Isolation from Rat Adipocytes of a Chemical Mediator for Insulin Activation of Pyruvate Dehydrogenase Free

Insulin treatment of adipocytes increased the amount or activity of a low molecular weight, acid-stable material which, when isolated from intact adipocytes by heat extraction and subsequent Sephadex G25 chromatography, yielded a single active fraction that stimulated mitochondrial pyruvate dehydrogenase by activating the phosphatase and not by altering the kinase activity. Phosphatase activation was demonstrated by the ability of the active material to increase pyruvate dehydrogenase activity in the absence of ATP and by the ability of NaF, a phosphatase inhibitor, to block this stimulation. Involvement of the kinase in this activation mechanism was eliminated by the fact that, in the presence of ATP, (1) NaF completely blocked the stimu ation of pyruvate dehydrogenase by the active fraction, and (2) the stimulation of pyruvate dehydrogenase by dichloroacetic acid, akinase inhibitor, was add itive to the stimulation caused by the active fraction. This active fraction may contain an intracellular chemical mediator or second messenger for insulin.