P2X(4) purinoceptors mediate an ATP-activated, non-selective cation current in rabbit osteoclasts

Extracellular nucleotides act as signaling molecules in numerous tissues. I n bone, nucleotides stimulate osteoclast formation and activity; however, t he receptors and signaling mechanisms underlying these effects have yet to be identified. To identify specific P2X purinoceptor subtypes in osteoclast s, degenerate oligonucleotide primers were used to PCR-amplify DNA fragment s from a rabbit osteoclast cDNA library. A 372-base-pair fragment was obtai ned that encoded an amino acid sequence with 88% identity to the rat P2X(4) purinoceptor, The presence of P2X(4) mRNA in purified osteoclasts was conf irmed by reverse transcription-PCR. Endogenous purinoceptors were functiona lly characterized in isolated rabbit osteoclasts by patch-clamp recording i n whole-cell configuration. At negative membrane potentials, application of ATP or ADP rapidly activated an inward current followed by an outward curr ent. In contrast, UTP or ADP beta S elicited only an outward current, due t o activation of a Ca2+-dependent K+ conductance. The initial inward current was nonselective for cations and inactivated during agonist application, F urthermore, the inward current was insensitive to suramin and Cibacron blue , and was potentiated by Zn2+, These characteristics are consistent with pr operties of P2X(4) purinoceptors. Activation of P2X(4) purinoceptors leads to cation influx and depolarization. Nucleotides, released at sites of trau ma or inflammation, may act through these receptors on osteoclasts to stimu late bone resorption.