been responsible for the phenotype (42).

To help address why the ₁-haplo-insufficient animals exhibit unexpected cardiac hypocontractility, mRNA microarray analysis was carried out on heart tissue from wild-type and ₁^+/– animals (40). Interestingly, the expression of a number of genes is altered, and these could be grouped into various clusters, including metabolic and ion transport pathways. The expression of a number of genes that are involved in enhancing cardiac contractility, such as atrial natriuretic peptide, are increased. Furthermore, the changes in gene expression in heart provided evidence of altered kidney and adrenal gland function. These indicate that multiple organs in these animals are compromised with the loss of one copy of the ₁-isoform, and this raised the possibility that the reduced cardiac contractility in ₁-heterozygous mice may be due, in part, to compensatory changes that accumulate during development. Therefore, it may be difficult to draw conclusions about ₁-isoform function from animals lacking one copy of the ₁-isoform gene.

Role of the ₁- and ₂-Isoforms in Cardiac Function: Isoform Specific Pharmacological Inhibition

Whereas the ablation and reduction of the ₁- or ₂-isoforms show the importance of these isoforms, they failed to definitively demonstrate a physiological function for them. Therefore, a different approach was used to examine the specific function of the ₁- and ₂-isoforms of the Na⁺-K⁺-ATPase.

An approach for investigating the role of each -isoform, without the long-term compensation that occurs during development in knockout animals, is to develop animals where only one isoform can be pharmacologically inhibited at a time and analyze the physiological consequence of this inhibition. All -isoforms of the Na⁺-K⁺-ATPase in most species, except for a few, such as the rat and mouse ₁-isoform, are inhibited by low concentrations of ouabain. We took advantage of our earlier studies defining the amino acids conferring differential sensitivity to ouabain. The basis for the relative sensitivity of the mouse and rat ₁-isoforms to ouabain resides in two amino acids in the first extracellular region of the -subunit (44). The mouse ₂-isoform, which is sensitive to ouabain, has a leucine at position 111 and an asparagine occurs at position 122. This is in contrast to the ₁-isoform, which has low affinty for ouabain, where arginine and aspartic acid appear at these sites. The positive charge of the latter two amino acids appears to confer resistance to ouabain (44). By using the mice with the combination of sensitive and insensitive isoforms, we can inhibit only one isoform by ouabain without altering the activity of other -isoforms.

With the use of a Cre-loxP gene-targeting strategy, mice were developed where the leucine and asparagine of the ₂-isoform gene were converted to arginine and aspartic acid, i.e., L111R and N122D (15). These animals express a portion of the -wild-type gene replaced with a sequence conferring ouabain resistance and are referred to as knockin mice.

The resulting knockin animals show no difference in the levels of expression and tissue distribution of the ₁-, ₂-, or ₃-isoforms (15). However, the muscle and heart of the ₂-ouabain-insensitive animals exhibit a loss of ouabain binding, as expected, because in wild-type mice, both tissues have a sensitive ₂-isoform in addition to the relatively insensitive ₁-isoform. This loss of ouabain binding indicates that the targeted ₂-isoform is successfully modified to a ouabain-resistant isoform. The gene-targeted knockin animals are born in normal Mendelian ratios and show no outward phenotype. Furthermore, their baseline cardiovascular hemodynamics are normal (15). The animals also respond normally to the -adenergic agonist dobutamine. Therefore, the conversion of the ₂-isoform to a ouabain-resistant protein does not alter essential properties of the Na⁺-K⁺-ATPase.

The analysis of cardiac contractility in intact mice and in ex vivo isolated heart preparations demonstrates that ouabain, at concentrations that inhibit the sensitive ₂-isoform in wild-type mice, fail to induce positive cardiac inotropy in knockin hearts. This is in contrast to the increased cardiac contractility observed in the wild-type heart following administration of the same concentrations of ouabain. This clearly indicates that the ₂-isoform is responsible for the ouabain-induced positive cardiac inotropy and demonstrates the importance of the ₂-isoform in the regulation of cardiac contractility (15). However, these studies do not provide information on the role of the ₁-isoform in these processes.

