Activity of adenosine diphosphates and triphosphates on a P2YT‐type receptor in brain capillary endothelial cells

Abstract

A P2Y (nucleotide) receptor activity in a clonal population (B10) of rat brain capillary endothelial cells is coupled to inhibition of adenylyl cyclase and has functional similarities to the P2Y_T (previously designated ‘P2T’) receptor for ADP of blood platelets. However, the only P2Y receptor which was detectable in a previous study of B10 cells by mRNA analysis was the P2Y₁ receptor, which elsewhere shows no transduction via cyclic nucleotides. We have sought here to clarify these issues. The inhibition of forskolin‐stimulated adenylyl cyclase induced by purified nucleotides was measured on B10 cells. The EC₅₀ value for 2‐methylthioADP (2‐MeSADP) was 2.2 nM and, surprisingly, 2‐MeSATP was an almost equally strong agonist (EC₅₀=3.5 nM). ATP and 2‐ClATP were weak partial agonists (EC₅₀=26 μM and 10 μM respectively) and under appropriate conditions could antagonise the activity on 2‐MeSADP. A known selective antagonist of the platelet P2Y_T receptor, 2‐propylthioadenosine‐5′‐(β,γ)‐difluoromethylene) triphosphonate (AR‐C 66096), was a competitive antagonist of this B10 cell receptor, with pK_B=7.6. That ligand is inactive at the P2Y₁ receptor in the same cells. Conversely, the competitive P2Y₁ receptor antagonists, the 3′, 5′‐ and 2′, 5′‐adenosine bis‐monophosphates, are, instead, weak agonists at the adenylyl cyclase‐inhibitory receptor. The inhibition of adenylyl cyclase by 2‐MeSADP was completely abolished by pertussis toxin. In summary, these brain endothelial cells possess a P2Y_T‐type receptor in addition to the P2Y₁ receptor. The two have similarities in agonist profiles but are clearly distinguishable by antagonists and by their second messenger activations. The possible relationships between the B10 and platelet P2Y_T receptors are discussed. British Journal of Pharmacology (2001) 132, 173–182; doi:10.1038/sj.bjp.0703816