No abstract

Release of surfactant from pulmonary type II epithelial cells was stimulated by the beta-adrenergic agonist terbutaline and the diterpene forskolin. Cytosolic cyclic adenosine monophosphate (cAMP) concentrations increased significantly following exposure to terbutaline or forskolin and reached maximal levels within 5 min after treatment. Terbutaline and forskolin had a synergistic effect on cytosolic cAMP levels when added simultaneously. cAMP-dependent protein kinase activity was identified in cytosolic preparations of type II pneumocytes by phosphorylation of the peptide substrate Kemptide (Leu-Arg-Arg-Ala-Ser-Leu-Gly) and binding of 3H-cAMP to the regulatory components of cAMP-dependent protein kinase. Type I and type II regulatory subunits of the cANP-dependent kinase were present in approximately equal concentrations in type II cell cytosol. Activation ratio of cAMP-dependent protein kinase in cultured type II cells increased significantly in the presence of terbutaline, forskolin, or terbutaline plus forskolin. Activation ratios increased from 0.45 +/- 0.03 for control cells to 0.96 +/- 0.06 for cells exposed to terbutaline (10 microM) plus forskolin (5 microM) for 20 min. Release of 3H-phosphatidylcholine was also stimulated by terbutaline and forskolin. Effects of terbutaline and forskolin on surfactant release were approximately additive. Our results demonstrated increased cytosolic cAMP levels, increased cAMP-dependent protein kinase activation ratios, and subsequent augmented surfactant release from isolated type II pneumocytes in response to terbutaline and forskolin. These data support a role for activation of cAMP-dependent protein kinase as a mediator of surfactant release and document the utility of forskolin for study of cAMP-mediated effects in isolated type II cells.