Of the five known dopamine receptors, DlA and D2 represent the major subtypes expressed in the striatum of the adult brain. Within the striatum, these two subtypes are differentially distributed in the two main neuronal populations that provide direct and indirect pathways between the striatum and the output nuclei ofthe basal ganglia. Movement The pivotal role played by dopamine receptors in the pathophysiology and treatment ofParkinson disease (1) and schizophrenia (2) and in the mode of action of addictive drugs such as amphetamine and cocaine (3, 4) is well established. Of the five known dopamine receptor subtypes (5), the DlA and D2 receptors account for the vast majority of dopamine receptors (6) expressed in the striatum. The DlA (also known as D, in the primate system) and D2 receptor subtypes are expressed mainly by spiny projection neurons, which account for 90-95% of the striatal neuron population (7). These striatal neurons may be subdivided into two major types on the basis of their axonal projections. One type provides a direct projection to the output nuclei of the basal ganglia: the substantia nigra and entopeduncular nucleus (the internal segment of the globus pallidus in primates). The other type provides projections to the globus pallidus (the external segment of the primate globus pallidus). As this latter type is connected indirectly to the output nuclei of the basal ganglia through connections with the subthalamic nucleus, the two output pathways are referred to as the direct and indirect output systems. Striatal neurons giving rise to the direct pathway express high levels of the DlA dopamine receptor subtype and the neuropeptides substance P and dynorphin, whereas neurons giving rise to the indirect pathway express high levels of the D2 dopamine receptor and the peptide enkephalin (7). The levels of peptide expression in these neurons provide an assay oftheir activity (7), as neuropeptide levels correlated with fuing rates in target neurons (1).Current models suggest that imbalanced activity in the direct and indirect pathways is responsible for clinical movement disorders (8). A number of studies have demonstrated that dopamine oppositely effects these two output pathways through their differential expression ofthe DlA and D2 receptors (7). For example, depletion of striatal dopamine with lesions of the nigrostriatal dopamine pathway in animal models of Parkinson disease results in reduced expression of substance P in direct output neurons and increased enkephalin expression in indirect striatal output neurons. Moreover, these changes may be selectively reversed with selective dopamine receptor agonist treatments, so that D1 agonist treatment normalizes substance P levels whereas D2 agonist treatment normalizes enkephalin levels (9). While these studies have demonstrated the differential role of DlA and D2 receptors in striatal function, important questions concerning the interaction between these neuronal pathways remain. To provide an experimental animal model to address the...

ABSTRACTderived from the 129/sv strain (Clontech) was screened using a mouse D3 cDNA probe (17). A positive clone encompassing exon 2 of the murine D3 gene was isolated and further characterized. A 7-kb Xho I-Asp718 fragment was engineered for targeted mutagenesis by introducing the GKNeo cassette (16) in antisense orientation at the Sal I site in exon 2 (17). Integration of sequences derived from the pGKNeo cassette generates a novel open reading frame, resulting in the following peptide sequence appended after Arg-148: PASDGIRT-WQNNTENEVYVEQRLLISFFRL Opal (Stop). The sequence of the mutant allele was confirmed by direct sequencing of reverse transcription-PCR (rPCR) products derived from brain mRNAs of -/-and +/-mice (data not shown).Transfection ofES Cells and Embryo Manipulations. J-1 ES cells (a kind gift of R. Jaenisch, Massachussetts Institute of Technology) at passage 13 were grown on mitomycin C-treated embryonic fibroblasts derived from a homozygous neomycin (Neo)-resistant transgenic mouse (16). Cells (2 x 107) were electroporated in a 1-ml cuvette (path length-0.2 cm) at 0.4 kV and 25 ,uF. Cells were plated onto 40 gelatin-coated Petri dishes (6 cm) on embryonic feeder cells. Selection with G418 (0.3 mg/ml; active concentration of 0.66 ,vg/mg of dry powder; GIBCO) was applied 24 hr after plating and was continued for 7-9 days. Individual Neo-resistant colonies were picked using a dissection microscope and expanded as described (16). Genomic DNA was prepared from an aliquot of cells for each clone using previously described techniques and analyzed by Southern blotting (18). Recovery, microinjection, and transfer of 3.5 day postcoitus embryos was performed as described (16).

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We describe the cloning and characterization of a human 5‐HT6 serotonin receptor. The open reading frame is interrupted by two introns in positions corresponding to the third cytoplasmic loop and the third extracellular loop. The human 5‐HT6 cDNA encodes a 440‐amino‐acid polypeptide whose sequence diverges significantly from that published for the rat 5‐HT6 receptor. Resequencing of the rat cDNA revealed a sequencing error producing a frame shift within the open reading frame. The human 5‐HT6 amino acid sequence is 89% similar to the corrected rat sequence. The recombinant human 5‐HT6 receptor is positively coupled to adenylyl cyclase and has pharmacological properties similar to the rat receptor with high affinity for several typical and atypical antipsychotics, including clozapine. The receptor is expressed in several human brain regions, most prominently in the caudate nucleus. The gene for the receptor maps to the human chromosome region 1p35–p36. This localization overlaps that established for the serotonin 5‐HT1Dα receptor, suggesting that these may be closely linked. Comparison of genomic and cDNA clones for the human 5‐HT6 receptor also reveals an RsaI restriction fragment length polymorphism within the coding region.