Necessity of inositol (1,4,5)-trisphosphate receptor 1 and μ-calpain in NO-induced osteoclast motility

Abstract

In skeletal remodeling, osteoclasts degrade bone, detach and move to new locations. Mechanical stretch and estrogen regulate osteoclast motility via nitric oxide (NO). We have found previously that NO stimulates guanylyl cyclase, activating the cGMP-dependent protein kinase 1 (PKG1), reversibly terminating osteoclast matrix degradation and attachment, and initiating motility. The PKG1 substrate vasodilator-stimulated protein (VASP), a membrane-attachment-related protein found in complexes with the integrin αvβ3 in adherent osteoclasts, was also required for motility. Here, we studied downstream mechanisms by which the NO-dependent pathway mediates osteoclast relocation. We found that NO-stimulated motility is dependent on activation of the Ca2+-activated proteinase μ-calpain. RNA interference (RNAi) showed that NO-dependent activation of μ-calpain also requires PKG1 and VASP. Inhibition of Src kinases, which are involved in the regulation of adhesion complexes, also abolished NO-stimulated calpain activity. Pharmacological inhibition and RNAi showed that calpain activation in this process is mediated by the inositol (1,4,5)-trisphosphate receptor 1 [Ins(1,4,5)P3R1] Ca2+ channel. We conclude that NO-induced motility in osteoclasts requires regulated Ca2+ release, which activates μ-calpain. This occurs via the Ins(1,4,5)P3R1.