Action of insulin on Na(+)-K(+)-ATPase and the Na(+)-K(+)-2Cl- cotransporter in 3T3-L1 adipocytes.
The Na(+)-K(+)-ATPase presents several different isoforms of its alpha- and beta-subunits. We detected alpha 1- and beta 1-mRNA transcripts and polypeptides in 3T3-L1 fibroblasts; during differentiation into adipocytes, alpha 1-mRNA decreased, alpha 2-mRNA was induced, beta 1-mRNA dropped to undetectable levels, and beta 2-mRNA was never expressed, suggesting that 3T3-L1 adipocytes may express an unidentified Na(+)-K(+)-ATPase beta-subunit isoform. Insulin rapidly increased ion pump activity [ouabain-sensitive 86Rb+(K+) uptake] in 3T3-L1 fibroblasts and adipocytes without changing the plasma membrane concentration of alpha 1- or alpha 2-subunits as determined by subcellular membrane fractionation and immunoblotting or by [3H]ouabain binding to intact cells. Monensin, which raises the concentration of intracellular Na+, increased Na(+)-K+ pump activity, and no further stimulation was achieved with insulin. The stimulation of the pump by insulin was reduced by bumetanide, an inhibitor of the Na(+)-K(+)-2Cl- cotransporter, and was prevented by omission of extracellular Cl-. Insulin increased both ouabain-sensitive and bumetanide-sensitive 86Rb+(K+) uptake. These results suggest that insulin activation of the Na(+)-K(+)-ATPase in 3T3-L1 adipocytes is mediated by an elevation in intracellular Na+ that is likely the consequence of Na(+)-K(+)-2Cl- cotransporter activation.
Sargeant
R J
RJ
Division of Cell Biology, Hospital for Sick Children, Toronto, Ontario, Canada.
Liu
Z
Z
Klip
A
A
D017667:Adipocytes / Q000187:drug effects* / Q000378:metabolism*; D002352:Carrier Proteins / Q000378:metabolism*; D002454:Cell Differentiation; D002460:Cell Line; D003600:Cytosol / Q000378:metabolism; D005347:Fibroblasts / Q000166:cytology; D007328:Insulin / Q000494:pharmacology*; D007527:Isoenzymes / Q000235:genetics; D010042:Ouabain / Q000378:metabolism / Q000494:pharmacology; D010455:Peptides / Q000378:metabolism; D011188:Potassium / Q000493:pharmacokinetics; D012333:RNA, Messenger / Q000378:metabolism; D012413:Rubidium / Q000493:pharmacokinetics; D012964:Sodium / Q000502:physiology; D028021:Sodium-Potassium-Chloride Symporters; D000254:Sodium-Potassium-Exchanging ATPase / Q000235:genetics / Q000378:metabolism*; D013347:Subcellular Fractions / Q000378:metabolism
D002352:Carrier Proteins; D007328:Insulin; D007527:Isoenzymes; D010455:Peptides; D012333:RNA, Messenger; D028021:Sodium-Potassium-Chloride Symporters; D010042:Ouabain; D012964:Sodium; D000254:Sodium-Potassium-Exchanging ATPase; D012413:Rubidium; D011188:Potassium