Cloning and expression of the A 2a adenosine receptor from guinea pig brain

Abstract

A full-length complementary DNA (cDNA) clone encoding the guinea pig brain A 2 adenosine receptor has been isolated by polymerase chain reaction (PCR) and low-stringency-hybridization screening of a guinea pig brain cDNA library. This cDNA contains a long open reading frame encoding a 409-amino acid-residue protein which is highly homologous to the A 2 adenosine receptors previously cloned from other species. Hydrophobicity analysis of the deduced protein sequence reveals seven hydrophobic regions, characteristic of a member of the G-protein-coupled receptor superfamily. Radioligand binding assay and functional (GTPase and cAMP) assays of the receptor, transiently expressed in mammalian cells, demonstrate typical characteristics of the A 2 type adenosine receptor. The messenger RNA (mRNA) of this A 2 receptor is found in the brain, heart, kidney and spleen. Receptor autoradiography with [ 3 H]CGS21680, a specific A 2 agonist, and in situ hybridization with A 2 cRNA probe in guinea pig brain indicate that the receptor is expressed exclusively in the caudate nucleus. The pharmacological profile and anatomical distribution of this receptor indicate that it is of the A 2a subtype. This work represents the first cloning of an A 2a receptor in a rodent species, offers a complete pharmacological characterization of the receptor and provides an anatomical comparison between binding profile and gene expression of the receptor.