Nav1.7, a peripheral neuron voltage-gated sodium channel, is essential for pain and olfaction in mice and humans. We examined the role of Nav1.7 as well as Nav1.3, Nav1.8, and Nav1.9 in different mouse models of chronic pain. Constriction-injury-dependent neuropathic pain is abolished when Nav1.7 is deleted in sensory neurons, unlike nerve-transection-related pain, which requires the deletion of Nav1.7 in sensory and sympathetic neurons for pain relief. Sympathetic sprouting that develops in parallel with nerve-transection pain depends on the presence of Nav1.7 in sympathetic neurons. Mechanical and cold allodynia required distinct sets of neurons and different repertoires of sodium channels depending on the nerve injury model. Surprisingly, pain induced by the chemotherapeutic agent oxaliplatin and cancer-induced bone pain do not require the presence of Nav1.7 sodium channels or Nav1.8-positive nociceptors. Thus, similar pain phenotypes arise through distinct cellular and molecular mechanisms. Therefore, rational analgesic drug therapy requires patient stratification in terms of mechanisms and not just phenotype.

A vast diversity of salient cues is sensed by numerous classes of primary sensory neurons, defined by specific neuropeptides, ion channels, or cytoskeletal proteins. Recent evidence has demonstrated a correlation between the expression of some of these molecular markers and transmission of signals related to distinct sensory modalities (eg, heat, cold, pressure). Voltage-gated sodium channel Na(v)1.8 has been reported to be preferentially expressed in small-diameter unmyelinated sensory afferents specialized for the detection of noxious stimuli (nociceptors), and Na(v)1.8-Cre mice have been widely used to investigate gene function in nociceptors. However, the identity of neurons in which Cre-mediated recombination occurs in these animals has not been resolved, and whether expression of Na(v)1.8 in these neurons is dynamic during development is not known, rendering interpretation of conditional knockout mouse phenotypes problematic. Here, we used genetics, immunohistochemistry, electrophysiology, and calcium imaging to precisely characterize the expression of Na(v)1.8 in the peripheral nervous system. We demonstrate that 75% of dorsal root ganglion (DRG) neurons express Na(v)1.8-Cre, including >90% of neurons expressing markers of nociceptors and, unexpectedly, a large population (∼40%) of neurons with myelinated A fibers. Furthermore, analysis of DRG neurons' central and peripheral projections revealed that Na(v)1.8-Cre is not restricted to nociceptors but is also expressed by at least 2 types of low-threshold mechanoreceptors essential for touch sensation, including those with C and Aβ fibers. Our results indicate that Na(v)1.8 underlies electrical activity of sensory neurons subserving multiple functional modalities, and call for cautious interpretation of the phenotypes of Na(v)1.8-Cre-driven conditional knockout mice.

Tissue acidosis is effective in causing chronic muscle pain. However, how muscle nociceptors contribute to the transition from acute to chronic pain is largely unknown.

Mechanistic insights into pain pathways are essential for a rational approach to treating this vast and increasing clinical problem. Sensory neurons that respond to tissue damage (nociceptors) may evoke pain sensations and are typically classified on the basis of action potential velocity. Electrophysiological studies have suggested that most of the C-fiber nociceptors are polymodal, responding to a variety of insults. In contrast, gene deletion studies in the sensory neurons of transgenic mice have frequently resulted in modality-specific deficits. We have used an in vivo imaging approach using the genetically encoded fluorescent calcium indicator GCaMP to study the activity of dorsal root ganglion sensory neurons in live animals challenged with painful stimuli. Using this approach, we can visualize spatially distinct neuronal responses and find that >85% of responsive dorsal root ganglion neurons are modality-specific, responding to either noxious mechanical, cold, or heat stimuli. These observations are mirrored in behavioral studies of transgenic mice. For example, deleting sodium channel Nav1.8 silences mechanical- but not heat-sensing sensory neurons, consistent with behavioral deficits. In contrast, primary cultures of axotomized sensory neurons show high levels of polymodality. After intraplantar treatment with prostaglandin E2, neurons in vivo respond more intensely to noxious thermal and mechanical stimuli, and additional neurons (silent nociceptors) are unmasked. Together, these studies define polymodality as an infrequent feature of nociceptive neurons in normal animals.

The skin has a dual function as a barrier and a sensory interface between the body and the environment. To protect against invading pathogens, the skin harbours specialized immune cells, including dermal dendritic cells (DDCs) and interleukin (IL)-17-producing γδ T (γδT17) cells, the aberrant activation of which by IL-23 can provoke psoriasis-like inflammation. The skin is also innervated by a meshwork of peripheral nerves consisting of relatively sparse autonomic and abundant sensory fibres. Interactions between the autonomic nervous system and immune cells in lymphoid organs are known to contribute to systemic immunity, but how peripheral nerves regulate cutaneous immune responses remains unclear. We exposed the skin of mice to imiquimod, which induces IL-23-dependent psoriasis-like inflammation. Here we show that a subset of sensory neurons expressing the ion channels TRPV1 and Nav1.8 is essential to drive this inflammatory response. Imaging of intact skin revealed that a large fraction of DDCs, the principal source of IL-23, is in close contact with these nociceptors. Upon selective pharmacological or genetic ablation of nociceptors, DDCs failed to produce IL-23 in imiquimod-exposed skin. Consequently, the local production of IL-23-dependent inflammatory cytokines by dermal γδT17 cells and the subsequent recruitment of inflammatory cells to the skin were markedly reduced. Intradermal injection of IL-23 bypassed the requirement for nociceptor communication with DDCs and restored the inflammatory response. These findings indicate that TRPV1(+)Nav1.8(+) nociceptors, by interacting with DDCs, regulate the IL-23/IL-17 pathway and control cutaneous immune responses.

