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This Article Full Text Full Text (PDF) Supplemental Figures All Versions of this Article: 295/3/C752    most recent 00228.2008v1 Alert me when this article is cited Alert me if a correction is posted Citation Map Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Google Scholar Articles by Iglesias, R. Articles by Scemes, E. PubMed PubMed Citation Articles by Iglesias, R. Articles by Scemes, E.

RECEPTORS AND SIGNAL TRANSDUCTION

P2X₇ receptor-Pannexin1 complex: pharmacology and signaling

¹The Dominick P. Purpura Department of Neuroscience, Albert Einstein College of Medicine, Bronx, New York; ²Department of Physiology and Biophysics, School of Medicine, University of Miami, Miami, Florida; and ³Department of Immunology, Instituto Oswaldo Cruz, Manguinhos, Rio de Janeiro, Brazil

Submitted 25 April 2008 ; accepted in final form 25 June 2008

Pannexin 1 (Panx1), an ortholog to invertebrate innexin gap junctions, has recently been proposed to be the pore induced by P2X₇ receptor (P2X₇R) activation. We explored the pharmacological action of compounds known to block gap junctions on Panx1 channels activated by the P2X₇R and the mechanisms involved in the interaction between these two proteins. Whole cell recordings revealed distinct P2X₇R and Panx1 currents in response to agonists. Activation of Panx1 currents following P2X₇R stimulation or by membrane depolarization was blocked by Panx1 small-interfering RNA (siRNA) and with mefloquine > carbenoxolone > flufenamic acid. Incubation of cells with KN-62, a P2X₇R antagonist, prevented current activation by 2'(3')-O-(4-benzoylbenzoyl)adenosine 5'-triphosphate (BzATP). Membrane permeabilization to dye induced by BzATP was also prevented by Panx1 siRNA and by carbenoxolone and mefloquine. Membrane permeant (TAT-P2X₇) peptides, provided evidence that the Src homology 3 death domain of the COOH-terminus of the P2X₇R is involved in the initial steps of the signal transduction events leading to Panx1 activation and that a Src tyrosine kinase is likely involved in this process. Competition assays indicated that 20 µM TAT-P2X₇ peptide caused 50% reduction in Src binding to the P2X₇R complex. Src tyrosine phosphorylation following BzATP stimulation was reduced by KN-62, TAT-P2X₇ peptide, and by the Src tyrosine inhibitor PP2 and these compounds prevented both large-conductance Panx1 currents and membrane permeabilization. These results together with the lack Panx1 tyrosine phosphorylation in response to P2X₇R stimulation indicate the involvement of an additional molecule in the tyrosine kinase signal transduction pathway mediating Panx1 activation through the P2X₇R.

gap junction blockers; permeabilization; P_2Z; src-tyrosine kinase