The role of the ₁-isoform in cardiac contractility was determined by developing animals with a ouabain-sensitive ₁-isoform (in rodents, the ₁-isoform is naturally ouabain insensitive) and a resistant ₂-isoform (16). The ouabain sensitivity of the ₁-isoform was conferred by introducing R111Q and D122N amino acid substitutions. The ouabain-sensitive ₁-isoform mice develop normally and show no visible phenotype. The expression and tissue distribution of each -isoform is also not altered by modification of the ₁-isoform. However, the ouabain sensitivity of the Na⁺-K⁺-ATPase in tissues, and specifically in kidney, where wild-type animals express only a ouabain-resistant ₁-isoform, is enhanced as expected in the gene-targeted animals. These animals were then mated to those with a ouabain-resistant ₂-isoform to give animals with the genotype 1^S/S 2^R/R, where S and R indicate sensitive and resistant to ouabain, respectively. In these animals only the ₁-isoform is inhibited by low concentrations of ouabain and the function of this isoform in cardiac contractility can be determined without inhibiting the ₂-isoform.

When ouabain is administered to ₁^S/S ₂^R/R-mice, a very large increase in cardiac contractility occurs (16). This indicates that the ₁-isoform, similar to the ₂-isoform when inhibited by ouabain, elicits an increase in cardiac contraction. It should be pointed out that the magnitude of the increase is much greater when ₁ is inhibited compared with inhibiting the ₂-isoform, which is likely because the ₁-isoform is more abundant than the ₂-isoform in the mouse heart.

Whereas these studies suggest that both ₁- and ₂-isoforms play a similar role in cardiac contractility, they do not address the mechanism by which these two isoforms regulate cardiac function. It has been postulated that the Na⁺-K⁺-ATPase, particularly the ₂-isoform, modulates Ca²⁺ levels during contraction through functional coupling with Na/Ca exchanger (28, 29, 51). Thus, we tested whether the ₁- or ₂-isoform functionally couples and physically interacts with the Na/Ca exchanger (16). Pretreatment with KB-R7943, an inhibitor of the reverse mode of the Na/Ca exchanger, abolished the cardiotonic effects of ouabain in isolated wild-type and the ouabain-sensitive ₁-isoform hearts. Also, immunoprecipitation analyses demonstrated that both the ₁- and ₂-isoforms colocalize with the Na/Ca exchanger in the heart. These studies clearly demonstrate that both the ₁- and ₂-isoforms regulate cardiac contraction through the functional and physical coupling with the Na/Ca exchanger. In summary, both isoforms colocalize with the Na/Ca exchanger, and their inhibition by ouabain leads to enhanced contractility.

Physiological Role of the ₂-Isoform in Blood Pressure Regulation

The importance of the ₂-isoform in regulation of cardiac contractility and the fact that the cardiac output and vascular resistance regulate blood pressure, led us to investigate the physiological role of the ₂-isoform in the regulation of blood pressure (17). Both wild-type and ouabain-resistant ₂-isoform mice were administered ouabain (once a day by intraperitoneal injection) over an extended period of time, and their blood pressure was measured daily via tail cuff. Chronic administration of ouabain induced hypertension in wild-type but not in knockin mice, indicating that the ₂-isoform is involved in the blood pressure response to ouabain. In addition, acute administration of ouabain increased blood pressure in wild-type but not in ₁^R/R ₂^R/R-mice. This shows that the ₂-isoform plays a role in maintenance of the overall cardiovascular hemodynamics and is further supported by data demonstrating that heterozygous ₂-isoform knockout mice are hypertensive (56).

Physiological Role of the Cardiac Glycoside Binding Site

One of the interesting features of the Na⁺-K⁺-ATPase is that it possesses a binding site for cardiac glycosides, which have been used for centuries in treatment of congestive heart failure. The observation that the cardiac glycoside binding site of the Na⁺-K⁺-ATPase is conserved through evolution, from hydra to human (10), suggests that this site may play an important biological role. The finding of endogenous cardiac glycoside-like compounds in mammals further strengthens this hypothesis (4, 8, 9, 14, 21–23, 37–39, 43, 45). Although the exact number and structure of endogenous cardiac glycosides are still unknown, elevated levels have been correlated with an increase in blood pressure (4, 13, 21, 22, 26, 32, 37–39, 53) and development of congestive heart failure (3, 25).

