Hypo‐osmotic potentiation of acetylcholine‐stimulated ciliary beat frequency through ATP release in rat tracheal ciliary cells

The ciliary beat frequency (CBF) of rat tracheal ciliary cells in a slice preparation was measured using video‐enhanced contrast (VEC) microscopy. Acetylcholine (ACh) increased CBF mediated via intracellular Ca 2+ concentration ([Ca 2+ ] i ) in a dose‐dependent manner. An adequate hypo‐osmotic stress (−40 mos m ) potentiated ACh‐stimulated CBF increase in tracheal ciliary cells and shifted the ACh dose–response curve to the left (lower concentration side). This potentiation was independent of hypo‐osmotic stresses applied ranging from −20 mosM to −90 mosM. A hypo‐osmotic stress induces ATP release in many cell types. The present study demonstrated that suramin (an inhibitor of purinergic receptors) and apyrase (an ATPase/ADPase) eliminate the hypo‐osmotic potentiation of ACh‐stimulated CBF increase and that ATP increased [Ca 2+ ] i and CBF, as well as potentiating ACh‐stimulated rises in [Ca 2+ ] i and CBF increase. Moreover, the apical surface of tracheal ciliary cells were stained immunopositive for the P2X 4 purinergic receptor. A hypo‐osmotic stress (−40 mosM) transiently increased [Ca 2+ ] i and potentiated the ACh‐stimulated [Ca 2+ ] i increase. The hypo‐osmotic potentiation of ACh‐stimulated CBF increase was not detected under Ca 2+ ‐free conditions. These observations suggest that a hypo‐osmotic stress stimulates ATP release from the trachea. The released ATP may induce further increases in [Ca 2+ ] i and CBF in ACh‐stimulated tracheal ciliary cells, which may be mediated by purinergic receptors, such as P2X 4 .