OBJECTIVE: The overproduction of vascular NO contributes toward the circulatory collapse observed in patients with septic shock. Dimethylarginine dimethylaminohydrolase (DDAH), which has 2 isoforms, metabolizes asymmetrically methylated arginines (asymmetric mono- or di-methylarginine), endogenously produced NO synthase inhibitors. We wished to investigate whether reducing DDAH1 activity, using genetic and pharmacological approaches, is protective during lipopolysaccharide-induced endotoxic shock. METHODS AND RESULTS: Experiments were conducted in DDAH1 heterozygous knockout mice (DDAH1(+/-)) or naive rats treated with a synthetic pharmacological DDAH inhibitor (L-257). We demonstrate for the first time that L-257 is DDAH1 selective using recombinant human DDAH proteins. DDAH1 mRNA was expressed in aortic but not macrophage cDNA, and consistent with this expression profile, L-257 selectively inhibited NO production from lipopolysaccharide-treated aorta but not macrophages, in culture. Conscious and anesthetized cardiovascular hemodynamics were monitored using implanted radiotelemetry devices or invasive catheters, respectively. Lipopolysaccharide was administered intravenously to model endotoxemia, and all animals presented with circulatory shock. DDAH1(+/-) mice or L-257-treated rats displayed attenuation in the rate of developed hypotension compared with wild-type littermates or vehicle control animals, respectively. CONCLUSIONS: Pharmacological and genetic reduction of DDAH1 activity is protective against the vascular changes observed during endotoxic shock.