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 three distinct NO synthase (NOS) isoforms (neuronal, inducible, and endothelial NOS), all of which are expressed in the human cardiovascular system. Although the regulatory roles of NOSs in cardiovascular diseases have been described in pharmacological studies with selective and non-selective NOS inhibitors, 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 animals, including singly, doubly, and triply NOS-deficient 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 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, providing pivotal insights into an understanding of the pathophysiology of NOSs in human cardiovascular diseases. Based on studies with the murine NOS genetic models, this review briefly summarizes the latest knowledge of NOSs and cardiovascular diseases.