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Compared to proprioceptive afferent collateral projections, less is known about the anatomical, neurochemical, and functional basis of nociceptive collateral projections modulating lumbar central pattern generators (CPG). Quick response times are critical to ensure rapid escape from aversive stimuli. Furthermore, sensitization of nociceptive afferent pathways can contribute to a pathological activation of motor circuits. We investigated the extent and role of collaterals of capsaicin-sensitive nociceptive sacrocaudal afferent (nSCA) nerves that directly ascend several spinal segments in Lissauer's tract and the dorsal column and regulate motor activity. Anterograde tracing demonstrated direct multisegmental projections of the sacral dorsal root 4 (S4) afferent collaterals in Lissauer's tract and in the dorsal column. Subsets of the traced S4 afferent collaterals expressed transient receptor potential vanilloid 1 (TRPV1), which transduces a nociceptive response to capsaicin. Electrophysiological data revealed that S4 dorsal root stimulation could evoke regular rhythmic bursting activity, and our data suggested that capsaicin-sensitive collaterals contribute to CPG activation across multiple segments. Capsaicin's effect on S4-evoked locomotor activity was potent until the lumbar 5 (L5) segments, and diminished in rostral segments. Using calcium imaging we found elevated calcium transients within Lissauer's tract and dorsal column at L5 segments when compared to the calcium transients only within the dorsal column at the lumbar 2 (L2) segments, which were desensitized by capsaicin. We conclude that lumbar locomotor networks in the neonatal mouse spinal cord are targets for modulation by direct multisegmental nSCA, subsets of which express TRPV1 in Lissauer's tract and the dorsal column. J. Comp. Neurol. 521:2870-2887, 2013. © 2013 Wiley Periodicals, Inc.

Dental pain, including toothache, is one of the most prevalent types of orofacial pain, causing severe, persistent pain that has a significant negative effect on quality of life, including eating disturbances, mood changes, and sleep disruption. As the primary cause of toothache pain is injury to the uniquely innervated dental pulp, rodent models of this injury provide the opportunity to study neurobiological mechanisms of tissue injury-induced persistent pain. Here we evaluated behavioral changes in mice with a dental pulp injury (DPI) produced by mechanically exposing the pulp to the oral environment. We monitored the daily life behaviors of mice with DPI, including measures of eating, drinking, and movement. During the first 48 hours, the only parameter affected by DPI was locomotion, which was reduced. There was also a significant short-term decrease in the amount of weight gained by DPI animals that was not related to food consumption. As cold allodynia is frequently observed in individuals experiencing toothache pain, we tested whether mice with DPI demonstrate an aversion to drinking cold liquids using a cold-sucrose consumption test. Surprisingly, mice with DPI increased their consumption of sucrose solution, to over 150% of baseline, regardless of temperature. Both the weight loss and increased sucrose intake in the first 2 days of injury were reversed by administration of indomethacin. These findings indicate that enhanced sucrose consumption may be a reliable measure of orofacial pain in rodents, and suggest that alterations in energy expenditure and motivational behaviors are under-recognized outcomes of tooth injury.

To what extent dorsal horn interneurons contribute to the modality specific processing of pain and itch messages is not known. Here, we report that loxp/cre-mediated CNS deletion of TR4, a testicular orphan nuclear receptor, results in loss of many excitatory interneurons in the superficial dorsal horn but preservation of primary afferents and spinal projection neurons. The interneuron loss is associated with a near complete absence of supraspinally integrated pain and itch behaviors, elevated mechanical withdrawal thresholds and loss of nerve injury-induced mechanical hypersensitivity, but reflex responsiveness to noxious heat, nerve injury-induced heat hypersensitivity, and tissue injury-induced heat and mechanical hypersensitivity are intact. We conclude that different subsets of dorsal horn excitatory interneurons contribute to tissue and nerve injury-induced heat and mechanical pain and that the full expression of supraspinally mediated pain and itch behaviors cannot be generated solely by nociceptor and pruritoceptor activation of projection neurons; concurrent activation of excitatory interneurons is essential.

We previously demonstrated that genetic and/or pharmacological ablation of the TRPV1+/peptidergic or the MrgprD+/non-peptidergic subset of nociceptors produced selective, modality-specific deficits in the behavioural responses to heat and mechanical stimuli, respectively. To assess whether this modality-specific contribution is also manifest at the level of spinal cord neuron responsiveness, here we made extracellular recordings from lumbar dorsal horn neurons of the mouse in response to graded thermal and mechanical stimulation. We found that, following intrathecal injection of capsaicin to eliminate the central terminals of TRPV1+ nociceptors, neurons in the region of laminae I and V of the spinal cord lost responsiveness to noxious heat (whether generated by a contact heat probe or diode laser), with no change in their response to noxious mechanical stimulation. In contrast, ablation of MrgprD+ afferents did not alter the response to noxious heat, but reduced the firing of superficial dorsal horn nociceptive-specific neurons in response to graded mechanical stimulation and decreased the relative number of wide dynamic range neurons that were exclusively mechanosensitive. Neither ablation procedure reduced the number of dorsal horn neurons that responded to noxious cold. These findings support the conclusion that TRPV1+ nociceptors are necessary and probably sufficient for noxious heat activation of dorsal horn neurons and that, despite their polymodal properties, TRPV1+ and MrgprD+ nociceptors provide modality-specific contributions to the response properties of spinal cord neurons.

