Activity of the Neuronal Cold Sensor TRPM8 Is Regulated by Phospholipase C via the Phospholipid Phosphoinositol 4,5-Bisphosphate

Abstract

Cold temperatures robustly activate a small cohort of somatosensory nerves, yet during a prolonged cold stimulus their activity will decrease, or adapt, over time. This process allows for the discrimination of subtle changes in temperature. At the molecular level, cold is detected by transient receptor potential melastatin 8 (TRPM8), a nonselective cation channel expressed on a subset of peripheral afferent fibers. We and others have reported that TRPM8 channels also adapt in a calcium-dependent manner when activated by the cooling compound menthol. Additionally, TRPM8 activity is sensitive to the phospholipid phosphoinositol 4,5-bisphosphate (PIP 2 ), a substrate for the enzyme phospholipase C (PLC). These results suggest an adaptation model whereby TRPM8-mediated Ca 2؉ influx activates PLC, thereby decreasing PIP 2 levels and resulting in reduced TRPM8 activity. Here we tested this model using pharmacological activation of PLC and by manipulating PIP 2 levels independent of both PLC and Ca 2؉ . PLC activation leads to adaptation-like reductions in cold-or menthol-evoked TRPM8 currents in both heterologous and native cells. Moreover, PLCindependent reductions in PIP 2 had a similar effect on cold-and menthol-evoked currents. Mechanistically, either form of adaptation does not alter temperature sensitivity of TRPM8 but does lead to a change in channel gating. Our results show that adaptation is a shift in voltage dependence toward more positive potentials, reversing the trend toward negative potentials caused by agonist. These data suggest that PLC activity not only mediates adaptation to thermal stimuli, but likely underlies a more general mechanism that establishes the temperature sensitivity of somatosensory neurons.The detection of temperature is a fundamental task of the nervous system. Temperature-sensing sensory afferent neurons reside in either the trigeminal (TG) 2 or dorsal root (DRG) sensory ganglia and project peripherally, terminating as free nerve endings that innervate areas of the head or trunk, respectively (1, 2). Subpopulations of these afferents respond to distinct sub-modalities of thermal stimuli, including noxious heat, innocuous cooling and warmth, and painfully cold temperatures. Each carries thermal information to the dorsal horn of the spinal cord, synapsing with neurons that project centrally (1, 3). The discovery of thermosensitive ion channels of the transient receptor potential (TRP) family demonstrated an underlying molecular mechanism for temperature detection (4). Cold temperature sensation is largely mediated by TRPM8, a nonselective cation channel expressed on a small subset of neurons (5, 6). TRPM8 is activated by cooling compounds, such as menthol, as well as cold temperatures below ϳ28°C, in vitro (7,8). Recent reports on the behavioral phenotype of TRPM8-null mice suggest that this lone channel is required for the majority of cold sensing in vivo (5, 9 -11). These and other data strongly implicate TRPM8 in not only the detection of both innocuous cool and some aspects of ...

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“…Notable examples are the mammalian TRPM8 and TRPV1 channels, activated by cold/menthol (Dhaka et al, 2007) and heat/vanilloid (Caterina et al, 1997) compounds, respectively, both involved in nociception. Interestingly, both of these channels are regulated by Ca 2+ -dependent and -independent pathways and cyclic nucleotides (Bhave et al, 2002;Daniels et al, 2008;De Petrocellis et al, 2007;Vanden Abeele et al, 2006). Other members of the TRP channel family, PKD1L3 and PKD2L1, have been recently implicated in the detection of sour compounds in mammals, while GPCRs are responsible for the detection of umami, sweet and bitter Huang et al, 2006;Ishimaru et al, 2006).…”

Complex Regulation of the TRPM8 Cold Receptor Channel

“…4, panel 5). In the event of M3 receptor-mediated TRPM8 inhibition, the involvement of the Gq/PLC pathway would be most anticipated, as the substrate of PLC activity, PIP 2 , is a well known co-factor in TRPM8 activation (15,16), whose depletion during PLC activity causes I TRPM8 suppression (35). However, our experiments with PLC inhibitor U73122 did not support the notion on Gq/PLC involvement, as this compound failed to impair in any essential way the inhibitory effects of Oxo-M on TRPM8 in HEK TRPM8/M3 cells.…”