Nitric oxide (NO) is produced in almost all tissues and organs, exerting a variety of biological actions under both physiological and pathological conditions. NO is synthesized by 3 distinct NO synthase (NOS) isoforms (neuronal, inducible, and endothelial NOS), all of which are expressed in the human cardiovascular system. The regulatory roles of NOSs in cardiovascular diseases have been described in pharmacological studies with selective and non-selective NOS inhibitors. However, the specificity of the NOS inhibitors continues to be an issue of debate. To overcome this issue, genetically engineered animals have been used. All types of NOS gene-deficient (knockout: KO) animals, including singly, doubly, and triply NOS-KO mice, and various types of NOS gene-transgenic (TG) animals, including conditional and non-conditional TG mice bearing endothelium-specific or cardiomyocyte-specific overexpression of each NOS gene, have thus far been developed. The roles of individual NOS isoforms, as well as the entire NOS system, in the cardiovascular system have been extensively investigated in those mice, and the results provide pivotal insights into the pathophysiology of NOSs in human cardiovascular diseases. Based on studies with murine NOS genetic models, this review summarizes the latest knowledge of NOSs and cardiovascular diseases. (Circ J 2009; 73: 986-993)