Our approach in determining whether the ouabain binding site plays a biological role, and, thereby, testing the physiological relevance of endogenous ligands, was to utilize the knockin mice described above in which the cardiac glycoside binding site of the ₁- and ₂-isoforms is altered. As indicated earlier, ₁^R/R ₂^R/R-animals show no observable phenotype under normal laboratory conditions nor do animals with a sensitive ₁-isoform. These animals are born in normal Mendelian ratios, their weight gain is normal, litter sizes are normal, and their life span is unchanged from wild-type animals. On the surface, these studies suggest that the ouabain binding site does not play a significant biological role under normal conditions. However, endogenous ouabain levels are present at very low concentrations under resting conditions but increase under stress, such as exercise (4) and ACTH administration (19, 21, 34, 35, 55) or a high-salt diet (26, 53). Therefore, animals with an ouabain-resistant ₂-isoformn i.e., ₁^R/R ₂^R/R, were compared with wild-type animals (₁^R/R ₂^S/S) following daily administration of ACTH (18). In wild-type animals, blood pressure began to increase on the second day of injection and plateaued at about 5 days. In animals that have the resistant ₂-isoform, this increase in blood pressure does not occur, indicating that ACTH-induced hypertension is mediated through the ouabain binding site of ₂-isoform of Na⁺-K⁺-ATPase. This suggests a role for this binding site, at least in this hypertensive condition, and further implicates the presence of an endogenous ligand that would interact with this site. During ACTH-induced hypertension, there is an increase in endogenous ouabain-like compounds, suggesting involvement of these compounds (18). Furthermore, it has been shown that adrenocortical cells secrete ouabain-like material following stimulation by ACTH (5, 33).

In humans, both the ₁- and ₂-isoforms are sensitive to ouabain, and it was of interest to determine whether the ouabain binding site of the ₁-isoform could also play a role in ACTH-induced hypertension in mice. There is a robust increase in blood pressure associated with administration of ACTH in ₁^S/S ₂^R/R, in contrast to no increase in blood pressure in ₁^R/R ₂^R/R-animals. This indicates that the ₁-isoform can also mediate ACTH-induced hypertension and suggests that in humans, where the ₁-isoform is sensitive to cardiac glycosides, regulation of the Na⁺-K⁺-ATPase through the ₁-isoform could also have physiological significance in the regulation of blood pressure.

Although these studies do not identify the ligand that is interacting with the Na⁺-K⁺-ATPase, they do demonstrate that the cardiac glycoside binding site is important and that an endogenous ligand must exist (Fig. 1). It is likely that the cardiac glycoside binding site of this enzyme will be found to play an important role in a number of biological functions.

View larger version (17K): [in this window] [in a new window]   Fig. 1. Ouabain-sensitive Na+-K+-ATPase mediates ACTH-induced hypertension. Chronic administration/exposure to ACTH releases naturally occurring ligand(s) for the Na+-K+-ATPase. Ouabain-sensitivity of the Na,K-ATPase determines its reactivity towards this endogenous ligand. The ouabain-sensitive Na+-K+-ATPase binds the ligand via the cardiac glycoside binding site. This interaction results in the development of hypertension. In contrast, ouabain-resistant Na+-K+-ATPase cannot interact with the endogenous ligand, and ACTH-induced hypertension does not occur. Q, glutamine; N, asparagine; R, arginine; and D, aspartic acid.

	ACKNOWLEDGMENTS

 
This research was supported by the National Institutes of Health Grants HL-28573 and HL-66062 (to J. B. Lingrel), and DK-57552 (to J. N. Lorenz), the Heart and Stroke Foundation of Ontario Grant NA-5102 and Canadian Institute of Health Research Grant 74572 (to J. W. Van Huysse).

