Signaling through Homomeric and Heteromeric Dopamine D2 and Cannabinoid Cb1 Receptors

Abstract

G protein-coupled receptors (GPCRs) play important roles in signal transduction and represent about half of the drug targets. Accumulating evidence suggests that GPCRs exist and function not only as homomers but as heteromers as well. Coimmunoprecipitation and fluorescence resonance energy transfer (FRET) results have provided evidence for heteromerization of D2-CB1 receptors in co-transfected cell lines. Besides, co-localization of CB1 and D2 receptors in the somata and dendrites of striato-pallidal GABA neurons and cortical-striatal glutamate terminals has been demonstrated. Studies have shown that homomeric D2 and CB1 receptors are Gi/o coupled able to inhibit upon stimulation the forskolin-induced adenylyl-cyclase activation leading to a decrease in cAMP levels. However, stimulation of the CB1 receptor upon the co-expression of D2-CB1 receptors in cell lines increased cAMP levels presumably because CB1 receptors switch their specificity to becoming Gs-coupled when complexed with the D2 receptor. In this study, we utilized Xenopus laevis oocytes as a heterologous expression system in which G protein-sensitive inwardly rectifying K+ (GIRK) channels are co-expressed with GPCRs. Stimulation of GPCRs with specific ligands activates GIRK channels via the betagamma subunits of the G proteins (Gbg). The GIRK channel current is monitored by two-electrode voltage clamp (TEVC) to access receptor activity. We have found that stimulation of homomeric D2 and CB1 receptors increases the GIRK currents. This increase could be blocked by pertussis toxin (PTX) that specifically ADP-ribosylates Gi/o. Overexpression of Gs in Xenopus oocytes biases signaling to couple Gs coupled receptors to GIRK channels via Gbg. Using this approach, we demonstrated that neither the CB1 nor the D2 homomeric receptors can couple to Gs. We are in the process of testing whether the channel response of CB1 receptor ligands through the heteromeric D2-CB1 receptors proceeds via Gs.