Animal Studies

Previous studies suggested that reactive oxygen species (ROS) produced by NADPH oxidase 4 (Nox4) affect the processing of neuropathic pain. However, mechanisms underlying Nox4-dependent pain signaling are incompletely understood. In this study, we aimed to identify novel Nox4 downstream interactors in the nociceptive system. Mice lacking Nox4 specifically in sensory neurons were generated by crossing Advillin-Cre mice with Nox4 mice. Tissue-specific deletion of Nox4 in sensory neurons considerably reduced mechanical hypersensitivity and neuronal action potential firing after peripheral nerve injury. Using a proteomic approach, we detected various proteins that are regulated in a Nox4-dependent manner after injury, including the small calcium-binding protein S100A4. Immunofluorescence staining and western blot experiments confirmed that S100A4 expression is massively up-regulated in peripheral nerves and dorsal root ganglia after injury. Furthermore, mice lacking S100A4 showed increased mechanical hypersensitivity after peripheral nerve injury and after delivery of a ROS donor. Our findings suggest that S100A4 expression is up-regulated after peripheral nerve injury in a Nox4-dependent manner and that deletion of S100A4 leads to an increased neuropathic pain hypersensitivity.

Although macrophages (MΦ) are known to play a central role in neuropathic pain, their contribution to cancer pain has not been established. Here we report that depletion of sciatic nerve resident MΦs (rMΦ) in mice attenuates mechanical/cold hypersensitivity and spontaneous pain evoked by intraplantar injection of melanoma or lung carcinoma cells. MΦ-colony stimulating factor (M-CSF) was upregulated in the sciatic nerve trunk and mediated cancer-evoked pain via rMΦ expansion, transient receptor potential ankyrin 1 (TRPA1) activation, and oxidative stress. Targeted deletion of Trpa1 revealed a key role for Schwann cell TRPA1 in sciatic nerve rMΦ expansion and pain-like behaviors. Depletion of rMΦs in a medial portion of the sciatic nerve prevented pain-like behaviors. Collectively, we identified a feed-forward pathway involving M-CSF, rMΦ, oxidative stress and Schwann cell TRPA1 that operates throughout the nerve trunk to signal cancer-evoked pain.

Accumulating evidence suggests hippocampal impairment under chronic pain phenotype. However, it is unknown whether neuropathic behaviors are related to dysfunction of the hippocampal circuitry. Here we enhanced hippocampal activity by pharmacological, opto- and chemo-genetic techniques to determine hippocampal influence on neuropathic pain behaviors. We found that excitation of the dorsal (DH), but not the ventral (VH), hippocampus induces analgesia in two rodent models of neuropathic pain (SNI and SNL), and in rats and mice. Optogenetic and pharmacological manipulations of DH neurons demonstrated that DH-induced analgesia was mediated by NMDA and μ-opioid receptors. In addition to analgesia, optogenetic stimulation of DH in SNI mice also resulted in enhanced real-time conditioned place preference for the chamber where DH was activated, a finding consistent with pain relief. Similar manipulations in VH were ineffective. Using chemo-fMRI, where awake resting-state fMRI was combined with viral vector mediated chemogenetic activation (PSAM/PSEM) of DH neurons, we demonstrated changes of functional connectivity between DH and thalamus and somatosensory regions that tracked the extent of relief from tactile allodynia. Moreover, we examined hippocampal functional connectivity in humans, and observe differential reorganization of its anterior and posterior subdivisions between subacute and chronic backpain. Altogether, these results imply that downregulation of DH circuitry during chronic neuropathic pain aggravates pain-related behaviors. Conversely, activation of DH reverses pain-related behaviors through local excitatory and opioidergic mechanisms affecting DH functional connectivity. Thus, the present study exhibits a novel causal role for the DH but not VH in controlling neuropathic pain-related behaviors.

Neuropeptide Y Y1 receptor-expressing neurons in the dorsal horn of the spinal cord contribute to chronic pain. For the first time, we characterized the firing patterns of Y1-expressing neurons in Y1eGFP reporter mice. Under hyperpolarized conditions, most Y1eGFP neurons exhibited fast A-type potassium currents and delayed, short-latency firing (DSLF). Y1eGFP DSLF neurons were almost always rapidly adapting and often exhibited rebound spiking, characteristics of spinal pain neurons under the control of T-type calcium channels. These results inspire future studies to determine whether tissue or nerve injury downregulates the channels that underlie A-currents, thus unmasking membrane hyperexcitability in Y1-expressing dorsal horn neurons, leading to persistent pain.

The two voltage gated sodium channels Nav1.7 and Nav1.8 are expressed in the peripheral nervous system and involved in various pain conditions including inflammatory and neuropathic pain. Rodent models bearing deletions or mutations of the corresponding genes, Scn9a and Scn10a, were created in order to understand the role of these channels in the pathophysiological mechanism underlying pain symptoms. This review summarizes the pain behavior profiles reported in Scn9a and Scn10a rodent models. The complete loss-of-function or knockout (KO) of Scn9a or Scn10a and the conditional KO (cKO) of Scn9a in specific cell populations were shown to decrease sensitivity to various pain stimuli. The Possum mutant mice bearing a dominant hypermorphic mutation in Scn10a revealed higher sensitivity to noxious stimuli. Several gain-of-function mutations were identified in patients with painful small fiber neuropathy. Future knowledge obtained from preclinical models bearing these mutations will allow understanding how these mutations affect pain. In addition, the review gives perspectives for creating models that better mimic patients' pain symptoms in view to developing novel analgesic strategies.