Detection, Analysis, and Quantification of GPCR Homo- and Heteroreceptor Complexes in Specific Neuronal Cell Populations Using the In Situ Proximity Ligation Assay


Por: Borroto-Escuela D.O., Narvaez M., Valladolid-Acebes I., Shumilov K., Di Palma M., Wydra K., Schaefer T., Reyes-Resina I., Navarro G., Mudó G., Filip M., Sartini S., Friedland K., Schellekens H., Beggiato S., Ferraro L., Tanganelli S., Franco R., Belluardo N., Ambrogini P., Pérez de la Mora M., Fuxe K.

Publicada: 1 ene 2018
Resumen:
GPCR's receptosome operates via coordinated changes between the receptor expression, their modifications and interactions between each other. Perturbation in specific heteroreceptor complexes and/or their balance/equilibrium with other heteroreceptor complexes and corresponding homoreceptor complexes is considered to have a role in pathogenic mechanisms. Such mechanisms lead to mental and neurological diseases, including drug addiction, depression, Parkinson's disease, and schizophrenia. To understand the associations of GPCRs and to unravel the global picture of their receptor-receptor interactions in the brain, different experimental detection techniques for receptor-receptor interactions have been established (e.g., co-immunoprecipitation based approach). However, they have been criticized for not reflecting the cellular situation or the dynamic nature of receptor-receptor interactions. Therefore, the detection and visualization of GPCR homo-and heteroreceptor complexes in the brain remained largely unknown until recent years, when a well-characterized in situ proximity ligation assay (in situ PLA) was adapted to validate the receptor complexes in their native environment. The in situ PLA protocol presented here can be used to visualize GPCR receptor-receptor interactions in cells and tissues in a highly sensitive and specific manner. We have developed a combined method using immunohistochemistry and PLA, particularly aimed to monitor interactions between GPCRs in specific neuronal cell populations. This allows the analysis of homo-and heteroreceptor complexes at a cellular and subcellular level. The method has the advantage that it can be used in clinical specimens, providing localized, quantifiable homo-and heteroreceptor complexes detected in single cells. We compare the advantages and limitations of the methods, underlining recent progress and the growing importance of these techniques in basic research. We discuss also their potential as tools for drug development and diagnostics.

Filiaciones:
Borroto-Escuela D.O.:
 Department of Neuroscience, Karolinska Institutet, Stockholm, Sweden

 Observatorio Cubano de Neurociencias, Grupo Bohío-Estudio, Yaguajay, Cuba

 Department of Biomolecular Science, Section of Physiology, University of Urbino, Urbino, Italy

Narvaez M.:
 Facultad de Medicina, Instituto de Investigación Biomédica de Málaga, Universidad de Málaga, Málaga, Spain

Valladolid-Acebes I.:
 The Rolf Luft Research Center for Diabetes and Endocrinology, Karolinska Institutet, Karolinska University Hospital L1, Stockholm, Sweden

Shumilov K.:
 Department of Neuroscience, Karolinska Institutet, Stockholm, Sweden

Di Palma M.:
 Department of Biomolecular Science, Section of Physiology, University of Urbino, Urbino, Italy

Wydra K.:
 Laboratory of Drug Addiction Pharmacology, Institute of Pharmacology, Polish Academy of Sciences, Kraków, Poland

Schaefer T.:
 Department of Neuroscience, Karolinska Institutet, Stockholm, Sweden

 Molecular & Clinical Pharmacy, Friedrich-Alexander-Universität Erlangen, Erlangen, Germany

Reyes-Resina I.:
 CIBERNED, Centro de Investigación Biomédica en Red sobre Enfermedades Neurodegenerativas, Instituto de Salud Carlos III, Madrid, Spain

 Institute of Biomedicine of the University of Barcelona (IBUB), Barcelona, Spain

 Department of Biochemistry and Molecular Biomedicine, Faculty of Biology, University of Barcelona, Barcelona, Spain

Navarro G.:
 CIBERNED, Centro de Investigación Biomédica en Red sobre Enfermedades Neurodegenerativas, Instituto de Salud Carlos III, Madrid, Spain

 Institute of Biomedicine of the University of Barcelona (IBUB), Barcelona, Spain

 Department of Biochemistry and Physiology, Faculty of Pharmacy, University of Barcelona, Barcelona, Spain

Mudó G.:
 Department of Experimental Biomedicine and Clinical Neurosciences, University of Palermo, Palermo, Italy

Filip M.:
 Laboratory of Drug Addiction Pharmacology, Institute of Pharmacology, Polish Academy of Sciences, Kraków, Poland

Sartini S.:
 Department of Biomolecular Science, Section of Physiology, University of Urbino, Urbino, Italy

Friedland K.:
 Molecular & Clinical Pharmacy, Friedrich-Alexander-Universität Erlangen, Erlangen, Germany

Schellekens H.:
 APC Microbiome Institute, University College Cork, Cork, Ireland

 Department of Anatomy and Neuroscience, University College Cork, Cork, Ireland

Beggiato S.:
 Department of Medical Sciences, University of Ferrara, Ferrara, Italy

Ferraro L.:
 Department of Medical Sciences, University of Ferrara, Ferrara, Italy

Tanganelli S.:
 Department of Life Sciences and Biotechnology (SVEB), University of Ferrara, Ferrara, Italy

Franco R.:
 CIBERNED, Centro de Investigación Biomédica en Red sobre Enfermedades Neurodegenerativas, Instituto de Salud Carlos III, Madrid, Spain

 Institute of Biomedicine of the University of Barcelona (IBUB), Barcelona, Spain

 Department of Biochemistry and Molecular Biomedicine, Faculty of Biology, University of Barcelona, Barcelona, Spain

Belluardo N.:
 Department of Experimental Biomedicine and Clinical Neurosciences, University of Palermo, Palermo, Italy

Ambrogini P.:
 Department of Biomolecular Science, Section of Physiology, University of Urbino, Urbino, Italy

Pérez de la Mora M.:
 Instituto de Fisiología Celular, Universidad Nacional Autónoma de México, Mexico City, Mexico

Fuxe K.:
 Department of Neuroscience, Karolinska Institutet, Stockholm, Sweden
ISSN: 08932336
Editorial
Humana Press Inc., 999 RIVERVIEW DR, STE 208, TOTOWA, NJ 07512-1165 USA, Estados Unidos America
Tipo de documento: Article
Volumen: 140 Número:
Páginas: 299-315
WOS Id: 000456573300021