Mechanisms underlying increases in rat soleus Na+-K+-ATPase activity by induced contractions

Mechanisms underlying increases in rat soleus Na+-K+-ATPase activity by induced contractions. J Appl Physiol 99: 2222-2232, 2005. First published August 18, 2005; doi: 10.1152/japplphysiol.00577.2005.-Acute regulation of the Na+-K+-ATPase activity in rat soleus muscle was investigated in response to 15 and 90 min of electrically induced contractile activity (500- ms trains at 30 Hz every 1.5 s). Kinetic measurements of Na+-K+-ATPase activity, assessed by the 3-O-methylfluorescein K+-stimulated phosphatase assay (3-O-MFP), were performed on crude homogenates (Hom) and on tissue separated into two membrane fractions, the sarcolemmal/particulate (SLP) and endosomal (En), in both stimulated (Stim) and contralateral control (Con) muscles. Maximal 3-O-MFP activity (V-max, nmol(.)mg protein(-1.) h(-1)) was elevated (P < 0.05) in Stim by 40% and by 53% in Hom and by 37 and 40% in SLP at 15 and 90 min, respectively. The 38% increase ( P < 0.05) in the alpha(2)-isoform subunit distribution in SLP at 15 min, as assessed by quantitative immunoblotting, persisted at 90 min, whereas for En a 42% decrease ( P < 0.05) was observed only at 15 min. For the alpha(1)-subunit at 15 min, a 27% decrease ( P < 0.05) was observed in En, whereas the 13% increase observed in SLP was not significant (P = 0.08). At 90 min, alpha(1) was increased (P < 0.05) by 14% in SLP and by 29% in En. No changes were observed in alpha(1)-subunit distribution in En and SLP regardless of time of stimulation. Immunoprecipitation with antiphosphotyrosine antibody and quantitative immunoblotting with alpha(1)- and alpha(2)-antibodies indicated increases ( P < 0.05) in tyrosine phosphorylation of 51% in alpha(2) at 15 min only. These results suggest that the increases in Vmax during contractile activity are mediated both by increased phosphorylation and by translocation of the enzyme to the plasma membrane.