Abstract
Hetero-oligomers of G-protein-coupled receptors have become the subject of intense investigation, because their purported potential to manifest signaling and pharmacological properties that differ from the component receptors makes them highly attractive for the development of more selective pharmacological treatments. In particular, dopamine D1 and D2 receptors have been proposed to form hetero-oligomers that couple to Gαq proteins, and SKF83959 has been proposed to act as a biased agonist that selectively engages these receptor complexes to activate Gαq and thus phospholipase C. D1/D2 heteromers have been proposed as relevant to the pathophysiology and treatment of depression and schizophrenia. We used in vitro bioluminescence resonance energy transfer, ex vivo analyses of receptor localization and proximity in brain slices, and behavioral assays in mice to characterize signaling from these putative dimers/oligomers. We were unable to detect Gαq or Gα11 protein coupling to homomers or heteromers of D1 or D2 receptors using a variety of biosensors. SKF83959-induced locomotor and grooming behaviors were eliminated in D1 receptor knockout (KO) mice, verifying a key role for D1-like receptor activation. In contrast, SKF83959-induced motor responses were intact in D2 receptor and Gαq KO mice, as well as in knock-in mice expressing a mutant Ala286-CaMKIIα that cannot autophosphorylate to become active. Moreover, we found that, in the shell of the nucleus accumbens, even in neurons in which D1 and D2 receptor promoters are both active, the receptor proteins are segregated and do not form complexes. These data are not compatible with SKF83959 signaling through Gαq or through a D1/D2 heteromer and challenge the existence of such a signaling complex in the adult animals that we used for our studies.
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Acknowledgements
This work was supported by NIH grants RO1MH086629 (GDS), F31DA029499 (to ALF), TL1 RR024158-04 (to HY), K05DA022413 and R01MH54137 (to JAJ), RO1MH093672 (to CK), R01NS078291 (to RJC), a Research scientist award from the Research Foundation for Mental Hygiene (to PT) and the Lieber Center for Schizophrenia Research and Treatment. Behavioral work was performed at the Vanderbilt Mouse Neurobehavioral Core, which is supported in part by P30HD15052. We thank Dr Celine Gales (Institut National de la Santé et de la Recherche Médicale, Toulouse, France) and Dr Nevin Lambert (Georgia Health Sciences University, Augusta, Georgia) for kindly sharing Rluc and Venus fusion G protein constructs, Dr Stefan Offermanns (University of Heidelberg, Germany) for supplying the Gαq mutant line and Dr Nicole Calakos (Duke University) for the Drd1a-tdTomato reporter line. We also thank Matt Buendia, Heather Durai and Dr John Allison for excellent technical assistance.
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Frederick, A., Yano, H., Trifilieff, P. et al. Evidence against dopamine D1/D2 receptor heteromers. Mol Psychiatry 20, 1373–1385 (2015). https://doi.org/10.1038/mp.2014.166
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DOI: https://doi.org/10.1038/mp.2014.166
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