Purified cyclic AMP-dependent protein kinase (cAK) catalytic subunit phosphorylated 180-, 49-, 31-, 19-, and 14-kilodalton (kDa) proteins of rabbit sciatic nerve membranes. The ability of cAK to phosphorylate these membrane substrate proteins was inhibited by gangliosides GM1, GD1a, and GT1b with half-maximal inhibitory concentration (I50) = 7-25 microM. Neutral glycolipids and lysophosphatidylcholine were much less effective. Cyclic AMP (cAMP) kinase phosphorylation of histone IIA was inhibited by GM1, GD1a, and GT1b (I50 = 115 microM, 75 microM, and 75 microM, respectively). Inhibition by GM1 was competitive with respect to histone (Ki = 108 microM). Autophosphorylation of cAMP kinase was inhibited by GM1 (I50 = 15 microM). GT1b, GD1a, and GM1 half-maximally stimulated calmodulin-dependent cyclic nucleotide phosphodiesterase at 0.1 microM, 0.2 microM, and 0.3 microM, respectively. Although GT1b stimulated phosphodiesterase by increasing Vmax and decreasing Km (similar to calmodulin), GD1a and GM1 produced only an increase in Vmax. These results suggest that ganglioside can modulate the activity of cAMP kinase by both direct inhibition of the enzyme and indirect reduction of cAMP levels through activation of phosphodiesterase. Through these mechanisms, gangliosides may alter cAMP-dependent protein phosphorylation and cell function within the nervous system.