Objective: The aim was to investigate the release of nitric oxide (NO) from porcine mitral valves. Methods: The valves were sandwiched between gauze in a chamber constantly perfused by Holman solution. The presence of NO in the perfusate of the valves was detected by two methods: a bioassay cascade system using preconstricted rings of pig coronary artery as the detector tissue, and a chemiluminescent method, specific for NO. Results: Porcine mitral valves release NO under basal conditions. Release can be increased by the agonists ADP, substance P, bradykinin, thrombin, and the calcium ionophore A23187. ADP was by far the most effective. NO release under basal conditions was inhibited by each of the three NO synthase inhibitors studied, but only NG-monomethyl L-arginine (L-NMMA) reduced ADP stimulated release. A reversible inhibition of NO release occurred when valves were perfused with calcium-free Holman solution and NO release was inhibited by removal of the endocardial monolayer. Conclusions: The endocardial endothelium covering the mitral valves releases NO under basal conditions and following stimulation with several agonists. The NO synthase enzyme is calcium dependent. Different mechanisms may exist for the synthesis of basal and agonist dependent NO release in these cells. Cardiovascular Research 1993;27:1657-1661