Co-immunoprecipitation of Membrane-Bound Receptors from Subsynaptic Compartments
The allosteric modulation in G protein-coupled heteroreceptors have opened a new vision of the physiology and pathophysiology of the central nervous system. Understanding the role of the receptor-receptor interactions in the synaptic compartments is essential to elucidate the function of these macromolecular complexes in the brain intercellular communication that occurs within wiring and volume transmission. Here, we describe a procedure to identify the receptor-receptor interactions in different enriched subsynaptic fractions. First, we present a protocol to obtain synaptosomes by means of a discontinuous sucrose gradient. The method continues with a subcellular fractionation to separate the pre-synaptic, the post-synaptic, and the extra-synaptic fractions. Finally, we describe the Co-immunoprecipitation as an accepted technique to identify the receptor-receptor interaction from isolated and validated subsynaptic compartments.
Key wordsG protein-coupled receptors Heteroreceptors Receptor-receptor interaction Synaptosome Sucrose gradient Subcellular fractionation Synaptic compartments Co-immunoprecipitation
This work was supported by Prometeo Program of the Higher Education, Science, Technology and Innovation Secretary and the Technical University of Ambato (grants 1243-CU-P-2014 to WR-F, 0912-CU-P-2016 and 0932-CU-P-2016 to WR-F and YP-B, and 0906-CU-P-2016 to MG-B).
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