1. Using the gene splicing technique a synthetic human dopamine (DA) D4.2 gene was constructed and subsequently stably expressed in CHO K1 cells. 2. Binding of [3H]spiperone to membranes prepared from human DA D4.2 CHO K1 cells was saturable with a Kd of 93 +/- 0.51 pM and a Bmax of 768 +/- 22 fmol per mg protein. 3. Clozapine, apomorphine, and S(+)-NPA were more selective for D4.2 than for D2L receptors, with D2L/D4.2 ratios of 5.7, 7.1, and 19.6, respectively. 4. Functional studies indicated that DA D4.2 receptors expressed in CHO K1 cells inhibited forskolin stimulated cAMP levels showing coupling to G-proteins. 5. Two reciprocal human D2L and D4.2 chimeric receptors (D2L/D4.2 and D4.2/D2L) were constructed by exchanging the amino-terminal end to the third transmembrane (TM) of one receptor with the counter part of the other receptor and expressing them transiently into COS-7 cells. The chimeric D2L/D4.2 receptor displayed non-detectable specific binding of [3H] spiperone and other ligands. The chimeric D4.2/D2L receptor binding affinities of DA agonists were more affected than that of antagonists, suggesting that binding affinities of agonists are more sensitive to changes in receptor conformation than that of antagonists. 6. This study characterized the pharmacology of a novel synthesized DA D4.2 receptor that provides a useful model for screening of potential D4.2 receptor agonist and antagonist.