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P2X 7 R‐mediated Ca 2+ ‐independent d ‐serine release via pannexin‐1 of the P2X 7 R‐pannexin‐1 complex in astrocytes
Author(s) -
Pan HanChi,
Chou YunChia,
Sun Synthia H.
Publication year - 2015
Publication title -
glia
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 2.954
H-Index - 164
eISSN - 1098-1136
pISSN - 0894-1491
DOI - 10.1002/glia.22790
Subject(s) - pannexin , serine , staurosporine , extracellular , protein kinase c , biology , glutamate receptor , microbiology and biotechnology , adenosine triphosphate , nmda receptor , biochemistry , intracellular , kinase , receptor , gap junction , phosphorylation , connexin
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 Ca 2+ ‐independent release mechanism mediated by P2X 7 receptors (P2X 7 R). Using [ 3 H] d ‐serine, which was loaded into astrocytes via the neutral amino acid transporter 2 (ASCT2), we observed that ATP and a potent P2X 7 R agonist, 2′(3′)‐ O ‐(4‐benzoylbenzoyl)adenosine‐5′‐triphosphate (BzATP), stimulated [ 3 H]D‐serine release and that were abolished by P2X 7 R selective antagonists and by shRNAs, whereas enhanced by removal of intracellular or extracellular Ca 2+ . The P2X 7 R‐mediated d ‐serine release was inhibited by pannexin‐1 antagonists, such as carbenoxolone (CBX), probenecid (PBN), and 10 Panx‐1 peptide, and shRNAs, and stimulation of P2X 7 R induced P2X 7 R‐pannexin‐1 complex formation. Simply incubating astrocytes in Ca 2+ /Mg 2+ ‐free buffer also induced the complex formation, and that enhanced basal d ‐serine release through pannexin‐1. The P2X 7 R‐mediated d ‐serine release assayed in Ca 2+ /Mg 2+ ‐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 Ca 2+ ‐dependent PKC inhibitor, Gö6976, inhibited the P2X 7 R‐mediated d ‐serine release. Thus, we conclude that in astrocytes, P2X 7 R‐pannexin‐1 complex formation is crucial for P2X 7 R‐mediated d ‐serine release through pannexin‐1 hemichannel. The release is Ca 2+ ‐independent and regulates by a Ca 2+ ‐independent PKC. The activated P2X 7 R per se is also functioned as a permeation channel to release d ‐serine in part. This P2X 7 R‐mediated d ‐serine release represents an important mechanism for activity‐dependent neuron‐glia interaction. GLIA 2015;63:877–893