Nitric oxide production by the Ca2+/calmodulin-dependent enzyme endothelial nitric oxide synthase primarily reflects changes in intracellular [Ca2+], increasing as Ca2+rises and decreasing as Ca2+falls. This simplistic bimodal mechanism of regulation has recently been refined by the finding that the binding of Ca2+/calmodulin to endothelial nitric oxide synthase involves the disruption of the association of endothelial nitric oxide synthase from the scaffolding protein caveolin and the subsequent translocation of the enzyme from plasmalemmal caveolae. Moreover, other endothelial nitric oxide synthase-associated proteins could account for a delayed Ca2+-independent activation of nitric oxide production, and may be involved with caveolin in the reversible trafficking of a large endothelial nitric oxide synthase-containing heterocomplex between the caveolae and cytosolic cell structures.