Ligand-Induced Regulation and Localization of Cannabinoid CB1 and Dopamine D2L Receptor Heterodimers

The cannabinoid CB1 (CB1) and dopamine D-2 (D-2) receptors are coexpressed in the basal ganglia, an area of the brain involved in such processes as cognition, motor function, and emotional control. Several lines of evidence suggest that CB1 and D-2 receptors may oligomerize, providing a unique pharmacology in vitro and in vivo. However, limited information exists on the regulation of CB1 and D-2 receptor dimers. We used a novel technique, multicolor bimolecular fluorescence complementation (MBiFC) to examine the subcellular localization of CB1-D-2L heterodimers as well as D-2L-D-2L homodimers in a neuronal cell model, Cath. a differentiated cells. MBiFC was then used to explore the effects of persistent ligand treatment on receptor dimerization at the plasma membrane and intracellularly. Persistent (20-h) agonist treatment resulted in increased formation of CB1-D-2L heterodimers relative to the D-2L-D-2L homodimers. The effects of the D-2 agonist quinpirole were restricted to the intracellular compartment and may reflect increased D-2L receptor expression. In contrast, treatment with the CB1 receptor agonist (2)-cis-3-[2-hydroxy-4-(1,1-dimethylheptyl) phenyl]-trans-4-(3-hydroxypropyl) cyclohexanol (CP55, 940) produced increases in both membrane and intracellular CB1 D-2L heterodimers independently of alterations in CB1 receptor expression. The effects of CB1 receptor activation were attenuated by the CB1 antagonist 1-(2,4-dichlorophenyl)-5-(4-iodophenyl)-4-methyl-N-4-morpholinyl-1H-pyrazole-3-carboxamide (AM281) and were both time-and dose-dependent. The effects of CB1 activation were examined further by combining MBiFC with a constitutively active CB1 receptor mutant, CB1 T210I. These studies demonstrated that the expression of CB1 T210I increased intracellular CB1-D-2L heterodimer formation. In summary, agonist-induced modulation of CB1-D-2L oligomerization may have physiological implications in diseases such as Parkinson's disease and drug abuse.