Pharmacological properties of ATP-sensitive purinergic receptors expressed in human G292 osteoblastic cells

Abstract

We characterized the pharmacological properties of P2 receptors expressed in G292 osteoblastic cells by studying the responses or changes in intracellular Ca 2+ level to P2 receptor agonists, antagonists and modulators. ATP induced robust responses in a concentration-dependent manner with EC 50 of 0.5 ± 0.07 μM. While α,β-methylene-ATP (αβmeATP) and 2',3'-O-(4-benzoylbenzoyl)-ATP (BzATP) were ineffective, ADP mimicked the action of ATP with EC 50 of 0.7 ± 0.2 μM. UTP and UDP also evoked responses with EC 50 of 2.0 ± 0.4 μM and 0.5 ± 0.1 μM respectively, but their responses were much smaller, resulting in an order of the response magnitude: ATP ~ ADP >> UTP ~ UDP. The responses evoked by ATP and ADP were blocked by pyridoxal-5'-phosphate-6-azophenyl-2,4,-disulfonate (PPADS) with IC 50 of 3.0 ± 0.05 μM and 5.0 ± 0.4 μM respectively, but not by suramin up to 30 μM. ATP-evoked responses were insensitive to inhibition by trinitrophenyl-ATP (TNP-ATP) and brilliant blue G. ADP-evoked responses were significantly inhibited by 2'-deoxy- N 6-methyladenosine-3',5'-biphosphate (MRS2179) and 2-chloro- N 6-methyl-( N)-methanocarba-2'-deoxyadenosine-3',5'-bisphosphate (MRS2279) with IC 50 of 48 ± 1.9 μM and 7.7 ± 0.9 μM respectively. Taken together, these results provide strong evidence for functional expression of ATP-sensitive P2Y receptors and particularly P2Y 1-like receptor in G292 cells.