Abstract

A range of ligands displayed agonism at the long isoform of the human dopamine D₂ receptor, whether using receptor-G protein fusions or membranes of cells in which pertussis toxin-resistant mutants of individual Gα_i-family G proteins could be expressed in an inducible fashion. Varying degrees of efficacy were observed for individual ligands as monitored by their capacity to load [³⁵S]GTPγS onto each of Gα_i1,Gα_i2,Gα_i3, and Gα_o1. By contrast, (S)-(–)-3-(3-hydroxyphenyl)-N-propylpiperidine was a partial agonist when Gα_o1 was the target G protein but an antagonist/inverse agonist at Gα_i1,Gα_i2, and Gα_i3. In ligand binding assays, dopamine identified both high- and low-affinity states at each of the dopamine D₂ receptor-G protein fusion proteins, and the high-affinity state was eliminated by guanine nucleotide. (S)-(–)-3-(3-Hydroxyphenyl)-N-propylpiperidine bound to an apparent single state of the constructs in which the D₂ receptor was fused to Gα_i1,Gα_i2, or Gα_i3. However, it bound to distinct high- and low-affinity states of the D₂ receptor-Gα_o1 fusion, with the high-affinity state being eliminated by guanine nucleotide. Likewise, although dopamine identified guanine nucleotide-sensitive high-affinity states of the D₂ receptor when expression of pertussis toxin-resistant forms of each of Gα_i1, Gα_i2, Gα_i3, and Gα_o1 was induced, (S)-(–)-3-(3-hydroxyphenyl)-N-propylpiperidine identified a high-affinity site only in the presence of Gα_o1. p-Tyramine displayed a protean ligand profile similar to that of (S)-(–)-3-(3-hydroxyphenyl)-N-propylpiperidine but with lower potency. These results demonstrate (S)-(–)-3-(3-hydroxyphenyl)-N-propylpiperidine to be a protean agonist at the D₂ receptor and may explain in vivo actions of this ligand.