CALMODULIN-DEPENDENT CYCLIC-AMP PHOSPHODIESTERASE IN LIVER PLASMA-MEMBRANES - STIMULATED BY INSULIN

In vivo insulin consistently stimulates the plasma membrane, high-affi nity (low K-m) cyclic AMP phosphodiesterase (PDE) from diabetic rat li ver or adipose tissue, lit vitro stimulation of membrane PDE by insuli n has been reported to be inconsistent, Also, the involvement of calmo dulin (CaM) in insulin stimulation of PDE has been controversial, In t his report, conditions for the isolation of rat liver plasma membranes containing PDE that is sensitive to in vitro insulin stimulation and the involvement of CaM in insulin stimulation of PDE were investigated , In vitro insulin raised the V-max of the enzyme without altering its apparent K-m and was dose dependent. Insulin stimulation was lost aft er freezing, sonication, solubilization with detergents, or storage of the membranes at 4 degrees C for 4 h after isolation, Insulin stimula tion was completely blocked by the CaM antagonist compound 48/80, EG;T A, or trifluoperazine, Two isoforms of membrane-bound PDE were separat ed by ion-exchange chromatography following solubilization of the plas ma membranes, The activities of both isoforms were stimulated by exoge nous CaM. Plasma membrane PDE eluted after the application of exogenou s CaM plus Ca2+. The data support the concept of a critical involvemen t of CaM in insulin activation of liver membrane PDE. (C) 1995 Academi c Press, Inc.