In response to mechanical stretch, airway smooth muscle exhibits various cellular functions such as contraction, proliferation, and cytoskeletal remodeling, all of which are implicated in the pathophysiology of asthma. We tested the hypothesis that mechanical stretch of airway smooth muscle cells increases intracellular Ca(2+) concentration ([Ca(2+)](i)) by activating stretch-activated (SA) nonselective cation channels. A single uniaxial stretch (3 s) was given to human bronchial smooth muscle cells cultured on an elastic silicone membrane. After the mechanical stretch, a transient increase in [Ca(2+)](i) was observed. The [Ca(2+)](i) increase was significantly dependent on stretch amplitude. The augmented [Ca(2+)](i) due to stretch was completely abolished by removal of extracellular Ca(2+) and was markedly attenuated by an application of Gd(3+), an inhibitor of SA channels, or ruthenium red, a transient receptor potential vanilloid (TRPV) inhibitor. In contrast, the stretch-induced rises of [Ca(2+)](i) were not altered by other Ca(2+) channel inhibitors such as nifedipine, BTP-2, and SKF-96365. Moreover, the [Ca(2+)](i) increases were not affected by indomethacin, a cyclooxygenase inhibitor, U-73122, a phospholipase C inhibitor, or xestospongin C, an inhibitor of the inositol-trisphosphate receptor. These findings demonstrate that a novel Ca(2+) influx pathway activated by mechanical stretch, possibly through the Ca(2+)-permeable SA channel activated directly by stretch rather than by indirect mechanisms via intracellular messenger production, is involved in human airway smooth muscle cells. A molecular candidate for the putative SA channel may be one of the members of the TRPV channel family. Thus, abnormal Ca(2+) homeostasis in response to excessive mechanical strain would contribute to the pathogenesis of asthma.