Functional Coupling of Human D₂, D₃, and D₄ Dopamine Receptors in HEK293 Cells

Abstract

The D₂ dopamine receptor is known to be functionally coupled to the inhibition of adenylate cyclase when expressed in a number of mammalian cell lines. However, functional coupling of the recently discovered D₃ and D₄ dopamine receptor subtypes has been more difficult to demonstrate. In this study, human D₂, D₃ and D₄ receptors were stably expressed separately in human embryonic kidney cells (HEK 293). In these cells, activation of D₂, D₃ or D₄ receptors resulted in the inhibition of forskolin-stirnulated adenylate cyclase activity in a dose responsive manner. This activation was prevented by pre-incubation of the cells expressing these receptors with the dopaminergic antagonist haloperidol. Radioligand binding studies using [³H]spiperone confirmed that the atypical neuroleptic clozapine has higher affinity for the human D₄ receptor than the D₃ or D₄ receptors, although only 6-fold higher than the D₂ receptor in this study. In addition, ribonuclease protection studies demonstrated the presence of D₄ dopamine receptor mRNA in human brain regions.