Acute Cocaine Enhances Dopamine D₂R Recognition and Signaling and Counteracts D₂R Internalization in Sigma1R-D₂R Heteroreceptor Complexes

The current study was performed to establish the actions of nanomolar concentrations of cocaine, not blocking the dopamine transporter, on dopamine D2 receptor (D₂R)-sigma 1 receptor (δ1R) heteroreceptor complexes and the D₂R protomer recognition, signaling and internalization in cellular models. We report the existence of D₂R-δ1R heteroreceptor complexes in subcortical limbic areas as well as the dorsal striatum, with different distribution patterns using the in situ proximity ligation assay. Also, through BRET, these heteromers were demonstrated in HEK293 cells. Furthermore, saturation binding assay demonstrated that in membrane preparations of HEK293 cells coexpressing D₂R and δ1R, cocaine (1 nM) significantly increased the D₂R B_max values over cells singly expressing D₂R. CREB reporter luc-gene assay indicated that coexpressed δ1R significantly reduced the potency of the D₂R-like agonist quinpirole to inhibit via D₂R activation the forskolin induced increase of the CREB signal. In contrast, the addition of 100 nM cocaine was found to markedly increase the quinpirole potency to inhibit the forskolin-induced increase of the CREB signal in the D₂R-δ1R cells. These events were associated with a marked reduction of cocaine-induced internalization of D₂R protomers in D₂R-δ1R heteromer-containing cells vs D₂R singly expressing cells as studied by means of confocal analysis of D₂R-δ1R trafficking and internalization. Overall, the formation of D₂R-δ1R heteromers enhanced the ability of cocaine to increase the D₂R protomer function associated with a marked reduction of its internalization. The existence of D₂R-δ1R heteromers opens up a new understanding of the acute actions of cocaine.