Aortic vascular smooth muscle cells (VSMC) were used to study the effect of age on responses to high glucose concentrations or the cytokine, tumor necrosis factor-alpha (TNF-α). Activator protein-1 (AP-1) binding to DNA increased more in VSMC from old versus young rats (P < 0.02) and was related to increased expression of its components, c-Fos, Fra-1, and JunD. The relationship to upstream signals, i.e., activities of mitogen-activated protein kinases (MAPK), was studied using antibodies to total and phosphorylated forms of extracellular signal-regulated kinases (ERK), c-Jun N-terminal kinases (JNK) and p38. High glucose and TNF-α increased ERK phosphorylation more in old (P < 0.05); whereas only TNF-α induced JNK activation in young (P < 0.04). PD98059, a MEK inhibitor, attenuated AP-1 activation, lowered c-Fos and Fra-1 protein levels and reduced cell number and cells positive for proliferating cell nuclear antigen in old. We concluded that age differentially influenced activation of signaling pathways in VSMC exposed to high glucose or TNF-α. This may contribute to the increased risk for vascular disease associated with aging and diabetes mellitus (DM). J. Cell. Physiol. 197: 418425, 2003© 2003 Wiley-Liss, Inc. [ABSTRACT FROM AUTHOR]