This study determined the influence of Rho in regulating phenotype in early passage rabbit aortic smooth muscle cell (SMC) culture. Extended confluence and serum-deprivation induced SMC differentiation, indicated by increased expression of contractile markers (α-SM actin, SM-MHC and SM-2). Activation of Rho by sphingosine-1-phosphate (S-1-P) further enhanced the expression of contractile proteins and their reorganisation such that cells adopted a more 'contractile' phenotype. Inhibition of Rho induced an extreme 'synthetic' morphology including increased vimentin expression.