Expression of dopamine D₃ receptor dimers and tetramers in brain and in transfected cells

The expression and characteristics of the dopamine D₃ receptor protein were studied in brain and in stably transfected GH3 cells. Monoclonal antibodies were used for immunoprecipitation and immunoblot experiments. Immunoprecipitates obtained from primate and rodent brain tissues contain a low molecular weight D₃ protein and one or two larger protein species whose molecular mass are integral multiples of the low molecular weight protein and thus appear to have resulted from dimerization and tetramerization of a D₃ monomer. Whereas D₃ receptor multimers were found to be abundantly expressed in brain, the major D₃ immunoreactivity expressed in stable D₃-expressing rat GH3 cells was found to be a monomer. However, multimeric D₃ receptor species with electrophoretic mobilities similar to those expressed in brain were also seen in D₃-expressing GH3 cells when a truncated D₃-like protein (named D(3nf))was co-expressed in these cells. Furthermore, results from immunoprecipitation experiments with D₃- and D(3nf)-specific antibodies show that the higher-order D₃ proteins extracted from brain and D₃/D(3nf) double transfectants also contain D(3nf)immunoreactivity, and immunocytochemical studies show that the expression of D₃ and D(3nf) immunoreactivities overlaps substantially in monkey and rat cortical neurons. Altogether, these data show oligomeric D₃ receptor protein expression in vivo and they suggest that at least some of these oligomers are heteroligomeric protein complexes containing D₃ and the truncated D(3nf) protein.