The purpose of this study was to determine whether expression of inducible nitric oxide synthase (iNOS) is altered in vascular smooth muscle cells of type 2 diabetes rats. We used cultured aortic smooth muscle cells (ASMCs) isolated from male Goto–Kakizaki diabetes rats (G–K rats) aged 27–28 weeks and age-matched Wistar rats (control rats). iNOS and extracellular signal-regulated kinase (ERK) were evaluated by immunoblot and/or immunochemical analyses, and NO production was evaluated by measuring NO X (NO 2 and NO 3). Expression of iNOS was not detected in ASMCs of either G–K or control rats under a resting condition. Stimulation with interleukin-1β (IL-1β) induced iNOS expression, which was much greater in ASMCs from G–K rats than in ASMCs from control rats. When ASMCs were stimulated with IL-1β, the number of iNOS-immunoreactive ASMCs from G–K rats increased more prominently than did the number of such ASMCs from control rats. IL-1β-induced NO production was also much greater in ASMCs from G–K rats than in those from control rats. Both IL-1β-induced iNOS expression and NO production in ASMCs of G–K and control rats were markedly reduced in the presence of an ERK inhibitor, U0126 or PD98059. Both basal and IL-1β-stimulated levels of ERK activity were significantly higher in ASMCs from G–K rats than in ASMCs from control rats. The results suggest that iNOS induction is enhanced in cultured ASMCs from G–K rats and that this enhancement is associated with increased ERK activity.