Dopamine D1 and adenosine A1 receptors form functionally interacting heteromeric complexes

The possible molecular basis for the previously described antagonistic interactions between adenosine A(1) receptors (A(1)R) and dopamine D-1 receptors (D1R) in the brain have been studied in mouse fibroblast Ltk(-) cells cotransfected with human A(1)R and D1R cDNAs or with human A(1)R and dopamine D-2 receptor (long-form) (D2R) cDNAs and in cortical neurons in culture. A(1)R and D1R, but not A(1)R and D2R, were found to coimmunoprecipitate in cotransfected fibroblasts. This selective A(1)R/D1R heteromerization disappeared after pretreatment with the D1R agonist, but not after combined pretreatment with D1R and A(1)R agonists. A high degree of A(1)R 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 A(1)R and D2R colocalization was observed in cotransfected fibroblasts. Pretreatment with the A(1)R agonist caused coclustering (coaggregation) of A(1)R and D1R. which was blocked by combined pretreatment with the D1R and A(1)R agonists in both fibroblast cells and in cortical neurons in culture. Combined pretreatment with D1R and A(1)R agonists, but not with either one alone, substantially reduced the D1R agonist-induced accumulation of cAMP. The A(1)R/D1R heteromerization may be one molecular basis for the demonstrated antagonistic modulation of A(1)R of D1R receptor signaling in the brain. The persistence of A(1)R/D1R heteromerization seems to be essential for the blockade of A(1)R agonist-induced A(1)R/D1R coclustering and for the desensitization of the D1R agonist-induced cAMP accumulation seen on combined pretreatment with D1R and A(1)R agonists, which indicates a potential role of A(1)R/D1R heteromers also in desensitization mechanisms and receptor trafficking.