Detection, analysis, and quantification of GPCR homo- and heteroreceptor complexes in specific neuronal cell populations using the in situ proximity ligation assay

Giuseppa Mudo', Natale Belluardo, Sarah Beggiato, Gemma Navarro, Dasiel O. Borroto-Escuela, Manuel Narvaez, Harriët Schellekens, Ismael Valladolid-Acebes, Malgorzata Filip, Kjell Fuxe, Michael Di Palma, Miguel Pérez De La Mora, Patrizia Ambrogini, Irene Reyes-Resina, Kristina Friedland, Kirill Shumilov, Thorsten Schaefer, Karolina Wydra, Stefano Sartini, Luca FerraroSergio Tanganelli, Rafael Franco

Risultato della ricerca: Chapter

3 Citazioni (Scopus)

Abstract

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.
Lingua originaleEnglish
Titolo della pubblicazione ospiteReceptor-Receptor Interactions in the Central Nervous System
Pagine299-315
Numero di pagine17
Stato di pubblicazionePublished - 2018

Serie di pubblicazioni

NomeNEUROMETHODS

All Science Journal Classification (ASJC) codes

  • Neuroscience(all)
  • Biochemistry, Genetics and Molecular Biology(all)
  • Pharmacology, Toxicology and Pharmaceutics(all)
  • Psychiatry and Mental health

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