FIG. 4.

Comparison of Akt and PI3K inhibition demonstrates that insulin regulates lipolysis independently of Akt kinase activity. Glycerol release was performed on 3T3-L1 adipocytes using the indicated additions: isoproterenol (Iso; 2 nM), insulin (Ins; 25 nM), Akt inhibitor (Akti; 10 μM), and LY294002 (LY; 50 and 100 μM). Western blotting was performed on whole-cell lysates as shown. Immunoprecipitation was performed using pan-Akt antibody. The immunoprecipitates were subjected to Western blot analysis, and Akt kinase activity was measured using GSK fusion protein. The degree of phosphorylation was assessed using the Licor Odyssey system and graphed as indicated.