A role for Ca²⁺-conducting ion channels in mechanically-induced signal transduction of airway epithelial cells

Mechanical stimulation of a single cell in a cultured monolayer of airway epithelial cells initiates an intercellularly communicated increase in intracellular Ca²⁺ concentration ([Ca²⁺](i)) that propagates radially through adjacent cells via gap junctions, forming an intercellular Ca²⁺ wave. Mechanically-induced intercellular Ca²⁺ waves also occur in the absence of extracellular Ca²⁺. However, in Ca²⁺-free medium an increase in [Ca²⁺](i) of the stimulated cell does not occur. Thus, mechanically-induced [Ca²⁺](i) changes in the stimulated cell are influenced by the extracellular Ca²⁺ concentration. To investigate if a channel-mediated Ca²⁺ flux across the plasma membrane contributes to the elevation of [Ca²⁺](i) in the stimulated cell we used digital image microscopy to measure mechanically-induced [Ca²⁺](i) changes in the presence of Ca²⁺ channel blockers. In Ca²⁺-free medium containing Gd³⁺ (20 μM) mechanical stimulation resulted in an [Ca²⁺](i) increase in the stimulated cell. The delay time between mechanical stimulation and increase in [Ca²⁺](i) of the stimulated cell was dependent on extracellular [Gd³⁺], with a half-maximal effective concentration of approximately 40 μM. Mechanical stimulation in Ca²⁺-free medium containing La³⁺ (10 μM) or Ni²⁺ (100 μM) gave similar results. Mechanical stimulation in Ca²⁺-free medium containing the dihydropyridine Ca²⁺ channel blockers nifedipine (10 μM) and nimodipine (10 μM) also resulted in an increase of [Ca²⁺](i) of the stimulated cell. Mechanical stimulation of cells treated with thapsigargin to deplete intracellular Ca²⁺ stores, in the presence of 1.3 mM extracellular Ca²⁺, results in an increase in [Ca²⁺](i) of the stimulated cell without the propagation of an intercellular Ca²⁺ wave. Mechanical stimulation of thapsigargin-treated cells in an extracellular medium buffered to 50 μM free Ca²⁺ still results in an increase in [Ca²⁺](i) in the stimulated cell. However, the mechanically-induced Ca²⁺ increase in the presence of 50 μM extracellular [Ca²⁺] can be reversibly blocked by 100 μM Ni²⁺. From these results we conclude that a flux of Ca²⁺ across the plasma membrane and through Ca²⁺-conducting channels contributes to the mechanically-induced [Ca²⁺](i) changes in the stimulated cell.