The outward K+ current induced by step depolarization of freshly dispersed myocytes of guinea-pig taenia coli decreased about 80% upon treatment with 3 mM tetraethylammonium chloride (TEA). Isoproterenol (ISO, 2-5 microM) restored it to a large extent. This restoration did not occur in the presence of propranolol (2 microM). In single-channel recordings from cell-attached patches, the activity of maxi-K+ channel is dominant. When 3 mM TEA is incorporated in the pipette solution, the dominant channel-openings observed had much smaller unitary conductance. On the addition of ISO (2 microM) to the bath solution, but not to the pipette solution, K(+)-channel openings with unitary conductance similar to that without TEA treatment appeared. Cyclic AMP incorporated into the cytoplasm through the pipette was ineffective. These results indicate that ISO release TEA decrease of maxi-K+ channel conductance through some intracellular second messenger system other than adenylyl cyclase-protein kinase A system.