	FOOTNOTES

 

Address for reprint requests and other correspondence: J. B Lingrel, Dept. of Molecular Genetics, Biochemistry and Microbiology, Univ. of Cincinnati College of Medicine, PO Box 670524, 231 Albert Sabin Way, Cincinnati, OH 45267-0524 (e-mail: Jerry.Lingrel{at}uc.edu)

	REFERENCES

TOP ABSTRACT REFERENCES

 

1. Arystarkhova E, Wetzel RK, Asinovski NK, and Sweadner KJ. The gamma subunit modulate Na(+) and K(+) affinity of the renal Na,K-ATPase. J Biol Chem 274: 33183–33185, 1999.[Abstract/Free Full Text]
2. Barcroft LC, Moseley AE, Lingrel JB, and Watson AJ. Deletion of the Na/K-ATPase alpha 1-subunit gene (Atp1a1) does not prevent cavitation of the preimplantation mouse embryo. Mech Dev 121: 417–426, 2004.[Web of Science][Medline]
3. Bagrov AY, Fedorova OV, Maslova MN, Roukoyatkina NI, Ukhanova MV, and Zhabko EP. Endogenous plasma Na,K-ATPase inhibitory activity and digoxin-like immunoreactivity after acute myocardial infarction. Cardiovasc Res 25: 371–377, 1991.[Web of Science][Medline]
4. Bauer N, Muller-Ehmsen J, Kramer U, Hambarchian N, Zobel C, Schwinger RH, Neu H, Kirch U, Grumbaun EG, and Schoner W. Ouabain-like compound changes rapidly on physical exercise in humans and dogs: effects of beta-blockade and angiotensin-converting enzyme inhibition. Hypertension 45: 1024–1028, 2005.[Abstract/Free Full Text]
5. Beck M, Szalay KS, Nagy GM, Toth M, and de Chatel R. Production of ouabain by rat adrenocortical cells. Endocr Res 22: 845–849, 1996.[Web of Science][Medline]
6. Beguin P, Cambert G, Guennoun S, Garty H, Horisberger JD, and Geering K. CHIF, A member of the FXYD protein family, is a regulator of Na,K-ATPase distinct from the gamma-subunit. EMBO J 20: 3993–4002, 2001.[CrossRef][Web of Science][Medline]
7. Blanco G, Sanchez G, Melton RJ, Tourtellotte WG, and Mercer RW. The alpha 4 isoform of the Na,K-ATPase is expressed in the germ cells of the testes. J Histochem Cytochem 48: 1023–1032, 2000.[Abstract/Free Full Text]
8. Blaustein MP. Physiological effects of endogenous ouabain: Control of intracellular Ca²⁺ stores and cell responsiveness. Am J Physiol Cell Physiol 264: C1367–C1387, 1993.[Abstract/Free Full Text]
9. Blaustein MP, Juhaszova M, and Golovina VA. The cellular mechanism of action of cardiotonic steroids: a new hypothesis. Clin Exp Hypertens 20: 691–703, 1998.[CrossRef][Web of Science][Medline]
10. Candield VA, Xu KY, D’Aquila T, Shyjan AW, and Levenson R. Molecular cloning and characterization of Na,K-ATPase from hydra vulgaris: implications for enzyme evolution and ouabain sensitivity. New Biol 4: 339–348, 1992.[Web of Science][Medline]
11. Crambert G, Fuzesi M, Garty H, Karlish S, and Geering K. Phospholemman (FXYD1) associates with Na,K-ATPase and regulates its transport properties. Proc Natl Acad Sci USA 99: 11476–11481, 2002.[Abstract/Free Full Text]
12. Crambert G, Li C, Claeys D, and Geering K. FXYD3 (Mat-8), a new regulator of Na,K-ATPase. Mol Biol Cell 16: 2363–2371, 2005.[Abstract/Free Full Text]
13. Delva P, Capra C, Degan M, Minuz P, Covi G, Milan L, Steele A, and Lechi A. High plasma levels of ouabain-like factor in normal pregnancy and in pre-eclampsia. Eur J Clin Invest 19: 95–100, 1989.[Web of Science][Medline]
