Marinobufagenin, an endogenous α-1 sodium pump ligand, in hypertensive Dahl salt-sensitive rats

Dahl salt-sensitive rats (DS), which have a mutation in the alpha -1 subunit of Na+/K+-ATPase, exhibit impaired pressure natriuresis and on a high-salt diet, retain Na+ and exhibit increased blood pressure. Recently, we have shown that mammalian tissues contain a bufadienolide Na+/K+-ATPase inhibitory factor, marinobufagenin (MBG), that exhibits greater affinity for the alpha -1 than alpha -3 sodium pump isoform. The present study investigated the possible role of MBG in hypertension in DS on a high NaCl intake. Eight DS and 8 Dahl salt-resistant rats (DR) were placed on an 8% NaCl diet. Within 2 weeks, systolic blood pressure increased in DS (162+/-9 mm Hg at week 2 versus 110+/-2 mm Hg in baseline, P<0.01), and increased less in DR (124+/-3 mm Hg at week 2 versus 112+/-2 mm Hg in baseline). Renal excretion of MBG increased rt-fold (38.9+/-7.6 pmol versus 9.1+/-1.3 pmol in baseline, P<0.01) in DS, but by only 25% in DR (13.2+/-0.9 pmol versus 10.3+/-0.7 pmol in baseline). Excretion of endogenous ouabain did not change in either strain. MBG-immunoreactive material was purified from the urine of hypertensive DS by means of 2 steps of reverse-phase high performance liquid chromatography (HPLC) and compared with plant ouabain and amphibian MBG for its ability to inhibit the Na+/K+-ATPase from rat kidney (which expresses only alpha -1 Na+/K+-ATPase isoform). Unlike ouabain (IC50=248 mu mol/L), serially diluted, HPLC-purified MBG immunoreactivity from DS and authentic MBG potently inhibited rat kidney Na+/K+-ATPase (IC50=70 and 78 nmol/L, respectively). Our results suggest that an alpha -1 Na+/K+-ATPase ligand, MEG, is elaborated to promote natriuresis in hypertensive DS. MBG acts as a selective inhibitor of the ouabain-resistant alpha -1 Na+/K+-ATPase subunit, ie, the major sodium pump isoform of the kidneys, as would be expected of a putative natriuretic hormone.