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Revealing Adenosine A2A-Dopamine D2 Receptor Heteromers in Parkinson's Disease Post-Mortem Brain through a New AlphaScreen-Based Assay

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Background: Several biophysical techniques have been successfully implemented to detect G protein-coupled receptors (GPCRs) heteromerization. Although these approaches have made it possible to ascertain the presence of GPCR heteromers in animal models of disease, no success has been accomplished in pathological human post-mortem brains. The AlphaScreen technology has been consistently used to quantify small analyte accumulation or depletion, bimolecular interactions, and post-translational modifications. The high signal-to-background, dynamic range and sensitivity exhibited by this technology support that it may be suitable to detect GPCR heteromers even under non-optimal conditions. Methods: Here, we describe the development of a new AlphaScreen assay to detect GPCR oligomers in human post-mortem brain. Results: Adenosine A2A-dopamine D2 receptor (A2AR/D2R) heteromer formation was monitored in caudate from healthy and Parkinson's disease (PD) subjects. The approach was first validated using striatal membranes from wild type and A2AR deficient mice. Secondly, we took advantage of the 6-hydroxydopamine hemiparkinsonian rat model to validate previous results. In addition, finally, A2AR/D2R heteromer formation was assessed in caudate membranes from human post-mortem brains. Importantly, our preliminary results revealed an increase in A2AR/D2R heteromer formation in PD brains. Conclusions: The new AlphaScreen assay allowed assessing GPCR heteromers in human post-mortem brains with high sensitivity.

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International Journal of Molecular Sciences, 2019, vol. 20, num. 14, p. 3600

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cc-by (c) Fernández Dueñas, Víctor et al., 2019

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Articles publicats en revistes (Patologia i Terapèutica Experimental)

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Revealing Adenosine A2A-Dopamine D2 Receptor Heteromers in Parkinson's Disease Post-Mortem Brain through a New AlphaScreen-Based Assay