Serum constituents can affect 2 '-& 3 '-O-(4-benzoylbenzoyl)-ATP potency at P2X(7) receptors

1 2 '-& 3 ' -O-(4-benzoylbenzoyl)-ATP (BzATP) is the prototypic agonist for P2X(7) receptors. In this study we demonstrate that bovine serum albumin ( BSA) can affect the potency of BzATP at P2X receptors. 2 BzATP potency (pEC(50)) to stimulate ethidium accumulation in cells expre ssing recombinant P2X7 receptors varied between 6.5 and 4, depending upon t he species orthologue studied and ionic conditions employed. BSA (0.1-1 mg ml(-1)) and foetal bovine serum (FBS, 1-10% v v(-1)) inhibited responses to BzATP but only when the BzATP pEC(50) exceeded 5. 3 BSA did not block ATP-stimulated ethidium accumulation, suggesting its ef fects were independent of P2X(7) receptor blockade. 4 BSA did not cause breakdown of nucleotides, although FBS (10% v v(-1)) ex hibited appreciable nucleotidase activity and caused significant breakdown of ATP. 5 In the presence of BSA, lipids such as 11-((5-dimethylaminonaphthalene-1- sulphonyl)amino)undecanoic acid (DAUDA) and arachidonic acid (AA) markedly increased BzATP potency. Lipids had no affect on ATP potency in the presenc e of BSA and had little effect on responses to BzATP in the absence of BSA. 6 These results suggested that the reduction in BzATP potency by BSA was du e to BzATP binding to BSA and that lipids prevented this binding. Consisten t with this hypothesis, BzATP inhibited binding of the fluorescent lipid, D AUDA, to BSA. 7 In conclusion, BSA and lipids can markedly affect BzATP potency at P2X(7) receptors but this is probably a consequence of BzATP binding to BSA. This finding has important implications when using BzATP in vivo or in the pres ence of albumin.