How cold is it? TRPM8 and TRPA1 in the molecular logic of cold sensation

Recognition of temperature is a critical element of sensory perception and allows us to evaluate both our external and internal environments. In vertebrates, the somatosensory system can discriminate discrete changes in ambient temperature, which activate nerve endings of primary afferent fibers. These thermosensitive nerves can be further segregated into those that detect either innocuous or noxious (painful) temperatures; the latter neurons being nociceptors. We now know that thermosensitive afferents express ion channels of the transient receptor potential (TRP) family that respond at distinct temperature thresholds, thus establishing the molecular basis for thermosensation. Much is known of those channels mediating the perception of noxious heat; however, those proposed to be involved in cool to noxious cold sensation, TRPM8 and TRPA1, have only recently been described. The former channel is a receptor for menthol, and links the sensations provided by this and other cooling compounds to temperature perception. While TRPM8 almost certainly performs a critical role in cold signaling, its part in nociception is still at issue. The latter channel, TRPA1, is activated by the pungent ingredients in mustard and cinnamon, but has also been postulated to mediate our perception of noxious cold temperatures. However, a number of conflicting reports have suggested that the role of this channel in cold sensation needs to be confirmed. Thus, the molecular logic for the perception of cold-evoked pain remains enigmatic. This review is intended to summarize our current understanding of these cold thermoreceptors, as well as address the current controversy regarding TRPA1 and cold signaling

Pharmacological Blockade of TRPM8 Ion Channels Alters Cold and Cold Pain Responses in Mice

TRPM8 (Transient Receptor Potential Melastatin-8) is a cold- and menthol-gated ion channel necessary for the detection of cold temperatures in the mammalian peripheral nervous system. Functioning TRPM8 channels are required for behavioral responses to innocuous cool, noxious cold, injury-evoked cold hypersensitivity, cooling-mediated analgesia, and thermoregulation. Because of these various roles, the ability to pharmacologically manipulate TRPM8 function to alter the excitability of cold-sensing neurons may have broad impact clinically. Here we examined a novel compound, PBMC (1-phenylethyl-4-(benzyloxy)-3-methoxybenzyl(2-aminoethyl)carbamate) which robustly and selectively inhibited TRPM8 channels in vitro with sub-nanomolar affinity, as determined by calcium microfluorimetry and electrophysiology. The actions of PBMC were selective for TRPM8, with no functional effects observed for the sensory ion channels TRPV1 and TRPA1. PBMC altered TRPM8 gating by shifting the voltage-dependence of menthol-evoked currents towards positive membrane potentials. When administered systemically to mice, PBMC treatment produced a dose-dependent hypothermia in wildtype animals while TRPM8-knockout mice remained unaffected. This hypothermic response was reduced at lower doses, whereas responses to evaporative cooling were still significantly attenuated. Lastly, systemic PBMC also diminished cold hypersensitivity in inflammatory and nerve-injury pain models, but was ineffective against oxaliplatin-induced neuropathic cold hypersensitivity, despite our findings that TRPM8 is required for the cold-related symptoms of this pathology. Thus PBMC is an attractive compound that serves as a template for the formulation of highly specific and potent TRPM8 antagonists that will have utility both in vitro and in vivo

Evolution of Thermal Response Properties in a Cold-Activated TRP Channel

Animals sense changes in ambient temperature irrespective of whether core body temperature is internally maintained (homeotherms) or subject to environmental variation (poikilotherms). Here we show that a cold-sensitive ion channel, TRPM8, displays dramatically different thermal activation ranges in frogs versus mammals or birds, consistent with variations in these species' cutaneous and core body temperatures. Thus, somatosensory receptors are not static through evolution, but show functional diversity reflecting the characteristics of an organism's ecological niche

Near-Membrane Dynamics and Capture of TRPM8 Channels within Transient Confinement Domains

