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

Abstract

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.

Inflammatory and neuropathic cold allodynia are selectively mediated by the neurotrophic factor receptor GFRα3

“…WT mice show robust cold allodynia 2 d after unilateral injections of the inflammatory agent complete Freund's adjuvant (CFA) (Fig. 1A) (P < 0.01, pre-vs. post-CFA or ipsilateral vs. contralateral), which we and others have previously reported (22)(23)(24). In contrast, Gfrα3 −/− mice show no differences in their hind paw lift latencies between the ipsilateral (inflamed) and the contralateral (control) sides, and there were no differences in their sensitivity compared with the basal, preinflamed state (Fig.…”

“…1B) (P < 0.01, preinjury vs. 7 d postinjury; P < 0.001, ipsilateral vs. contralateral), but cold sensitivity was remarkably unchanged in Gfrα3 −/− mice (ipsilateral vs. contralateral; preinjury vs. 7 d postinjury; P > 0.05). Lastly, one of the major side effects of platin-based chemotherapeutics is cold pain (26), a phenotype that can be modeled in mice given a single systemic injection of oxaliplatin (22,23). As with the previous pain models, the cold allodynia observed in WT mice (P < 0.001, basal vs. 7 d postinjection) was completely absent in mice null for GFRα3 ( Fig.…”

“…Artemin-and NGF-induced sensitization of cold responses is TRPM8-dependent, and multiple studies have shown that TRPM8 plays a role in the development of cold allodynia (8,(22)(23)(24)57). However, inflammatory cold allodynia is also diminished by both an TRPA1 antagonist and reduced TRPA1 transcript expression, and cold hypersensitivity can be induced by TRPA1 agonism in WT but not Trpa1 −/− mice (58, 59).…”

“…Next, to test the role of GFRα3 in pathological cold pain, we examined adult WT and Gfrα3 −/− littermates in classical models of inflammation, nerve injury, and chemotherapeutic-induced neuropathic pain (22,23). WT mice show robust cold allodynia 2 d after unilateral injections of the inflammatory agent complete Freund's adjuvant (CFA) (Fig.…”

Tryptamine-Based Derivatives as Transient Receptor Potential Melastatin Type 8 (TRPM8) Channel Modulators

“…2 Notably, both TRPM8 agonists and antagonists exert analgesic effects: in particular, while TRPM8 agonists produce profound analgesia 2 at very low concentrations, even greater effects have been reported for TRPM8 antagonists. 3 This analgesia modulation represents a novel approach in a largely unmet therapeutic need. 4 Moreover, many studies have recently revealed novel potential pharmacological implications for TRPM8 modulators.…”