Aging is associated with an increase in insulin resistance in skeletal muscle, yet the underlying mechanism is not well established. We hypothesize that with aging, a chronic increase in stress kinase activation, coupled with a decrease in oxidative capacity, leads to insulin resistance in skeletal muscle. In aged (24 month-old) and young (3 month-old) Fischer 344 rats, 2-deoxy glucose uptake and insulin signaling (as measured by phosphorylation of IRS-1, Akt and AS160) decreased significantly with age. Activation of c-Jun NH2-terminal kinase (JNK), glycogen serine kinase-3 (GSK-3), and degradation of IB by the upstream inhibitor of kappa B kinase (IKK), as measured by Western blot analysis, were increased with age in both soleus and Epi muscles. However, much higher activation of these kinases in Epi muscles from young rats compared to soleus results in a greater effect of these kinases on insulin signaling in fast-twitch muscle with age. Heat shock protein 72 (HSP72) expression and phosphorylation of HSP25 were higher in soleus compared to Epi muscles and both parameters decreased with age. Age and fiber type differences in cytochrome oxidase activity are consistent with observed changes in HSP expression and activation. Our results demonstrate a significant difference in the ability of slow-twitch and fast-twitch muscles to respond to insulin and regulate glucose with age. A greater constitutive HSP expression and lower stress kinase activation may account for the ability of slow-twitch muscles to preserve the capacity to respond to insulin and maintain glucose homeostasis with age.