Fluorescence-based, high-throughput assays for μ-opioid receptor activation using a membrane potential-sensitive dye

Alisa Knapman, Mark Connor

    Research output: Chapter in Book/Report/Conference proceedingOther chapter contribution

    7 Citations (Scopus)

    Abstract

    The development of new and improved opioid analgesics requires high-throughput screening (HTS) methods to identify potential therapeutics from large libraries of lead compounds. Here we describe two simple, real-time fluorescence-based assays of μ-opioid receptor activation that may be scaled up for HTS. In AtT-20 cells expressing the μ-opioid receptor (MOPr), opioids activate endogenous G protein gated inwardly rectifying K channels (GIRK channels), leading to membrane hyperpolarization. In Chinese hamster ovary cells expressing MOPr, adenylyl cyclase activation via forskolin results in membrane hyperpolarization, which is inhibited by opioids. Changes in membrane potential can be measured using a proprietary membrane potential-sensitive dye. In contrast to many HTS methods currently available, these assays reflect naturalistic coupling of the receptor to effector molecules.

    Original languageEnglish
    Title of host publicationOpioid Receptors
    Subtitle of host publicationMethods and Protocols
    EditorsSanti M. Spampinato
    Place of PublicationTotowa, NJ
    PublisherSpringer, Springer Nature
    Pages177-185
    Number of pages9
    Edition1
    ISBN (Electronic)9781493917082
    ISBN (Print)9781493917075
    DOIs
    Publication statusPublished - 2015

    Publication series

    NameMethods in Molecular Biology: Methods and Protocols
    PublisherSpringer
    Volume1230
    ISSN (Print)1064-3745

    Keywords

    • adenylyl cyclase
    • AtT-20
    • CHO
    • fluorescent assay
    • GIRK
    • high-throughput screening
    • membrane potential
    • μ-opioid receptor

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