The sodium pump and hypertension: A physiological role for the cardiac glycoside binding site of the Na,K-ATPase

The Na,K-ATPase (or Na pump) is an integral membrane protein that transports Na⁺ and K⁺ across the plasma membrane of almost all animal cells and couples this work to the hydrolysis of the terminal phosphate bond of ATP (1). A significant fraction (up to ≈30%) of the ATP generated by cell metabolism is dedicated to this active transport process. The electrical gradient created by the Na pump is essential for the excitable activity of muscle and nerve tissue, and the inwardly directed Na gradient maintained by the Na pump is used in most cells and organs to drive the uptake and accumulation of a wide range of essential nutrients and cellular substrates and to reduce cell calcium and proton concentrations.

The Na,K-ATPase also serves as the unique binding site for cardiac glycosides, such as ouabain, digoxin, and digitoxin. Plant extracts containing cardiac glycosides have been used therapeutically since the 18th century. The cardiac glycoside site is the pharmacological target for digoxin, which is currently used widely in the treatment of congestive heart failure. However, the strong evolutionary conservation of this binding site among almost all species has suggested that important physiological functions may involve recognition of this site by endogenous agents. The work of Dostanic-Larson et al. in this issue of PNAS (2) demonstrates that the cardiac glycoside binding site of Na,K-ATPase plays an important physiological role in blood pressure regulation. It is reported that ACTH-induced hypertension is abolished when the interaction of cardiac glycosides with the Na,K-ATPase is disrupted. The interaction was abolished by developing genetically engineered knock-in mice in which two amino acids (located in the first extracellular domain of the Na,K-ATPase α-subunit) that are critical for ouabain sensitivity were replaced (3, 4). By using this approach, the normally ouabain-sensitive α2 isoform of the Na,K-ATPase was rendered resistant, whereas the normally insensitive α1 isoform was made ouabain-responsive (5, 6). Although the amino acid substitutions altered the sensitivity to cardiac glycosides of the α1 and α2 isoforms, they did not affect the expression of these two isoforms or the total Na,K-ATPase activity in tissues from the knock-in mice. It was noted that chronic administration of ACTH, which caused hypertension in wild-type mice, did not have such an effect in mice in which the α2 isoform was made resistant to cardiac glycosides. In an observation that completes the circle, knock-in animals in which the normally ouabain-resistant α1 isoform was made sensitive to cardiac glycosides and the sensitive α2 isoform was made resistant also developed ACTH-induced hypertension. These results provide a clear demonstration that an endogenous ligand must exist that mediates its action by binding to the Na,K-ATPase and is one of the regulators of blood pressure.

There has been a considerable controversy during the last two decades over the existence, identity, and physiological significance of a natural ligand that might interact with the cardiac glycoside binding site on the Na,K-ATPase. The identification of cardiac glycoside-like molecules in mammals has led to the proposal that these compounds are natural regulators of Na,K-ATPase in vivo (7–10). Several studies have shown a correlation between these cardiac glycoside-like molecules and certain pathological conditions, such as hypertension and heart failure (11–17). Plasma levels of these putative ligands for the Na,K-ATPase are high in several animal hypertension models, as well as in human essential hypertension and preeclampsia (11–14, 18, 19). In addition, marked increases in peripheral and central endogenous ouabain-like compounds occur in animal models of congestive heart failure as well as in human patients (15–17). However, it is still difficult to draw definitive conclusions regarding the specific function of these molecules in pathophysiology. Although the studies by Dostanic-Larson et al. (2) strongly support the suggestion that endogenous cardiac glycoside-like compounds are the naturally occurring ligand, they do not eliminate the possibility that other steroid-like (or, indeed, nonsteroidal) molecules, either previously identified or novel, may play this functional role. Nevertheless, this important work does clearly demonstrate that a naturally occurring regulator must exist and that its interaction with the cardiac glycoside binding site of the Na,K-ATPase plays a role in blood pressure regulation.

According to the National Health and Nutrition Examination Survey (NHANES) at least 50 million Americans have elevated blood pressure and ≈7.1 million annual deaths can be attributed to hypertension (20). The individual and societal costs of the prevalence of this condition are enormous. The present work brings a new dimension to an already complex physiological system of blood pressure regulation. Recently, the same authors have provided evidence that the Na,K-ATPase is involved in blood pressure regulation by demonstrating that inhibition of this enzyme, specifically of the α2 isoform, by ouabain leads to hypertension (21). In this issue of PNAS, these authors go beyond this finding and show that the cardiac glycoside binding site of the Na,K-ATPase modulates the development of hypertension. What we have so far learned from the work by Dostanic-Larson and coworkers (2) is that ACTH-induced hypertension in mice, which is a model system for hypertension associated with the ectopic ACTH and Cushing syndrome in humans, is due to modulation of Na,K-ATPase via the cardiac glycoside binding site (22–23). According to the model presented in Fig. 1, high levels of ACTH, which result from adenomas or an over-active adrenal gland, lead to elevation of a naturally occurring ligand for the Na,K-ATPase. Interaction of this ligand with Na,K-ATPase by means of the cardiac glycoside binding site results in hypertension.

Fig. 1.

Model for ACTH-induced hypertension by means of the cardiac glycoside binding site of the Na,K-ATPase. Chronic and excessive circulation of ACTH cased by adenoma or an overproductive adrenal gland results in the continuous elevation of a natural ligand for the Na,K-ATPase. Interaction of this ligand with the high-affinity cardiac glycoside binding site of the Na,K-ATPase, specifically an ouabain-sensitive α1 or an ouabain-sensitive α2 isoform, results in hypertension. The mechanism of how this interaction increases blood pressure is still unknown and could involve alteration of intracellular ion concentrations and/or induction of signaling pathway, such as Src/PKC/MAPKK.

We now need to understand how ACTH induces the elevation of the natural ligand. In addition, it is imperative to determine what the natural ligand for Na,K-ATPase is and whether its interaction with the cardiac glycoside binding site mediates other forms of hypertension. Another intriguing question is how interaction of this natural ligand with Na,K-ATPase causes ACTH-induced hypertension. A number of possible mechanisms can be proposed. The inhibition of Na,K-ATPase may change a local intracellular ion concentration, specifically of Na⁺ and Ca²⁺, or binding of the ligand may initiate a complex intracellular signaling cascade, any of which could result in elevation of blood pressure (24, 25). However, by using the elegant experimental approach employing cardiac glycoside-resistant or -sensitive knock-in mice, most of these questions can now be addressed.

It is both instructional and important to appreciate that the approach used by the authors of this study relied on the results of their own basic research on structure–function correlates in an ion transporter. These findings were then employed in the context of an animal model system to shed light on the mechanism of an exceptionally important disease state in humans. This logical sequence of events required both the basic studies and their “translation” to a disease-relevant context.