Unsuitability of the 86Rb+ uptake method for estimation of (Na+ + K+)-ATPase activity in innervated tissues.
(Na+ + K+)-ATPase activity was estimated by 86Rb+ uptake in dog saphenous vein to determine the validity of the technique in tissues that have a sympathetic innervation. When saphenous vein rings were incubated at 37 degrees C in Krebs' solution containing 86Rb+, the cardenolide acetylstrophanthidin caused a concentration-dependent inhibition of Rb+ uptake. The threshold for inhibition was approx. 10 nM acetylstrophanthidin and the maximum effect was obtained at 9 microM. In the upper part of this concentration range (greater than 1 microM) acetylstrophanthidin released noradrenaline from the sympathetic nerve terminals associated with the tissue. In this upper part of the acetylstrophanthidin concentration range the alpha-adrenoceptor antagonist phentolamine (8 microM) reduced, by up to 25%, the degree of 86Rb+ uptake inhibition caused by the cardenolide. In other experiments, saphenous vein strips were loaded with 86Rb+ and perifused with Krebs' solution containing acetylstrophanthidin. At concentrations which release noradrenaline, acetylstrophanthidin increased the efflux of 86Rb+. Phentolamine (8 microM) prevented the acetylstrophanthidin-evoked efflux of the isotope as did prior in vitro denervation of 86Rb+ loaded strips with 6-hydroxydopamine. Exogenous noradrenaline (1-100 microM) added to the perifusing fluid also caused an efflux of 86Rb+ that was attenuated by phentolamine. The data indicate for dog saphenous vein that with low concentrations of acetylstrophanthidin the extent of 86Rb+ accumulation might accurately reflect prevailing (Na+ + K+)-ATPase activity. At higher concentrations of acetylstrophanthidin, however, noradrenaline is released from the nerve endings and causes 86Rb+ efflux from the smooth muscle cells consequent upon alpha-adrenoceptor activation. Since this efflux reduces the extent of Rb+ accumulation, measurement of the latter does not adequately reflect uptake mediated by the activity of (Na+ + K+)-ATPase. This is significant because in most applications of the 86Rb+ uptake method it is the estimate of Rb+ accumulation made in the presence of a high concentration of cardenolide that forms the basis of all subsequent calculations with respect to (Na+ + K+)-ATPase activity.
D000818:Animals; D004285:Dogs; D005188:False Negative Reactions; D006892:Hydroxydopamines / Q000494:pharmacology; D009131:Muscle, Smooth, Vascular / Q000187:drug effects / Q000294:innervation; D009638:Norepinephrine / Q000378:metabolism / Q000494:pharmacology; D016627:Oxidopamine; D010646:Phentolamine / Q000494:pharmacology; D011868:Radioisotopes / Q000378:metabolism; D011942:Receptors, Adrenergic, alpha / Q000187:drug effects; D012413:Rubidium / Q000378:metabolism; D012501:Saphenous Vein / Q000201:enzymology / Q000294:innervation; D000254:Sodium-Potassium-Exchanging ATPase / Q000032:analysis*; D013327:Strophanthidin / Q000031:analogs & derivatives / Q000494:pharmacology; D013564:Sympathetic Nervous System / Q000187:drug effects / Q000378:metabolism
D006892:Hydroxydopamines; D011868:Radioisotopes; D011942:Receptors, Adrenergic, alpha; D013327:Strophanthidin; D016627:Oxidopamine; D000254:Sodium-Potassium-Exchanging ATPase; C004681:acetylstrophanthidin; D012413:Rubidium; D009638:Norepinephrine; D010646:Phentolamine