[H-3]-SCH 58261 LABELING OF FUNCTIONAL A(2A) ADENOSINE RECEPTORS IN HUMAN NEUTROPHIL MEMBRANES

1 The present study describes the direct labelling of A(2A) adenosine receptors in human neutrophil membranes with the potent and selective antagonist radioligand, -(2-phenylethyl)-2-(2-furyl)-pyrazolo[4,3-e]-1 ,2,4 triazolo[1,5-c]pyrimidine, ([H-3]-SCH 58261). In addition, both r eceptor affinity and potency of a number of adenosine receptor agonist s and antagonists were determined in binding, adenylyl cyclase and sup eroxide anion production assays. 2 Saturation experiments revealed a s ingle class of binding sites with K-d and B-max values of 1.34 nM and 75 fmol mg(-1) protein, respectively. Adenosine receptor ligands compe ted for the binding of 1 nM [H-3]-SCH 58261 to human neutrophil membra nes, with a rank order of potency consistent with that typically found for interactions with the A(2A) adenosine receptors. In the adenylyl cyclase and in the superoxide anion production assays the same compoun ds exhibited a rank order of potency identical to that observed in bin ding experiments. 3 Thermodynamic data indicated that [H-3]-SCH 58261 binding to human neutrophils is entropy and enthalpy-driven. This find ing is in agreement with the thermodynamic behaviour of antagonists bi nding to rat striatal A(2A) adenosine receptors. 4 It was concluded th at in human neutrophil membranes, [H-3]-SCH 58261 directly labels bind ing sites with pharmacological properties similar to those of A(2A) ad enosine receptors of other tissues. The receptors labelled by [H-3]-SC H 58261 mediated the effects of adenosine and adenosine receptor agoni sts to stimulate cyclic AMP accumulation and inhibition of superoxide anion production in human neutrophils.