Dopamine D₂ receptors (D₂R) are known to form transient homodimer complexes, of which the increased formation has already been associated with development of schizophrenia. Pharmacological targeting and modulation of the equilibrium of these receptor homodimers might lead to a better understanding of the critical role played by these complexes in physiological and pathological conditions. Whereas agonist addition has shown to prolong the D₂R dimer lifetime and increase the level of dimer formation, the possible influence of D₂R antagonists on dimerization has remained rather unexplored. Here, using a live-cell reporter assay based on the functional complementation of a split Nanoluciferase, a panel of six D₂R antagonists were screened for their ability to modulate the level of D2LR dimer formation. Incubation with the D₂R antagonist spiperone decreased the level of D2LR dimer formation significantly by 40-60% in real-time and after long-term (≥16 h) incubations. The fact that dimer formation of the well-studied A2a-D2LR dimer was not altered following incubation with spiperone supports the specificity of this observation. Other D₂R antagonists, such as clozapine, risperidone, and droperidol did not significantly evoke this dissociation event. Furthermore, molecular modeling reveals that spiperone presents specific Tyr1995.48 and Phe3906.52 conformations, compared to clozapine, which may determine D₂R homodimerization.
Keywords: G protein-coupled receptor (GPCR); NanoLuc binary technology (NanoBiT); dimerization; dopamine D2 receptor; oligomerization; protein complementation assay.