Functional histamine H₃ and adenosine A_2A receptor heteromers in recombinant cells and rat striatum

In the striatum, histamine H₃ receptors (H₃Rs) are co-expressed with adenosine A_2A receptors (A_2ARs) in the cortico-striatal glutamatergic afferents and the GABAergic medium-sized spiny neurons that originate the indirect pathway of the basal ganglia. This location allows H₃Rs and A_2ARs to regulate the striatal GABAergic and glutamatergic transmission. However, whether these receptors can physically interact has not yet been assessed. To test this hypothesis, a heteromer-selective in vitro assay was used to detect functional complementation between a chimeric A_2AR₃₀₂-Gα_qi4 and wild-type H₃Rs in transfected HEK-293T cells. H₃R activation with the agonist RAMH resulted in Ca²⁺ mobilization (pEC₅₀ 7.31 ± 0.23; maximal stimulation, Emax 449 ± 25% of basal) indicative of receptor heterodimerization. Functional H₃R-A_2AR heteromers were confirmed by co-immunoprecipitation and observations of differential cAMP signaling when both receptors were co-expressed in the same cells. In membranes from rat striatal synaptosomes, H₃R activation decreased A_2AR affinity for the agonist CGS-21680 (pKi values 8.10 ± 0.04 and 7.70 ± 0.04). Moreover, H₃Rs and A_2ARs co-immunoprecipitated in protein extracts from striatal synaptosomes. These results support the existence of a H₃R-A_2AR heteromer with possible physiological implications for the modulation of the intra-striatal transmission.