Ablation of TrpV1 neurons reveals their selective role in thermal pain sensation

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Abstract

Here we make use of neural ablation to investigate the properties of the TrpV1-expressing neurons in the trigeminal and dorsal root ganglia of mice. Resiniferotoxin (RTX), a potent TrpV1 agonist, administered either by direct injection in the ganglion or intrathecally killed approximately 70% of TrpV1 cells and resulted in modest thermal analgesia. Interestingly, after carageenan injection in the hind paw, the analgesic effects of RTX were dramatically increased with mice now paradoxically showing far less response to heat applied at sites of inflammation. This additional carageenan and RTX-induced analgesia was transient, lasting less than 2 days, and likely resulted from deafferentation of remaining TrpV1 neurons. Remarkably, although RTX affected sensitivity to heat, mechanical sensitivity (both of normal and inflamed tissue) was completely unaltered by toxin-mediated silencing of the TrpV1 sensory input. Thus, our data demonstrate that TrpV1 neurons are selectively tuned nociceptors that mediate responses to thermal but not mechanical pain and insinuate a labeled line model for somatosensory coding.

Introduction

In mammals, a family of Trp ion channels has been implicated in mediating thermosensation (Jordt et al., 2003). TrpV1, the capsaicin receptor, plays an important role in sensation of noxious heat (Caterina et al., 1997). TrpV1−/− mice have reduced thermal sensitivity but importantly loss of TrpV1 does not eliminate responses to noxious heat (Woodbury et al., 2004). TrpV1 is also believed to play a role in increased pain sensitivity following inflammation at least in part because inflammation induced phosphorylation of TrpV1 results in increased sensitivity of this ion channel to temperature (Bhave et al., 2002, Cesare et al., 1999, Nilius et al., 2005, Numazaki et al., 2002). Indeed, carageenan-induced inflammation strongly enhances the response of normal mice to noxious heat but does not sensitize TrpV1−/− mutants (Caterina et al., 2000). Thus, attention has focused on TrpV1 and TrpV1 cells for their role in pain sensation and as targets for pain therapies. Notably, TrpV1 antagonists have been shown to be efficacious in animal models of chronic pain (Szallasi et al., 2007, Wong and Gavva, 2009) but, as yet, have not been exploited in human pain management.

An alternate strategy for pain management involves ablation of TrpV1-expressing cells or TrpV1 fibers. This approach has a long history: more than 25 years ago, capsaicin injection was reported to dramatically decrease the level of inflammatory mediators (e.g., substance P) and pain sensation for the life of animals (Jancso et al., 1977, Nagy et al., 1981, Yaksh et al., 1979). More recently, the potent TrpV1 agonist, resiniferotoxin (RTX) has been demonstrated to kill TrpV1-expressing neurons by affecting long-term Ca2+ entry (Olah et al., 2001). RTX has also been used therapeutically in animals and has been reported to alleviate chronic pain with no effects on discriminative touch or proprioception (Brown et al., 2005, Karai et al., 2004, Neubert et al., 2003). In a rat model, RTX was shown to eliminate many TrpV1 neurons, dramatically decrease responses to capsaicin and thermal pain (Karai et al., 2004), and affect mechanosensation (Tender et al., 2008).

Here, we set out to determine the function of TrpV1 cells and explored the use of RTX as a means to selectively eliminate these neurons in mice. We carried out a detailed investigation into the efficacy of RTX in ablating sensory neurons and demonstrate that although a large subset of TrpV1 neurons are killed by a single injection, other TrpV1 cells are not affected even by repeated RTX treatments. RTX reduces but does not eliminate capsaicin responses and thermal sensation of normal mice. In contrast, after inflammation, RTX was far more effective and resulted in a profound loss of thermal sensation without any corresponding effect on mechanosensation (including to painful stimuli). Thus, our data strongly support a model where TrpV1 neurons selectively mediate thermal nociception and a distinct labeled line transmits mechanical pain (see also Cavanaugh et al., 2009).

Section snippets

Results

RTX is a potent TrpV1 agonist that has been reported to effectively kill TrpV1-expressing cells in vitro and selectively ablate capsaicin-sensitive sensory neurons in animal models (Karai et al., 2004). Thus, RTX is an attractive agent to help reveal the role of TrpV1 neurons in vivo and to generate ganglia depleted in this subset of neurons for transcriptome analysis. In initial experiments, we administered large amounts of RTX to mice (10-fold higher doses than previously reported effective

Discussion

Recently, considerable attention has focused on RTX as a novel therapeutic agent for treating chronic pain by ablation of TrpV1-expressing cells in select sensory ganglia. Here, we demonstrate that although RTX treatment is effective at killing many TrpV1-expressing sensory neurons, a population of large-diameter TrpA1/TrpV1-expressing cells (approximately 30% of the total TrpV1 cells in the trigeminal ganglion) is resistant to the toxin (Fig. 1, Fig. 2). We have shown that TrpA1 is simply a

Animal models

Mice were 25–30 g (2–4 months old) male C57/BL6, TrpV1−/− (Caterina et al., 2000), or TrpA1−/− (Kwan et al., 2006). Procedures followed the NIH guidelines for the care and use of laboratory animals and were approved by the National Institute of Dental and Craniofacial Research Animal Care and Use Committee.

Acknowledgments

We thank N. Ryba, T. Usdin, and S. Gutkind for helpful discussions and critical review of the manuscript. This work was supported by the intramural research program of the NIH, NIDCR.

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