[Porcine kidney extract contains a specific inhibitor of the ouabain-sensitive alpha2-isoform of Na, K-ATPase present in rat diaphragm fibres].
In experiments on isolated rat diaphragm muscle, acetylcholine (100 nmol/l) hyperpolarized muscle fibres due to activation of the alpha 2 isoform of Na,K-ATPase. This hyperpolarization was blocked in a dose-dependent manner by ouabain (K0.5 = 8 +/- 4 nmol/l) as well as by a solution of porcine kidney extract (10 kDa cut-off filtration), with the K0.5 approximately equal to a 1:20,000-fold dilution. The inhibitory activity of the developed slowly over a period of 3 hours and, in contrast to ouabain, was still present after 1 hour of washing. Ouabain, but not the extract, inhibits Rb+ uptake in human erythrocytes that only express the alpha = 1 isoform of Na, K-ATPase. Our data suggest that in rat skeletal muscle the alpha 1 isoform of Na,K-ATPase is primarily responsible for ionic homeostasis, while the alpha 2 isoform provides a "regulatable" function and may be controlled by cholinergic stimulation and/or endogenous digitalis-like factors (EDLFs). Porcine kidney extract contains a factor (M. W. < 10 kDa) that selectively inhibits the rat alpha 2 isoform and differs from ouabain. Our experimental protocol can be used as a highly sensitive physiological assay for factors that selectively inhibit the alpha 2 isoform of Na,K-ATPase.
Krivoĭ
I I
II
St. Petersburg State University, 199034, St. Petersburg, 7/9 Universitetskaya Nab., Russia.
Vasil'ev
A N
AN
Gromova
V V
VV
Prytkov
A E
AE
Marakhova
I I
II
Kravtsova
V V
VV
Dobretsov
M G
MG
Mandel
F
F
D000818:Animals; D003964:Diaphragm / Q000187:drug effects / Q000201:enzymology / Q000502:physiology; D004791:Enzyme Inhibitors / Q000302:isolation & purification / Q000494:pharmacology*; D066298:In Vitro Techniques; D007668:Kidney / Q000737:chemistry*; D008297:Male; D008564:Membrane Potentials / Q000187:drug effects; D018482:Muscle, Skeletal / Q000187:drug effects / Q000201:enzymology* / Q000502:physiology; D010042:Ouabain / Q000494:pharmacology*; D051381:Rats; D000254:Sodium-Potassium-Exchanging ATPase / Q000037:antagonists & inhibitors*; D013552:Swine; D014020:Tissue Extracts / Q000302:isolation & purification / Q000494:pharmacology*
D004791:Enzyme Inhibitors; D014020:Tissue Extracts; D010042:Ouabain; C498532:Atp1a2 protein, rat; D000254:Sodium-Potassium-Exchanging ATPase