title: Pharmacological characterization of P2X1 and P2X3 purinergic receptors in bovine chondrocytes.
abstract: OBJECTIVE
  The aim of the present study is that of characterizing, for the first time in a quantitative way, from a biochemical, physico chemical and functional point of view P2X(1) and P2X(3) purinergic receptors in bovine chondrocytes. The affinity and the potency of typical purinergic ligands were studied through competition binding experiments and their role in modulating chondrocyte actvities was investigated by analyzing nitric oxide (NO) and prostaglandin E2 (PGE(2)) release.
  
  
  METHODS
  Saturation, competition binding experiments, western blotting and immunohistochemistry assays on the P2X(1) and P2X(3) purinergic receptors in bovine chondrocytes were performed. Thermodynamic analysis of the P2X(1) and P2X(3) purinergic binding was studied to investigate the forces driving drug-receptor coupling. In the functional assays (NO and PGE(2) release) the potency of purinergic agonists and antagonists was evaluated.
  
  
  RESULTS
  Bovine chondrocytes expressed P2X(1) and P2X(3) purinergic receptors and thermodynamic parameters indicated that purinergic binding is enthalpy- and entropy-driven for agonists and totally entropy-driven for antagonists. Typical purinergic agonists such as adenosine 5'-triphosphate (ATP) and alpha,beta-methyleneATP were able to increase NO and PGE(2) release. A purinergic antagonist, A317491, was able to block the stimulatory effect on functional experiments mediated by the agonists.
  
  
  CONCLUSIONS
  These data demonstrate for the first time the presence of functional P2X(1) and P2X(3) purinergic receptors in bovine chondrocytes. Agonists and antagonists are thermodynamically discriminated and are able to modulate functional responses such as NO and PGE(2) release. These results suggest the potential role of novel purinergic antagonists in the treatment of pathophysiological diseases linked to the inflammation and involved in articular cartilage resorption.
authors.affiliation: Department of Clinical and Experimental Medicine, University of Ferrara, Italy.
authors.forename: K
authors.initials: K
authors.lastname: Varani
authors.forename: M
authors.initials: M
authors.lastname: De Mattei
authors.forename: F
authors.initials: F
authors.lastname: Vincenzi
authors.forename: A
authors.initials: A
authors.lastname: Tosi
authors.forename: S
authors.initials: S
authors.lastname: Gessi
authors.forename: S
authors.initials: S
authors.lastname: Merighi
authors.forename: A
authors.initials: A
authors.lastname: Pellati
authors.forename: F
authors.initials: F
authors.lastname: Masieri
authors.forename: A
authors.initials: A
authors.lastname: Ongaro
authors.forename: P A
authors.initials: PA
authors.lastname: Borea
mesh_terms: D000818:Animals; D001667:Binding, Competitive; D015415:Biomarkers / Q000378:metabolism; D002417:Cattle; D002478:Cells, Cultured; D019902:Chondrocytes / Q000378:metabolism*; D015232:Dinoprostone / Q000378:metabolism*; D009569:Nitric Oxide / Q000378:metabolism*; D018048:Receptors, Purinergic P2 / Q000378:metabolism*; D058469:Receptors, Purinergic P2X; D058477:Receptors, Purinergic P2X3
chemical_list: D015415:Biomarkers; D018048:Receptors, Purinergic P2; D058469:Receptors, Purinergic P2X; D058477:Receptors, Purinergic P2X3; D009569:Nitric Oxide; D015232:Dinoprostone