The thienopyridine A-769662 and benzimidazole 991 inhibit human TASK-3 potassium channels in an AMPK-independent manner

Esraa A. Said, Ryan W. Lewis, Mark L. Dallas, Chris Peers, Fiona A. Ross, Asier Unciti-Broceta, D. Grahame Hardie, A. Mark Evans (Lead / Corresponding author)

    Research output: Contribution to journalArticlepeer-review

    Abstract

    Heteromeric Tandem pore domain Acid Sensitive (TASK)-1/3 channels are critical to oxygen-sensing by carotid body type 1 cells, where hypoxia-induced inhibition of TASK-3 and/or TASK-1/3 potassium currents leads to voltage-gated calcium entry, exocytotic transmitter release and increases in carotid body afferent input responses that initiate corrective changes in breathing patterns. It was proposed that, in response to hypoxia, the AMP–activated protein kinase (AMPK) might directly phosphorylate and inhibit TASK channels, in particular TASK–3, but studies on rat type I cells questioned this view. However, sequence alignment identified a putative AMPK recognition motif in human (h) TASK-3, but not hTASK–1, with Ser55 representing a potential phosphorylation site. We therefore studied the effects of five different AMPK activators on recombinant hTASK–3 potassium channels expressed in human embryonic kidney (HEK)–293 cells. Two structurally unrelated AMPK activators, the thienopyridine A–769662 (100–500 µM) and the benzimidazole 991 (3–30 µM) inhibited hTASK–3 currents in a concentration–dependent manner, while the 4-azabenzimidazole MK–8722 (3–30 µM) partially inhibited hTASK–3 at concentrations above those required for maximal AMPK activation. By contrast, the 4-azabenzimidazole, BI-9774 (10–100 µM; a closely related analogue of MK8722) and the pro-drug AICA-riboside (1 mM; metabolised to ZMP, an AMP-mimetic) had no significant effect on hTASK–3 currents at concentrations sufficient to maximally activate AMPK. Importantly, A–769662 (300 µM) also inhibited hTASK–3 channel currents in HEK–293 cells that stably over-expressed an AMPK–β1 subunit mutant (S108A) that renders AMPK insensitive to activators that bind to the Allosteric Drug and Metabolite site, such as A–769662. We therefore identify A–769662 and 991 as novel hTASK–3 channel inhibitors and provide conclusive evidence that AMPK does not regulate hTASK–3 channel currents.

    Original languageEnglish
    Article number116562
    Number of pages9
    JournalBiochemical Pharmacology
    Volume230
    Issue number1
    Early online date30 Oct 2024
    DOIs
    Publication statusPublished - Dec 2024

    Keywords

    • 4-azabenzimidazole
    • 991
    • A-769662
    • AICA-riboside
    • AMPK
    • Benzimidazole
    • BI-9774
    • human TASK-3
    • MK-8722
    • Thienopyridine

    ASJC Scopus subject areas

    • Biochemistry
    • Pharmacology

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