Caldesmon (CaD) is a smooth muscle regulatory protein, which consists of a C-terminal actin binding domain known to inhibit actomyosin ATPase activity, a central spacer region, and a N-terminal myosin binding domain the funcion of which is sill uncertain. To assess the function of this domain for regulation of arterial tone a mouse model was generated which expresses a truncated CaD lacking the myosin binding domain due to ablation of exon2 (ex2-/-). In A. saphena isolated from ex2-/- mice, there was a trend for an increased reactivity towards phenylephrine (PE). Relaxtion induced by acetylcholine of isometrically preconstricted arteries (0.5 mM PE) was augmented (pEC50 7.9±0.1, n=5 versus pEC50 6.8±0.05; n=3 in wild type, WT; p <0.001). The effect of acetylcholine was also similarly enhanced in tail arteries and was completely inhibited by 100 mM L-NAME in ex2-/- and WT arteries. In both types of arteries relaxation induced by DEA-NO was not significantly different between Ex2-/- and WT. In pressurized A. gracilis the response to increasing pressure was not changed in ex2-/- mice, while there was small increase in the sensitivity to acethylcholine but not to DEA-NO. In conclusion, CaD through its myosin-binding domain appears to be involved in the regulation of arterial tone but surprisingly not by affecting the tone of the smooth muscle cells themselves but by augmenting NO-mediated endothelial relaxation.