Multiple DI dopaminergic receptor subtypes have been postulated on the basis of pharmacological, biochemical, and genetic studies. We describe the isolation and characterization of a rat gene encoding a dopamine receptor that is structurally and functionally similar to the D1 dopamine receptor. The coding region, which is intronless, encodes a protein of 475 amino acids (Mr 52,834) with structural features that are consistent with receptors coupled to guanine nucleotide-binding regulatory proteins. The expressed protein binds dopaminergic ligands and mediates stimulation of adenylyl cyclase with pharmacological properties similar to those of the DI dopamine receptor. The gene encoding the human homologue of this receptor subtype is located to the short arm of chromosome 4 (4p16.3), the same region as the Huntington disease gene. In striking contrast to the previously cloned DI receptor, little or no mRNA for the receptor described here was observed in striatum, nucleus accumbens, olfactory tubercle, and frontal cortex. High levels of mRNA for this receptor were found in distinct layers of the hippocampus, the mammillary nuclei, and the anterior pretectal nuclei, brain regions that have been shown to exhibit little or no DI dopamine receptor binding. On the basis of its properties we propose that this dopamine receptor subtype be called DIB1The actions of dopamine were originally thought to be mediated by an interaction with two distinct receptor subtypes: D1 receptors, which were coupled to the stimulation of adenylyl cyclase, and D2 receptors, which were either linked or not linked to the inhibition of adenylyl cyclase (1). Recently, molecular biological approaches have supported and extended this pharmacological classification. At least three different receptor genes code for dopamine receptor subtypes-namely, D1, D2, and D3 (2-9). These receptors belong to the large family of receptors coupled to guanine nucleotide-binding regulatory protein (G protein) and are believed to contain seven membrane-spanning domains (10).Traditionally, the central actions of dopaminergic compounds have been attributed to their interactions with D2 dopamine receptors (11). However, recent studies have indicated that D1 dopamine receptors have important functions in the central nervous system (12). The distribution of D1 receptors in brain has been studied by using the selective ligand SCH 23390. Although this ligand binds to a single class ofreceptor sites, recent evidence has suggested the existence of multiple D1 receptor subtypes (13). For instance, it has been demonstrated (14) that injection of rat striatal mRNA into Xenopus oocytes directs the expression of a D1 dopamine receptor coupled to activation of phospholipase C and phosphatidylinositol phosphate metabolism. Furthermore, dopamine does not stimulate adenylyl cyclase in the amygdala, a tissue known to contain specific binding sites for the radiolabeled Dl-selective antagonist SCT{ 23390 (13). In the periphery, D1 receptors have been shown to stimulate adenyly...

The sub-family of dopamine D1-like receptors is now known to be comprised of at least two members: the originally cloned D1 receptor (herein referred to as the D1a receptor) and a related receptor referred to as the D1b, D1 beta or D5 dopamine receptor (herein referred to as the D1b/D5 receptor). Here, we characterize the D1b/D5 receptor expressed transiently in COS-7 cells and permanently in Ltk- cells. Transiently expressed human D1b/D5 receptors bind the D1 specific ligand [125I]SCH 23982 saturably and with high affinity (KD = 500 pM). Competition for [125I]SCH 23982 binding to rat D1b/D5 and human D1a and D1b/D5 receptors supports the contention that the two D1b/D5 receptors are species homologues. Furthermore, in COS-7 cells, as previously observed, dopamine competes for the binding of [125I]SCH 23982 to human D1b/D5 receptors with a higher affinity than that seen at the human D1a receptor. These results are similar to those seen in Ltk- cells permanently transfected with the human D1b/D5 receptor. In these cells, dopamine competition for [125I]SCH 23982 binding is complex, sensitive to guanine nucleotides and of a higher affinity than that observed for dopamine binding to the human D1a receptor expressed in these same cells. In both D1a and D1b/D5 expressing Ltk- cells, dopamine stimulates adenylyl cyclase with an EC50 of approximately 200 nM. Furthermore, preincubation of Ltk- cells expressing the D1a and D1b/D5 receptors with dopamine results in desensitization of the response of adenylyl cyclase to subsequent agonist stimulation.