Triphenylphosphine Oxide Is a Potent and Selective Inhibitor of the Transient Receptor Potential Melastatin-5 Ion Channel

Abstract

Transient receptor potential melastatin-5 (TRPM5) is a calcium-gated monovalent cation channel expressed in highly specialized cells of the taste bud and gastrointestinal tract, as well as in pancreatic β-cells. Well established as a critical signaling protein for G protein-coupled receptor-mediated taste pathways, TRPM5 also has recently been implicated as a regulator of incretin and insulin secretion. To date, no inhibitors of practical use have been described that could facilitate investigation of TRPM5 functions in taste or secretion of metabolic hormones. Using recombinant TRPM5-expressing cells in a fluorescence imaging plate reader-based membrane potential assay, we identified triphenylphosphine oxide (TPPO) as a selective and potent inhibitor of TRPM5. TPPO inhibited both human (IC₅₀ = 12 μM) and murine TRPM5 (IC₅₀ = 30 μM) heterologously expressed in HEK293 cells, but had no effect (up to 100 μM) on the membrane potential responses of TRPA1, TRPV1, or TRPM4b. TPPO also inhibited a calcium-gated TRPM5-dependent conductance in taste cells isolated from the tongues of transgenic TRPM5(+/)⁻ mice. In contrast, TPP had no effect on TRPM5 responses, indicating a strict requirement of the oxygen atom for activity. Sixteen additional TPPO derivatives also inhibited TRPM5 but none more potently than TPPO. Structure-activity relationship of tested compounds was used for molecular modeling-based analysis to clarify the positive and negative structural contributions to the potency of TPPO and its derivatives. TPPO is the most potent TRPM5 inhibitor described to date and is the first demonstrated to exhibit selectivity over other channels.

Bitter triggers acetylcholine release from polymodal urethral chemosensory cells and bladder reflexes

“…4 B and C). Although TPRM5 itself is a monovalent-specific cation channel activated by Ca 2+ rather than a Ca 2+ channel itself (18), its inhibition by triphenylphosphine oxide (TPPO) is known to significantly reduce the tastant-induced rise in [Ca 2+ ] i in mouse taste bud cells (19); we found that this action applies to denatonium responses in urethral cells as well (Fig. 4D).…”

TRPM5 is critical for linoleic acid-induced CCK secretion from the enteroendocrine cell line, STC-1

“…Because the LA-induced TRP-like inward currents were selective for monovalent cations, we investigated the possibility of TRPM5 involvement in LA-induced signaling in STC-1 cells. TPPO, a specific antagonist for TRPM5 (19), significantly reduced LA-induced and, importantly, denatonium benzoate-induced inward currents. We saw very similar levels of inhibition of LA-induced and denatonium benzoate-induced inward currents upon application of TPPO.…”

“…TRPM5, as well as GPCR taste receptors and other G protein signaling elements, is expressed in STC-1 cells (4, 16, 21). On the basis of our results that LA-activated currents in STC-1 cells are primarily monovalent cation selective, we investigated specific involvement of TRPM5 channels in LA-activated inward current by using the TRPM5 channel blocker triphenylphosphine oxide (TPPO) (17,19). In whole cell voltage-clamp experiments, we compared LA-induced inward currents in control STC-1 cells and in TPPO-treated (100 M, 2-min pretreatment) STC-1 cells.…”

Bitter and sweet taste receptors regulate human upper respiratory innate immunity

“…Furthermore, denatonium-induced calcium responses were blocked by inhibition of PLCβ2 (Supplemental Figure 3A), an important downstream component of taste signal transduction (18). The duration of denatonium-induced calcium responses was reduced by either removal of extracellular calcium or application of triphenylphosphine oxide (Supplemental Figure 3B), an inhibitor of the TRPM5 component of taste signaling (19). Additionally, the inositol triphosphate receptor (IP 3 R) inhibitor xestospongin B significantly inhibited the denatonium-induced calcium response (Supplemental Figure 3C).…”