Evidence for a G protein-coupled diadenosine-5 ',5 '''-P-1,P-4-tetraphosphate (Ap(4)A) receptor binding site in lung membranes from rat

Nucleotide receptors are of considerable importance in the treatment of lun g diseases, such as cystic fibrosis. Because diadenosine polyphosphates may also be of significance as signalling molecules in lung, as they are in a variety of tissues, in the present work we investigated the binding sites f or [H-3]diadenosine-5',5"'-P-1, P-4-tetraphosphate (Ap(4)A) in plasma membr anes from rat lung and studied their possible coupling to G proteins. We pr esent evidence for a single high-affinity binding site for [H-3]Ap(4)A with similar affinity for other diadenosine polyphosphates Ap(n)A (n = 2 to 6). Displacement studies with different nucleotides revealed that the [H-3]Ap( 4)A binding site was different from P2X and P2Y(2) receptor binding sites. Pretreatment of lung membranes with GTP gamma S or GTP in the presence of M g2+ increased the K-i for Ap(4)A from 91 nM to 5.1 mu M, which is indicativ e of G protein coupling. The putative coupling to G proteins was further co nfirmed by the enhancement of [S-35]GTP gamma S binding (to G(alpha) protei ns) to lung membranes by Ap(4)A (63% increase over basal) in a concentratio n-dependent manner. Therefore, our data for the first time provide evidence of a G protein-coupled Ap(4)A binding site in lung membranes. (C) 1999 Els evier Science B.V. All rights reserved.