Gαq sensitizes TRPM8 to inhibition by PI(4,5)P2 depletion upon receptor activation

Activation of G-protein coupled receptors (GPCRs) was proposed to inhibit the cold and menthol sensitive Transient Receptor Potential Melastatin 8 (TRPM8) channels via direct binding of Gq to the channel. It is well documented that TRPM8 requires the plasma membrane phospholipid phosphatidylinositol 4,5-bisphosphate [PI(4,5)P2 or PIP2] for activity. It was claimed however that a decrease in cellular levels of this lipid does not contribute to channel inhibition upon receptor activation. Here we show that supplementing the whole cell patch pipette with PI(4,5)P2 reduced inhibition of TRPM8 by activation of Gq-coupled receptors in mouse dorsal root ganglion (DRG) neurons. Activation of the same receptors induced Phospholipase C (PLC) activation and decreased plasma membrane PI(4,5)P2 levels in these neurons. PI(4,5)P2 also reduced inhibition of TRPM8 by activation of heterologously expressed Gq-coupled muscarinic M1 receptors. Co-expression of a constitutively active Gq protein that does not couple to PLC inhibited TRPM8 activity, and in cells expressing this protein decreasing PI(4,5)P2 levels using a voltage sensitive 5-phosphatase induced a stronger inhibition of TRPM8 activity than in control cells. Our data indicate that PI(4,5)P2 depletion plays an important role in TRPM8 inhibition upon GPCR activation, and Gq inhibits the channel by reducing its apparent affinity for PI(4,5)P2 and thus sensitizes the channel to inhibition by decreasing PI(4,5)P2 levels.