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P2X7R‐mediated Ca2+‐independent d‐serine release via pannexin‐1 of the P2X7R‐pannexin‐1 complex in astrocytes

d‐serine is a coagonist of N‐methyl‐d‐aspartate (NMDA) subtype of glutamate receptor and plays a role in regulating activity‐dependent synaptic plasticity. In this study, we examined the mechanism by which extracellular ATP triggers the release of d‐serine from astrocytes and discovered a novel Ca2+‐independent release mechanism mediated by P2X7 receptors (P2X7R). Using [3H] d‐serine, which was loaded into astrocytes via the neutral amino acid transporter 2 (ASCT2), we observed that ATP and a potent P2X7R agonist, 2′(3′)‐O‐(4‐benzoylbenzoyl)adenosine‐5′‐triphosphate (BzATP), stimulated [3H]D‐serine release and that were abolished by P2X7R selective antagonists and by shRNAs, whereas enhanced by removal of intracellular or extracellular Ca2+. The P2X7R‐mediated d‐serine release was inhibited by pannexin‐1 antagonists, such as carbenoxolone (CBX), probenecid (PBN), and 10Panx‐1 peptide, and shRNAs, and stimulation of P2X7R induced P2X7R‐pannexin‐1 complex formation. Simply incubating astrocytes in Ca2+/Mg2+‐free buffer also induced the complex formation, and that enhanced basal d‐serine release through pannexin‐1. The P2X7R‐mediated d‐serine release assayed in Ca2+/Mg2+‐free buffer was enhanced as well, and that was inhibited by CBX. Treating astrocytes with general protein kinase C (PKC) inhibitors, such as chelerythrine, GF109203X, and staurosporine, but not Ca2+‐dependent PKC inhibitor, Gö6976, inhibited the P2X7R‐mediated d‐serine release. Thus, we conclude that in astrocytes, P2X7R‐pannexin‐1 complex formation is crucial for P2X7R‐mediated d‐serine release through pannexin‐1 hemichannel. The release is Ca2+‐independent and regulates by a Ca2+‐independent PKC. The activated P2X7R per se is also functioned as a permeation channel to release d‐serine in part. This P2X7R‐mediated d‐serine release represents an important mechanism for activity‐dependent neuron‐glia interaction. GLIA 2015;63:877–893