14. Doris PA. Regulation of Na,K-ATPase by endogenous ouabain-like materials. Proc Soc Exp Biol Med 205: 202–212, 1994.[CrossRef][Medline]
15. Dostanic I, Lorenz JN, Schultz JEJ, Grupp IL, Neumann JC, Wani MA, and Lingrel J. B The 2 isoform of Na,K-ATPase mediates ouabain-induced cardiac inotropy in mice. J Biol Chem 276: 53026–53034, 2003.
16. Dostanic I, Schultz Jl. J, Lorenz JN, and Lingrel J. B The 1 Isoform of Na,K-ATPase regulates cardiac contractility and functionally interacts and co-localizes with the Na/Ca exchanger in heart. J Biol Chem 279: 54053–54061, 2004.[Abstract/Free Full Text]
17. Dostanic I, Paul RJ, Lorenz JN, Theriault S, Van Huysse JW, and Lingrel JB. The ₂-isoform of Na⁺-K⁺-ATPase mediates ouabain-induced hypertension in mice and increased vascular contractility in vitro. Am J Physiol Heart Circ Physiol 288: H477–H485, 2005.[Abstract/Free Full Text]
18. Dostanic-Larson I, Van Huysse JW, Lorenz JN, and Lingrel JB. The highly conserved cardiac glycoside binding site of Na,K-ATPase plays a role in blood pressure regulation. Proc Natl Acad Sci USA 102: 15845–15850, 2005.[Abstract/Free Full Text]
19. Fedorova OV, Anderson DE, and Bagrov AY. Plasma marinobufagenin-like and ouabain-like immunoreactivity in adrenocorticotropin-treated rats. Am J Hypertens 11: 796–802, 1998.[CrossRef][Web of Science][Medline]
20. Feschenko MS, Donnet C, Wetzel RK, Asinovski NK, Jones LR, and Sweadner KJ. Phospholemman, a single-span membrane protein, is an accessory protein of Na,K-ATPase in cerebellum and choroid plexus. J Neurosci 23: 2161–2169, 2003.[Abstract/Free Full Text]
21. Goto A, Yamada K, Hazama H, Uehara Y, Atarashi K, Hirata Y, Kimura K, and Omata M. Ouabain-like compound in hypertension associated with ectopic corticotropin syndrome. Hypertension 28: 421–425, 1996.[Abstract/Free Full Text]
22. Hamlyn JM, Ringel R, Schaeffer J, Levinson PD, Hamilton BP, Kowarski AA, and Blaustein MP. A circulating inhibitor of (Na⁺, K⁺) ATPase associated with essential hypertension. Nature 300: 650–652, 1982.[CrossRef][Medline]
23. Hamlyn JM, Blaustein MP, Bova S, DuCharme DW, Harris DW, Mandel F, Mathews WR, and Ludens JH. Identification and characterization of a ouabain-like compound from human plasma. Proc Natl Acad Sci USA 88: 6259–6263, 1991.[Abstract/Free Full Text]
24. He S, Shelly DA, Moseley AE, James PF, James JH, Paul RJ, and Lingrel J. B The ₁- and ₂-isoforms of Na⁺-K⁺-ATPase play different roles in skeletal muscle contractility. Am J Physiol Regul Integr Comp Physiol 281: R917–R925, 2001.[Abstract/Free Full Text]
25. Huang BS, Yuan B, and Leenen FH. Chronic blockade of brain "ouabain" prevents sympathetic hyper-reactivity and impairment of acute baroreflex resetting in rats with congestive heart failure. Can J Physiol Pharmacol 78: 45–53, 2000.[CrossRef][Web of Science][Medline]
26. Huang BS and Leenen FH. Brain "ouabain" mediates the sympathoexcitatory and hypertensive effects of high sodium intake in Dahl salt-sensitive rats. Circ Res 74:586–95, 1994.[Abstract/Free Full Text]
27. Ikeda K, Onimaru H, Yamada J, Inoue K, Ueno S, Onaka T, Toyoda H, Arata A, Ishikawa T, Taketo MM, Fukuda A, and Kawaakami K. Malfunction of respiratory-related neuronal activity in Na,K-ATPase 2 subunit-deficient mice is attributable to abnormal C1-homeostasis in brainstem neurons. J Neurosci 24: 10693–10701, 2004.[Abstract/Free Full Text]
