Mammalian transient receptor potential ankyrin 1 (TRPA1) is a polymodal nociceptor that plays an important role in pain generation, but its role as a cold nociceptor is still controversial. Here, we propose that TRPA1 can sense noxious cold via transduction of reactive oxygen species (ROS) signalling. We show that inhibiting hydroxylation of a proline residue within the N-terminal ankyrin repeat of human TRPA1 by mutation or using a prolyl hydroxylase (PHD) inhibitor potentiates the cold sensitivity of TRPA1 in the presence of hydrogen peroxide. Inhibiting PHD in mice triggers mouse TRPA1 sensitization sufficiently to sense cold-evoked ROS, which causes cold hypersensitivity. Furthermore, this phenomenon underlies the acute cold hypersensitivity induced by the chemotherapeutic agent oxaliplatin or its metabolite oxalate. Thus, our findings provide evidence that blocking prolyl hydroxylation reveals TRPA1 sensitization to ROS, which enables TRPA1 to convert ROS signalling into cold sensitivity.

Recent evidence suggests that transient receptor potential melastatin 2 (TRPM2) expressed in immune cells plays an important role in immune and inflammatory responses. We recently reported that TRPM2 expressed in macrophages and spinal microglia contributes to the pathogenesis of inflammatory and neuropathic pain aggravating peripheral and central pronociceptive inflammatory responses in mice. To further elucidate the contribution of TRPM2 expressed by peripheral immune cells to neuropathic pain, we examined the development of peripheral nerve injury-induced neuropathic pain and the infiltration of immune cells (particularly macrophages) into the injured nerve and spinal cord by using bone marrow (BM) chimeric mice by crossing wildtype (WT) and TRPM2-knockout (TRPM2-KO) mice. Four types of BM chimeric mice were prepared, in which irradiated WT or TRPM2-KO recipient mice were transplanted with either WT-or TRPM2-KO donor mouse-derived green fluorescence protein-positive (GFP+) BM cells (TRPM2BM+/Rec+, TRPM2BM–/Rec+, TRPM2BM+/Rec–, and TRPM2BM–/Rec– mice). Mechanical allodynia induced by partial sciatic nerve ligation observed in TRPM2BM+/Rec+ mice was attenuated in TRPM2BM–/Rec+, TRPM2BM+/Rec–, and TRPM2BM–/Rec– mice. The numbers of GFP+ BM-derived cells and Iba1/GFP double-positive macrophages in the injured sciatic nerve did not differ among chimeric mice 14 days after the nerve injury. In the spinal cord, the number of GFP+ BM-derived cells, particularly GFP/Iba1 double-positive macrophages, was significantly decreased in the three TRPM2-KO chimeric mouse groups compared with TRPM2BM+/Rec+ mice. However, the numbers of GFP–/Iba1+ resident microglia did not differ among chimeric mice. These results suggest that TRPM2 plays an important role in the infiltration of peripheral immune cells, particularly macrophages, into the spinal cord, rather than the infiltration of peripheral immune cells into the injured nerves and activation of spinal-resident microglia. The spinal infiltration of macrophages mediated by TRPM2 may contribute to the pathogenesis of neuropathic pain.

Dysesthesia is an unpleasant abnormal sensation, which is often accompanied by peripheral neuropathy or vascular impairment. Here, we examined the roles of transient receptor potential ankyrin 1 (TRPA1) in dysesthesia-like behaviours elicited by transient hindlimb ischemia (15–60 min) by tightly compressing the hindlimb, and reperfusion by releasing the ligature. The paw-withdrawal responses to tactile stimulation were reduced during ischemia and lasted for a while after reperfusion. Hindlimb ischemia/reperfusion elicited spontaneous licking of the ischemic hindpaw that peaked within 10 min. The licking was inhibited by reactive oxygen species (ROS) scavengers, a TRPA1 antagonist, or TRPA1 deficiency, but not by TRPV1 deficiency. In human TRPA1-expressing cells as well as cultured mouse dorsal root ganglion neurons, the H2O2-evoked TRPA1 response was significantly increased by pretreatment with hypoxia (80 mmHg) for 30 min. This hypoxia-induced TRPA1 sensitisation to H2O2 was inhibited by overexpressing a catalytically-inactive mutant of prolyl hydroxylase (PHD) 2 or in a TRPA1 proline mutant resistant to PHDs. Consistent with these results, a PHD inhibitor increased H2O2-evoked nocifensive behaviours through TRPA1 activation. Our results suggest that transient hindlimb ischemia/reperfusion-evoked spontaneous licking, i.e. painful dysesthesia, is caused by ROS-evoked activation of TRPA1 sensitised by hypoxia through inhibiting PHD-mediated hydroxylation of a proline residue in TRPA1.

Some chemotherapeutic agents including platinum compounds, taxanes, vinca alkaloids, and bortezomib, cause peripheral neuropathy. However, unlike other chemotherapeutic agents, oxaliplatin, a platinum-based chemotherapeutic agent, induces a peculiar acute peripheral neuropathy, such as paresthesia and dysesthesia, which are often triggered or enhanced by exposure to cold, whereas cumulative and chronic neuropathy induced by oxaliplatin includes pain sensation (1). The acute peripheral neuropathy is induced in almost all patients during or within hours after the infusion. However, an effective pharmacological strategy for its management remains controversial (1, 2).Many studies in animal models focus on the oxaliplatininduced chronic and/or subacute painful peripheral neuro pathy that appear several days to several weeks after oxaliplatin administration (3 -5), while oxaliplatininduced acute peripheral neuropathy is poorly characterized. Recently, we have reported a mouse model of rapid-onset cold hypersensitivity induced by oxaliplatin (6). When mice are given a single administration of oxaliplatin, cold but not mechanical hypersensitivity is induced within 2 h after the administration, while it is not induced by other chemotherapeutic agents such as cisplatin and paclitaxel. These findings suggest that rapid-onset cold hypersensitivity is representative of the acute peripheral neuropathy characteristic to oxaliplatin in mice. In the present study, we assessed the effects of standard analgesics, such as non-steroidal anti-inflammatory agent (NSAID), opioid analgesics, tricyclic antidepressant, serotonin and noradrenaline reuptake inhibitor (SNRI), calcium channel a 2 -d ligand, local anesthetic, and calcium gluconate (1, 2), on the oxaliplatin-induced acute cold hypersensitivity in mice. 54 Shogoin-Kawahara-cho, Sakyo-ku, Kyoto 606-8507, Japan Received November 26, 2013; Accepted February 17, 2014 Abstract. Oxaliplatin, a platinum-based chemotherapeutic agent, causes an acute peripheral neuropathy triggered by cold in almost all patients during or within hours after its infusion. We recently reported that a single administration of oxaliplatin induced cold hypersensitivity 2 h after the administration in mice. In this study, we examined whether standard analgesics relieve the oxaliplatin-induced acute cold hypersensitivity. Gabapentin, tramadol, mexiletine, and calcium gluconate significantly inhibited and morphine and milnacipran decreased the acute cold hypersensitivity, while diclofenac and amitriptyline had no effects. These results suggest that gabapentin, tramadol, mexiletine, and calcium gluconate are effective against oxaliplatin-induced acute peripheral neuropathy. Pharmacological Characterization of Standard Analgesics on OxaliplatinInduced Acute Cold Hypersensitivity in Mice