The cold and menthol receptor, TRPM8, is a non-selective cation channel expressed in a subset of peripheral neurons that is responsible for neuronal detection of environmental cold stimuli. It was previously shown that members of the transient receptor potential (TRP) family of ion channels are translocated toward the plasma membrane (PM) in response to agonist stimulation. Because the spatial and temporal dynamics of cold receptor cell-surface residence may determine neuronal activity, we hypothesized that the movement of TRPM8 to and from the PM might be a regulated process. Single particle tracking (SPT) is a useful tool for probing the organization and dynamics of protein constituents in the plasma membrane.We used SPT to study the receptor dynamics and describe membrane/near-membrane behavior of particles containing TRPM8-EGFP in transfected HEK-293T and F-11 cells. Cells were imaged using total internal reflection fluorescence (TIRF) microscopy and the 2D and 3D trajectories of TRPM8 molecules were calculated by analyzing mean-square particle displacement against time. Four characteristic types of motion were observed: stationary mode, simple Brownian diffusion, directed motion, and confined diffusion. In the absence of cold or menthol to activate the channel, most TRPM8 particles move in network covering the PM, periodically lingering for 2–8 s in confined microdomains of about 800 nm radius. Removing cholesterol with methyl-beta-cyclodextrin (MβCD) stabilizes TRPM8 motion in the PM and is correlated with larger TRPM8 current amplitude that results from an increase in the number of available channels without a change in open probability.These results reveal a novel mechanism for regulating TRPM8 channel activity, and suggest that PM dynamics may play an important role in controlling electrical activity in cold-sensitive neurons

Citral Sensing by TRANSient Receptor Potential Channels in Dorsal Root Ganglion Neurons

Transient receptor potential (TRP) ion channels mediate key aspects of taste, smell, pain, temperature sensation, and pheromone detection. To deepen our understanding of TRP channel physiology, we require more diverse pharmacological tools. Citral, a bioactive component of lemongrass, is commonly used as a taste enhancer, as an odorant in perfumes, and as an insect repellent. Here we report that citral activates TRP channels found in sensory neurons (TRPV1 and TRPV3, TRPM8, and TRPA1), and produces long-lasting inhibition of TRPV1–3 and TRPM8, while transiently blocking TRPV4 and TRPA1. Sustained citral inhibition is independent of internal calcium concentration, but is state-dependent, developing only after TRP channel opening. Citral's actions as a partial agonist are not due to cysteine modification of the channels nor are they a consequence of citral's stereoisoforms. The isolated aldehyde and alcohol cis and trans enantiomers (neral, nerol, geranial, and geraniol) each reproduce citral's actions. In juvenile rat dorsal root ganglion neurons, prolonged citral inhibition of native TRPV1 channels enabled the separation of TRPV2 and TRPV3 currents. We find that TRPV2 and TRPV3 channels are present in a high proportion of these neurons (94% respond to 2-aminoethyldiphenyl borate), consistent with our immunolabeling experiments and previous in situ hybridization studies. The TRPV1 activation requires residues in transmembrane segments two through four of the voltage-sensor domain, a region previously implicated in capsaicin activation of TRPV1 and analogous menthol activation of TRPM8. Citral's broad spectrum and prolonged sensory inhibition may prove more useful than capsaicin for allodynia, itch, or other types of pain involving superficial sensory nerves and skin

Transient receptor potential canonical 4 and 5 proteins as targets in cancer therapeutics

Novel approaches towards cancer therapy are urgently needed. One approach might be to target ion channels mediating Ca²+ entry because of the critical roles played by Ca²+ in many cell types, including cancer cells. There are several types of these ion channels, but here we address those formed by assembly of transient receptor potential canonical (TRPC) proteins, particularly those which involve two closely related members of the family: TRPC4 and TRPC5. We focus on these proteins because recent studies point to roles in important aspects of cancer: drug resistance, transmission of drug resistance through extracellular vesicles, tumour vascularisation, and evoked cancer cell death by the TRPC4/5 channel activator (−)-englerin A. We conclude that further research is both justified and necessary before these proteins can be considered as strong targets for anti-cancer cell drug discovery programmes. It is nevertheless already apparent that inhibitors of the channels would be unlikely to cause significant adverse effects, but, rather, have other effects which may be beneficial in the context of cancer and chemotherapy, potentially including suppression of innate fear, visceral pain and pathological cardiac remodelling