28. James PF, Grupp IL, Grupp G, Woo AL, Askew GR, Croyle ML, Walsh RA, and Lingrel JB. Identification of a specific role for the Na,K-ATPase 2 isoform as a regulator of calcium in the heart. Mol Cell 3: 555–563, 1999.[CrossRef][Web of Science][Medline]
29. Juhaszova M and Blaustein MP. Na⁺ pump low and high ouabain affinity alpha subunit isoforms are differently distributed in cells. Proc Natl Acad Sci USA 94: 1800–1805, 1997.[Abstract/Free Full Text]
30. Juhaszova M, Shimizu ML, Borin RK, Santiago Yip EM, Lindenmayer GE, and Blaustein MP. Localization of the Na⁺-Ca²⁺ exchanger in vascular smooth muscle, and in neurons and astrocytes. Ann NY Acad Sci 779: 318–335, 1996.[Web of Science][Medline]
31. Kaplan JH. Biochemistry of Na,K-ATPase. Annu Rev Biochem 71:511–535, 2002.[CrossRef][Web of Science][Medline]
32. Kunes J, Stolba P, Pohlova I, Jelinek J, and Zicha J. The importance of endogenous digoxin-like factors in rats with various forms of experimental hypertension. Clin Exp Hypertens A 7: 707–720, 1985.[Web of Science][Medline]
33. Laredo J, Hamilton BP, and Hamlyn JM. Ouabain is secreted by bovine adrenocortical cells. Endocrinology 135: 794–797, 1994.[Abstract]
34. Li M, Wong KS, Martin A, and Whitworth JA. Adrenocorticotrophin-induced hypertension in rats. Role of progesterone and digoxin-like substances. Am J Hypertens 7: 59–68, 1994.[Web of Science][Medline]
35. Li M, Wen C, and Whitworth JA. Hemodynamic effects of the Fab fragment of digoxin antibody (Digibind) in corticotropic (ACTH)-induced hypertension. Am J Hypertens 10: 332–336, 1997.[CrossRef][Web of Science][Medline]
36. Lingrel JB and Kuntzweiler T. Na⁺,K⁺-ATPase. J Biol Chem 269: 19659–19662, 1994.[Free Full Text]
37. Manunta P, Stella P, Rivera R, Ciurlino D, Cusi D, Ferrandi M, Hamlyn JM, and Bianchi G. Left ventricular mass, stroke volume, and ouabain-like factor in essential hypertension. Hypertension 34: 450–456, 1999.[Abstract/Free Full Text]
38. Manunta P, Iacoviello M, Forleo C, Messaggio E, Hamlyn JM, Lucarelli K, Guida P, Romito R, De Tommasi E, Bianchi G, Rizzion P, and Pitzalis MV. High circulating levels of endogenous ouabain in the offspring of hypertensive and normotensive individuals. J Hypertens 23: 1677–1681, 2005.[Web of Science][Medline]
39. Manunta P, Messaggio E, Ballabeni C, Sciarrone MT, Lanzani C, Ferrandi M, Hamlyn JM, Cusi D, Galletti F, and Bianchi G. Plasma ouabain-like factor during acute and chronic changes in sodium balance in essential hypertension. Hypertension 38: 198–203, 2001.[Abstract/Free Full Text]
40. Moseley AE, Huddleson JP, Bohanan CS, James PF, Lorenz JN, Aronow BJ, and Lingrel JB. Genetic profiling reveals global changes in multiple biological pathways in the hearts of Na,K-ATPase alpha 1 isoform haploinsufficient mice. Cell Physiol Biochem 15: 145–158, 2005.[CrossRef][Web of Science][Medline]
41. Moseley AE, Lieske SP, Wetzel RK, James PF, He S, Shelly DA, Paul RJ, Boivin GP, Witte DP, Ramirez JM, Sweadner KJ, and Lingrel JB. The Na,K-ATPase 2 Isoform is Expressed in Neurons, and its Absence Disrupts Neuronal Activity in Newborn Mice. J Biol Chem 278: 5317–5324, 2003.[Abstract/Free Full Text]
