Insulin stimulates both the alpha 1 and the alpha 2 isoforms of the rat adipocyte (Na+,K+) ATPase. Two mechanisms of stimulation.
Results obtained with adipocyte ghosts indicated that the relative pumping activities of the alpha 1 and alpha 2 isoforms of the (Na+,K+) pump depend strongly on intracellular sodium concentration, [Na+]i (McGill, D. (1991) J. Biol. Chem. 266, 15817-15823). Accordingly, [Na+]i was determined in rat adipocytes as a function of ouabain concentration and found to increase gradually as the concentration of ouabain increased. Incubation conditions were therefore designed such that the [Na+]i at 0 M and 10(-5) M ouabain were identical, in order to study the activities of both forms of the pump under identical conditions. Under these conditions, the alpha 2 isozyme accounts for 42% of the total pumping activity; these data prove that the activity of the alpha 2 isozyme is suppressed to a much greater extent than that of the alpha 1 isozyme, in relation to maximally obtainable activities measured in plasma membranes (Lytton J., Lin, J.C., and Guidotti, G. (1985) J. Biol. Chem. 260, 1177-1184). Furthermore, insulin stimulation of 86Rb+/K+ uptake in adipocytes results from a 58 and a 128% increase in the activities of the alpha 1 and alpha 2 isozymes (Na+,K+) pump, respectively. In addition, it is shown that under the conditions used to determine the [Na+]i dependence of 86Rb+/K+ uptake into adipocytes (0 mM KCl, various [NaCl]), [Na+]i decreases rapidly upon the addition of KCl/86RbCl for the initiation of the uptake measurement. By making uptake measurements quickly after the addition of KCl to eliminate the effect of a decreasing [Na+]i, we demonstrate that the stimulation of the alpha 1 isozyme is due to a small decrease in the K0.5Na+ whereas the stimulation of the alpha 2 isozyme results from a decrease in the K0.5Na+ and an increase in the Vmax.
McGill
D L
DL
Department of Biochemistry and Molecular Biology, Harvard University, Cambridge, Massachusetts 02138.
Guidotti
G
G
D000273:Adipose Tissue / Q000166:cytology / Q000187:drug effects / Q000201:enzymology*; D000818:Animals; D002462:Cell Membrane / Q000187:drug effects / Q000201:enzymology; D007328:Insulin / Q000494:pharmacology*; D007527:Isoenzymes / Q000378:metabolism*; D008297:Male; D010042:Ouabain / Q000494:pharmacology; D051381:Rats; D012964:Sodium / Q000378:metabolism; D000254:Sodium-Potassium-Exchanging ATPase / Q000378:metabolism*
D007328:Insulin; D007527:Isoenzymes; D010042:Ouabain; D012964:Sodium; D000254:Sodium-Potassium-Exchanging ATPase