Natural products that elicit discomfort or pain represent invaluable tools for probing molecular mechanisms underlying pain sensation. Plant-derived irritants have predominated in this regard, but animal venoms have also evolved to avert predators by targeting neurons and receptors whose activation produces noxious sensations. As such, venoms provide a rich and varied source of small molecule and protein pharmacophores that can be exploited to characterize and manipulate key components of the pain-signalling pathway. With this in mind, here we perform an unbiased in vitro screen to identify snake venoms capable of activating somatosensory neurons. Venom from the Texas coral snake (Micrurus tener tener), whose bite produces intense and unremitting pain, excites a large cohort of sensory neurons. The purified active species (MitTx) consists of a heteromeric complex between Kunitz- and phospholipase-A2-like proteins that together function as a potent, persistent and selective agonist for acid-sensing ion channels (ASICs), showing equal or greater efficacy compared with acidic pH. MitTx is highly selective for the ASIC1 subtype at neutral pH; under more acidic conditions (pH < 6.5), MitTx massively potentiates (>100-fold) proton-evoked activation of ASIC2a channels. These observations raise the possibility that ASIC channels function as coincidence detectors for extracellular protons and other, as yet unidentified, endogenous factors. Purified MitTx elicits robust pain-related behaviour in mice by activation of ASIC1 channels on capsaicin-sensitive nerve fibres. These findings reveal a mechanism whereby snake venoms produce pain, and highlight an unexpected contribution of ASIC1 channels to nociception.

Mechanical pain contributes to the morbidity associated with inflammation and trauma, but primary sensory neurons that convey the sensation of acute and persistent mechanical pain have not been identified. Dorsal root ganglion (DRG) neurons transmit sensory information to the spinal cord using the excitatory transmitter glutamate, a process that depends on glutamate transport into synaptic vesicles for regulated exocytotic release. Here we report that a small subset of cells in the DRG expresses the low abundance vesicular glutamate transporter VGLUT3 (also known as SLC17A8). In the dorsal horn of the spinal cord, these afferents project to lamina I and the innermost layer of lamina II, which has previously been implicated in persistent pain caused by injury. Because the different VGLUT isoforms generally have a non-redundant pattern of expression, we used Vglut3 knockout mice to assess the role of VGLUT3(+) primary afferents in the behavioural response to somatosensory input. The loss of VGLUT3 specifically impairs mechanical pain sensation, and in particular the mechanical hypersensitivity to normally innocuous stimuli that accompanies inflammation, nerve injury and trauma. Direct recording from VGLUT3(+) neurons in the DRG further identifies them as a poorly understood population of unmyelinated, low threshold mechanoreceptors (C-LTMRs). The analysis of Vglut3(-/-) mice now indicates a critical role for C-LTMRs in the mechanical hypersensitivity caused by injury.

The anti- migraine action of "triptan" drugs involves the activation of serotonin subtype 1D (5-HT1D) receptors expressed on "painresponsive" trigeminal primary afferents. In the central terminals of these nociceptors, the receptor is concentrated on peptidergic dense core vesicles (DCVs) and is notably absent from the plasma membrane. Based on this arrangement, we hypothesized that in the resting state the receptor is not available for binding by a triptan, but that noxious stimulation of these afferents could trigger vesicular release of DCVs, thus externalizing the receptor. Here we report that within 5 min of an acute mechanical stimulus to the hindpaw of the rat, there is a significant increase of 5-HT1D-immunoreactivity (IR) in the ipsilateral dorsal horn of the spinal cord. We suggest that these rapid immunohistochemical changes reflect redistribution of sequestered receptor to the plasma membrane, where it is more readily detected. We also observed divergent changes in 5-HT1D-IR in inflammatory and nerve-injury models of persistent pain, occurring at least in part through the regulation of 5-HT1D-receptor gene expression. Finally, we found that 5-HT1D-IR is unchanged in the spinal cord dorsal horn of mice with a deletion of the gene encoding the neuropeptide substance P. This result differs from that reported for the partial derivative-opioid receptor, which is also sorted to DCVs, but is greatly reduced in preprotachykinin mutant mice. We suggest that a "pain"-triggered regulation of 5-HT1D-receptor expression underlies the effectiveness of triptans for the treatment of migraine. Moreover, the widespread expression of 5-HT1D receptor in somatic nociceptive afferents suggests that triptans could, in certain circumstances, treat pain in nontrigeminal regions of the body.

Author(s): Jeremiah D. Osteen [1]; Volker Herzig [2]; John Gilchrist [3]; Joshua J. Emrick [1]; Chuchu Zhang [1]; Xidao Wang [4]; Joel Castro [5, 6]; Sonia Garcia-Caraballo [5, 6]; Luke [...], Voltage-gated sodium (Na[sub.v]) channels initiate action potentials in most neurons, including primary afferent nerve fibres of the pain pathway. Local anaesthetics block pain through non-specific actions at all Na[sub.v] channels, but the discovery of selective modulators would facilitate the analysis of individual subtypes of these channels and their contributions to chemical, mechanical, or thermal pain. Here we identify and characterize spider (Heteroscodra maculata) toxins that selectively activate the Na[sub.v]1.1 subtype, the role of which in nociception and pain has not been elucidated. We use these probes to show that Na[sub.v]1.1-expressing fibres are modality-specific nociceptors: their activation elicits robust pain behaviours without neurogenic inflammation and produces profound hypersensitivity to mechanical, but not thermal, stimuli. In the gut, high-threshold mechanosensitive fibres also express Na[sub.v]1.1 and show enhanced toxin sensitivity in a mouse model of irritable bowel syndrome. Together, these findings establish an unexpected role for Na[sub.v]1.1 channels in regulating the excitability of sensory nerve fibres that mediate mechanical pain.