The application of mechanical stresses to the airway smooth muscle (ASM) cell causes time-dependent cytoskeletal stiffening and remodeling (Deng L, Fairbank NJ, Fabry B, Smith PG, and Maksym GN. Am J Physiol Cell Physiol 287: C440-C448, 2004). We investigated here the extent to which these behaviors are modulated by the state of cell activation (tone). Localized mechanical stress was applied to the ASM cell in culture via oscillating beads (4.5 mum) that were tightly bound to the actin cytoskeleton (CSK). Tone was reduced from baseline level using a panel of relaxant agonists (10(-3) M dibutyryl cAMP, 10(-4) M forskolin, or 10(-6) M formoterol). To assess functional changes, we measured cell stiffness (G') using optical magnetic twisting cytometry, and to assess structural changes of the CSK we measured actin accumulation in the neighborhood of the bead. Applied mechanical stress caused a twofold increase in G' at 120 min. After cessation of applied stress, G' diminished only 24 +/- 6% (mean +/- SE) at 1 h, leaving substantial residual effects that were largely irreversible. However, applied stress-induced stiffening could be prevented by ablation of tone. Ablation of tone also inhibited the amount of actin accumulation induced by applied mechanical stress (P < 0.05). Thus the greater the contractile tone, the greater was applied stress-induced CSK stiffening and remodeling. As regards pathobiology of asthma, this suggests a maladaptive positive feedback in which tone potentiates ASM remodeling and stiffening that further increases stress and possibly leads to worsening airway function.