42. Moseley AE, Cougnon MH, Grupp IL, Schultz JE, and Lingrel JB. Attenuation of cardiac contractility in Na,K-ATPase 1 isoform-deficient hearts under reduced calcium conditions. J Mol Cell Cardiol 37: 913–919, 2004.[CrossRef][Web of Science][Medline]
43. Parhami-Seren B, Haberly R, Margolies MN, Haupert GT Jr. Ouabain-binding protein(s) from human plasma. Hypertension 40: 220–228, 2002.[Abstract/Free Full Text]
44. Price EM and Lingrel JB. Structure-function relationships in the Na,K-ATPase alpha subunit: site directed mutagenesis of glutamine-111 to arginine and asparagines-122 to aspartic acid generates a ouabain-resistant enzyme. Biochemistry 27: 8400–8408, 1988.[CrossRef][Medline]
45. Schoner W. Endogenous cardiac glycosides, a new class of steroid hormones. Eur J Biochem 269: 2440–2448, 2002.[Web of Science][Medline]
46. Shamraj OI, Melvin D, and Lingrel JB. Expression of Na,K-ATPase isoforms in human heart. Biochem Biophys Res Commun 179: 1434–1440, 1991.[CrossRef][Web of Science][Medline]
47. Shamraj OI and Lingrel JB. A putative fourth Na,K-ATPase alpha-subunit gene is expressed in testis. Proc Natl Acad Sci USA 91: 12952–12956, 1994.[Abstract/Free Full Text]
48. Stengelin MK and Hoffman JF. Na,K-ATPase subunit isoforms in human reticulocytes: evidence from reverse transcription-PCR for the presence of alpha1, alpha3, beta2, beta3, and gamma. Proc Natl Acad Sci USA 94: 5943–5948, 1997.[Abstract/Free Full Text]
49. Sweadner KJ, Arystarkhova E, Donnet C, and Wetzel RK. FXYD proteins as regulators of the Na,K-ATPase in the kidney. Ann NY Acad Sci 986: 382–387, 2003.[Web of Science][Medline]
50. Sweadner KJ. Phospholemman: a new force in cardiac contractility. Circ Res 97: 510–511, 2005.[Free Full Text]
51. Terracciano CM. Rapid inhibition of the Na⁺-K⁺ pump affects Na⁺-Ca²⁺ exchanger-mediated relaxation in rabbit ventricular myocytes. J Physiol 533: 165–173, 2001.[Abstract/Free Full Text]
52. Therien AG, Goldshleger R, Karlish SJD, and Blostein R. Tissue-specific distribution and modulatory role of the subunit of the Na,K-ATPase. J Biol Chem 272: 32628–32634, 1997.[Abstract/Free Full Text]
53. Van Huysse JW and Hou X. Pressor response to CSF sodium in mice: mediation by a ouabain-like substance and renin-angiotensin system in the brain. Brain Res 1021: 219–231, 2004.[CrossRef][Web of Science][Medline]
54. Woo AL, James PF, and Lingrel JB. Sperm motility is dependent on a unique isoform of the Na,K-ATPase. J Biol Chem 275: 20693–20699, 2000.[Abstract/Free Full Text]
55. Yamada K, Goto A, and Onata M. Adrenocorticotropin-induced hypertension in rats. Am J Hypertens 10: 403–408, 1997.[Web of Science][Medline]
56. Zhang J, Lee MY, Cavalli M, Chen L, Berra-Romani R, Balke CW, Bianchi G, Ferrari P, Hamlyn JM, Iwamoto T, Lingrel JB, Matteson DR, Wier WG, and Blaustein MP. Sodium pump (alpha)2 subunits control myogenic tone and blood pressure in mice. J Physiol 569: 243–256, 2005.[Abstract/Free Full Text]
57. Zouzoulas Thereien A, AG, Scanzano R, Deber CM, and Blostein R. Modulation of Na,K-ATPase by the subunit. J Biol Chem 278: 40437–40441, 2003.[Abstract/Free Full Text]

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