Characterization and Channel Coupling of the P2Y12Nucleotide Receptor of Brain Capillary Endothelial Cells

• Published in: The Journal of biological chemistry Vol. 277; no. 35; pp. 31390 - 31400
• Main Authors: Simon, Joseph, Filippov, Alexander K., Göransson, Sara, Wong, Yung H., Frelin, Christian, Michel, Anton D., Brown, David A., Barnard, Eric A.
• Format: Journal Article
• Language: English
• Published: American Society for Biochemistry and Molecular Biology 01.08.2002
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1074/jbc.M110714200

Rat brain capillary endothelial (B10) cells express an unidentified nucleotide receptor linked to adenylyl cyclase inhibition. We show that this receptor in B10 cells is identical in sequence to the P2Y 12 ADP receptor (“ P2Y T ”) of platelets. When expressed heterologously, 2-methylthio-ADP (2-MeSADP; EC 50 , 2 n m ), ADP, and adenosine 5′- O -(2-thio)diphosphate were agonists of cAMP decrease, and 2-propylthio- d -β,γ-difluoromethylene-ATP was a competitive antagonist ( K B , 28 n m ), as in platelets. However, 2-methylthio-ATP (2-MeSATP) (EC 50 , 0.4 n m ), ATP (1.9 μ m ), and 2-chloro-ATP (190 n m ), antagonists in the platelet, were also agonists. 2-MeSADP activated (EC 50 , 0.1 n m ) GIRK1/GIRK2 inward rectifier K + channels when co-expressed with P2Y 12 receptors in sympathetic neurons. Surprisingly, P2Y 1 receptors expressed likewise gave that response; however, a full inactivation followed, absent with P2Y 12 receptors. A new P2Y 12 -mediated transduction was found, the closing of native N-type Ca 2+ channels; again both 2-MeSATP and 2-MeSADP are agonists (EC 50 , 0.04 and 0.1 n m , respectively). That action, like their cAMP response, was pertussis toxin-sensitive. The Ca 2+ channel inhibition and K + channel activation are mediated by βγ subunit release from a heterotrimeric G-protein. Gα subunit types in B10 cells were also identified. The presence in the brain capillary endothelial cell of the P2Y 12 receptor is a significant extension of its functional range.