Mechanical and osmotic sensitivity of the transient receptor potential vanilloid 4 (TRPV4) channel depends on phospholipase A(2) (PLA(2)) activation and the subsequent production of the arachidonic acid metabolites, epoxyeicosatrienoic acid (EET). We show that both high viscous loading and hypotonicity stimuli in native ciliated epithelial cells use PLA(2)-EET as the primary pathway to activate TRPV4. Under conditions of low PLA(2) activation, both also use extracellular ATP-mediated activation of phospholipase C (PLC)-inositol trisphosphate (IP(3)) signaling to support TRPV4 gating. IP(3), without being an agonist itself, sensitizes TRPV4 to EET in epithelial ciliated cells and cells heterologously expressing TRPV4, an effect inhibited by the IP(3) receptor antagonist xestospongin C. Coimmunoprecipitation assays indicated a physical interaction between TRPV4 and IP(3) receptor 3. Collectively, our study suggests a functional coupling between plasma membrane TRPV4 channels and intracellular store Ca(2+) channels required to initiate and maintain the oscillatory Ca(2+) signal triggered by high viscosity and hypotonic stimuli that do not reach a threshold level of PLA(2) activation.

This work was supported by grants from the Spanish Ministries of Education and Science (SAF2006-04973 and SAF2006-13893-C02-02), and Health (Fondo de Investigación Sanitaria, Red HERACLES RD06/0009), the Generalitat de Catalunya (SGR05-266), and Fundació la Marató de TV3 (061331). J.M. Fernández-Fernández is a Ramón y Cajal Fellow.