K+-induced hyperpolarization in rat mesenteric artery:: identification, localization and role of Na+/K+-ATPases

1 Mechanisms underlying K+-induced hyperpolarizations in the presence and absence of phenylephrine were investigated in endothelium-denuded rat mesenteric arteries (for all mean values, n = 4). 2 Myocyte resting membrane potential (m.p.) was -58.8 +/- 0.8 mV. Application of 5 rum KCl produced similar hyperpolarizations in the absence (17.6 +/- 0.7 mV) or presence (15.8 +/- 1.0 mV) of 500 nM ouabain. In the presence of ouabain +30 muM barium, hyperpolarization to 5 mm KCl was essentially abolished. 3 In the presence of 10 pm phenylephrine (m.p. -33.7 +/- 3 mV), repolarization to 5 mm KCl did not occur in the presence or absence of 4-aminopyridine but was restored (-26.9 +/- 1.8 mV) on addition of iberiotoxin (100 nM). Under these conditions the K+-induced repolarization was insensitive to barium (30 muM) but abolished by 500 muM ouabain alone. 4 In the presence of phenylephrine + iberiotoxin the hyperpolarization to 5 mm K+ was inhibited in the additional presence of 300 nM levcromakalim, an action which was reversed by 10 Pm glibenclamide. 5 RT-PCR, Western blotting and immunohistochemical techniques collectively showed the presence of alpha(1)-, alpha(2),- and alpha3-subunits of Na+/K+-ATPase in the myocytes. 6 In K+-free solution, re-introduction of K+ (to 4.6 mM) hyperpolarized myocytes by 20.9 +/- 0.5 mV, an effect unchanged by 500 nM ouabain but abolished by 500 muM ouabain. 7 We conclude that under basal conditions, Na+/K+-ATPases containing alpha(2)- and/or alpha(3)-subunits are partially responsible for the observed K+-induced effects. The opening of myocyte K+ channels (by levcromakalim or phenylephrine) creates a 'K+ cloud' around the cells which fully activates Na+/K+-ATPase and thereby abolishes further responses to [K+](o) elevation.