EFFECTS OF 2 ENDOGENOUS NA+,K+-ATPASE INHIBITORS, MARINOBUFAGENIN AND OUABAIN, ON ISOLATED RAT AORTA

Previously, we reported that the venom of Bufo marinus toad contains a Na+,K+-ATPase inhibitor with potent vasoconstrictor activity. In the present study, using thin-layer chromatography in Silicagel 60 F-254+366, we separated a vasoactive substance from a mixture of steroids from Bufo marinus venom. Based on chromatographic mobility of this substance and typical color reaction after its vizualization with SbCl3, we identified it as a previously described steroid, marinobufagenin. Vasoconstrictor and Na+,K+ pump inhibitory properties of marinobufagenin were studied in isolated rat aortic rings and compared with those of ouabain. Ouabain (10-100 mu mol.l(-1)) produced weak vasoconstriction, which was blocked by 2 mu mol.l(-1) phentolamine. 10 mu mol.l(-1) ouabain stimulated, and at higher concentrations inhibited, the Na+,K+ pump. 2 mu mol.l(-1) phentolamine abolished the activating effect of 10 mu mol.l(-1) ouabain on the Na+,K+ pump, but did not alter the inhibitory action of higher concentrations of ouabain. By contrast, marunibufagenin elicited rapid and strong vasoconstriction and inhibited ouabain-sensitive Rb-86 uptake. Antidigoxin antibody antagonized the vasoconstrictor responses to marinobufagenin, but not to ouabain. 2 mu mol.l(-1) phentolamine did not alter the constrictor effect of marinobufagenin. In solid-phase digoxin immunoassay, marinobufagenin demonstrated higher digoxin-like immunoreactivity than ouabain.