Cannabinoid receptor interactions with the antagonists SR 141716A and SR 144528

The G protein-coupled cannabinoid receptor subtypes CB1 and CB2 have been c loned from several species. The CB1 receptor is highly conserved across spe cies, whereas the CB2 receptor shows considerable cross-species variations. The two human receptors share only 44% overall identity, ranging from 35% to 82% in the transmembrane regions. Despite this structural disparity, the most potent cannabinoid agonists currently available are largely undiscrim inating and are therefore unsatisfactory tools for investigating the archit ecture of ligand binding sites. However, the availability of two highly spe cific antagonists, SR 141716A for the CB1 receptor and SR 144528 for the CB 2 receptor, has allowed us to adopt a systematic approach to defining their respective binding sites through the use of chimeric CB1 receptor/CB2 rece ptor constructs, coupled with site-directed mutagenesis. We identified the region encompassed by the fourth and fifth transmembrane helices as being c ritical for antagonist specificity. Both the wild type human receptors over expressed in heterologous systems are autoactivated; SR 141716A and SR 1445 28 exhibit classical inverse agonist properties with their respective targe t receptors. In addition, through its interaction with the CB1 receptor SR 141716A blocks the G(i) protein-mediated activation of mitogen-activated pr otein kinase stimulated by insulin or insulinlike growth factor I. An in-de pth analysis of this discovery has led to a modified three-state model for the CB1 receptor, one of which implicates the SR 141716A-mediated sequestra tion of G(i) proteins, with the result that the growth factor-stimulated in tracellular pathways are effectively impeded.