NITRIC oxide is the major endothelium-derived relaxing factor (EDRF)^1–3, and it is thought to relax smooth muscle cells by stimulation of guanylate cyclase, accumulation of its product cyclic GMP, and cGMP-dependent modification of several intracellular processes^4,5, including activation of potassium channels through cGMP-dependent protein kinase^6,7. Here we present evidence that both exogenous nitric oxide and native EDRF can directly activate single Ca²⁺-dependent K⁺ channels (K⁺_Ca) in cell-free membrane patches without requiring cGMP. Under conditions when guanylate cyclase was inhibited by methylene blue, considerable relaxation of rabbit aorta to nitric oxide persisted which was blocked by charybdotoxin, a specific inhibitor of K⁺_Ca channels. These studies demonstrate a novel direct action of nitric oxide on K⁺_Ca channels.