Changes in sodium channel activity and neuronal hyperexcitability contribute to neuropathic pain, a major clinical problem. There is strong evidence that the re-expression of the embryonic voltage-gated sodium channel subunit Nav1.3 underlies neuronal hyperexcitability and neuropathic pain. Here we show that acute and inflammatory pain behaviour is unchanged in global Nav1.3 mutant mice. Surprisingly, neuropathic pain also developed normally in the Nav1.3 mutant mouse. To rule out any genetic compensation mechanisms that may have masked the phenotype, we investigated neuropathic pain in two conditional Nav1.3 mutant mouse lines. We used Nav1.8-Cre mice to delete Nav1.3 in nociceptors at E14 and NFH-Cre mice to delete Nav1.3 throughout the nervous system postnatally. Again normal levels of neuropathic pain developed after nerve injury in both lines. Furthermore, ectopic discharges from damaged nerves were unaffected by the absence of Nav1.3 in global knock-out mice. Our data demonstrate that Nav1.3 is neither necessary nor sufficient for the development of nerve-injury related pain.

Human acute and inflammatory pain requires the expression of voltage-gated sodium channel Nav1.7 but its significance for neuropathic pain is unknown. Here we show that Nav1.7 expression in different sets of mouse sensory and sympathetic neurons underlies distinct types of pain sensation. Ablating Nav1.7 gene (SCN9A) expression in all sensory neurons using Advillin-Cre abolishes mechanical pain, inflammatory pain and reflex withdrawal responses to heat. In contrast, heat-evoked pain is retained when SCN9A is deleted only in Nav1.8-positive nociceptors. Surprisingly, responses to the hotplate test, as well as neuropathic pain, are unaffected when SCN9A is deleted in all sensory neurons. However, deleting SCN9A in both sensory and sympathetic neurons abolishes these pain sensations and recapitulates the pain-free phenotype seen in humans with SCN9A loss-of-function mutations. These observations demonstrate an important role for Nav1.7 in sympathetic neurons in neuropathic pain, and provide possible insights into the mechanisms that underlie gain-of-function Nav1.7-dependent pain conditions.

Chronic visceral pain affects millions of individuals worldwide and remains poorly understood, with current therapeutic options constrained by gastrointestinal adverse effects. Visceral pain is strongly associated with inflammation and distension of the gut. Here we report that the voltage-gated sodium channel subtype NaV1.9 is expressed in half of gut-projecting rodent dorsal root ganglia sensory neurons. We show that NaV1.9 is required for normal mechanosensation, for direct excitation and for sensitization of mouse colonic afferents by mediators from inflammatory bowel disease tissues, and by noxious inflammatory mediators individually. Excitatory responses to ATP or PGE2 were substantially reduced in NaV1.9(-/-) mice. Deletion of NaV1.9 substantially attenuates excitation and subsequent mechanical hypersensitivity after application of inflammatory soup (IS) (bradykinin, ATP, histamine, PGE2, and 5HT) to visceral nociceptors located in the serosa and mesentery. Responses to mechanical stimulation of mesenteric afferents were also reduced by loss of NaV1.9, and there was a rightward shift in stimulus-response function to ramp colonic distension. By contrast, responses to rapid, high-intensity phasic distension of the colon are initially unaffected; however, run-down of responses to repeat phasic distension were exacerbated in NaV1.9(-/-) afferents. Finally colonic afferent activation by supernatants derived from inflamed human tissue was greatly reduced in NaV1.9(-/-) mice. These results demonstrate that NaV1.9 is required for persistence of responses to intense mechanical stimulation, contributes to inflammatory mechanical hypersensitivity, and is essential for activation by noxious inflammatory mediators, including those from diseased human bowel. These observations indicate that NaV1.9 represents a high-value target for development of visceral analgesics.

The molecular basis of mechanosensory transduction by primary sensory neurones remains poorly understood. Amongst candidate transducer molecules are members of the acid-sensing ion channel (ASIC) family; nerve fibre recordings have shown ASIC2 and ASIC3 null mutants have aberrant responses to suprathreshold mechanical stimuli. Using the neuronal cell body as a model of the sensory terminal we investigated if ASIC2 or 3 contributed to mechanically activated currents in dorsal root ganglion (DRG) neurones. We cultured neurones from ASIC2 and ASIC3 null mutants and compared response properties with those of wild-type controls. Neuronal subpopulations [categorized by cell size, action potential duration and isolectin B4 (IB4) binding] generated distinct responses to mechanical stimulation consistent with their predicted in vivo phenotypes. In particular, there was a striking relationship between action potential duration and mechanosensitivity as has been observed in vivo. Putative low threshold mechanoreceptors exhibited rapidly adapting mechanically activated currents. Conversely, when nociceptors responded they displayed slowly or intermediately adapting currents that were smaller in amplitude than responses of low threshold mechanoreceptor neurones. No differences in current amplitude or kinetics were found between ASIC2 and/or ASIC3 null mutants and controls. Ruthenium red (5 microm) blocked mechanically activated currents in a voltage-dependent manner, with equal efficacy in wild-type and knockout animals. Analysis of proton-gated currents revealed that in wild-type and ASIC2/3 double knockout mice the majority of putative low threshold mechanoreceptors did not exhibit ASIC-like currents but exhibited a persistent current in response to low pH. Our findings are consistent with another ion channel type being important in DRG mechanotransduction.