FIG. 3.

Effects of recombinant human adiponectin on cAMP content of murine primary skeletal muscle cells. Primary skeletal muscle cells, cultured in 96-well plates, were exposed to 50 μg/ml of adiponectin and 5 μmol/l forskolin for 12 h. Adiponectin-treated cells were also pretreated with 1 μmol/l H89, a PKA inhibitor, for 1 h before the addition of adiponectin. The cAMP assay was performed according to the manufacturer's instructions. Cell lysates from primary cultured skeletal muscle cells were subjected to SDS-PAGE, followed by electrophoretic transfer to a nitrocellulose membrane. Membranes were incubated for 1 h at room temperature with antibody against mouse FHC. Detection was performed using an electrochemiluminescence chemiluminescent kit according to the manufacturer's instructions. Representative results from four independent experiments are presented. Values are means ± SE. *Significant difference (P < 0.05) relative to control cells in the absence of ligands (lane 1). N.S.; not significant relative to paired control cells in the absence of forskolin (lane 1) or H89 (lane 3).