Chronic treatment of rats with R-PIA in vivo desensitized adenosine A 1 receptor-mediated inhibition of adenylyl cyclase in brain plasma membranes and increased basal and forskolin-stimulated adenylyl cyclase. The adenosine A 1 receptor agonist CHA (cyclohexyl adenosine) inhibited forskolin-stimulated adenylyl cyclase in synaptic plasma membranes from control rats but failed to do so in membranes isolated from rats treated with R-PIA. This loss of response was accompanied with a significant decrease in both, total number of adenosine A 1 receptors and steady-state level of α-G i in synaptic plasma membranes. An increase in the steady-state level of α-G s in synaptic plasma membranes was also observed by R-PIA treatment. Concurrently, a significant increase of adenosine A 1 receptors was observed in microsomes and coated vesicles. These results demonstrate adenosine A 1 receptor desensitization in rat brain by in vivo treatment with R-PIA and suggest a role for coated vesicles in the internalization of G-protein coupled receptors.