Dopamine D₁ and adenosine A₁ receptors form functionally interacting heteromeric complexes

Abstract

The possible molecular basis for the previously described antagonistic interactions between adenosine A₁ receptors (A₁R) and dopamine D₁ receptors (D₁R) in the brain have been studied in mouse fibroblast Ltk⁻ cells cotransfected with human A₁R and D₁R cDNAs or with human A₁R and dopamine D₂ receptor (long-form) (D₂R) cDNAs and in cortical neurons in culture. A₁R and D₁R, but not A₁R and D₂R, were found to coimmunoprecipitate in cotransfected fibroblasts. This selective A₁R/D₁R heteromerization disappeared after pretreatment with the D₁R agonist, but not after combined pretreatment with D₁R and A₁R agonists. A high degree of A₁R and D₁R colocalization, demonstrated in double immunofluorescence experiments with confocal laser microscopy, was found in both cotransfected fibroblast cells and cortical neurons in culture. On the other hand, a low degree of A₁R and D₂R colocalization was observed in cotransfected fibroblasts. Pretreatment with the A₁R agonist caused coclustering (coaggregation) of A₁R and D₁R, which was blocked by combined pretreatment with the D₁R and A₁R agonists in both fibroblast cells and in cortical neurons in culture. Combined pretreatment with D₁R and A₁R agonists, but not with either one alone, substantially reduced the D₁R agonist-induced accumulation of cAMP. The A₁R/D₁R heteromerization may be one molecular basis for the demonstrated antagonistic modulation of A₁R of D₁R receptor signaling in the brain. The persistence of A₁R/D₁R heteromerization seems to be essential for the blockade of A₁R agonist-induced A₁R/D₁R coclustering and for the desensitization of the D₁R agonist-induced cAMP accumulation seen on combined pretreatment with D₁R and A₁R agonists, which indicates a potential role of A₁R/D₁R heteromers also in desensitization mechanisms and receptor trafficking.