T. Sheahan

Voluntary Exercise Training: Analysis of Mice in Uninjured, Inflammatory, and Nerve-Injured Pain States.

Both clinical and animal studies suggest that exercise may be an effective way to manage inflammatory and neuropathic pain conditions. However, existing animal studies commonly use forced exercise paradigms that incorporate varying degrees of stress, which itself can elicit analgesia, and thus may complicate the interpretation of the effects of exercise on pain. We investigated the analgesic potential of voluntary wheel running in the formalin model of acute inflammatory pain and the spared nerve injury model of neuropathic pain in mice. In uninjured, adult C57BL/6J mice, 1 to 4 weeks of exercise training did not alter nociceptive thresholds, lumbar dorsal root ganglia neuronal excitability, or hindpaw intraepidermal innervation. Further, exercise training failed to attenuate formalin-induced spontaneous pain. Lastly, 2 weeks of exercise training was ineffective in reversing spared nerve injury-induced mechanical hypersensitivity or in improving muscle wasting or hindpaw denervation. These findings indicate that in contrast to rodent forced exercise paradigms, short durations of voluntary wheel running do not improve pain-like symptoms in mouse models of acute inflammation and peripheral nerve injury.

Surgical extraction of human dorsal root ganglia from organ donors and preparation of primary sensory neuron cultures

Primary cultures of rodent sensory neurons are widely used to investigate the cellular and molecular mechanisms involved in pain, itch, nerve injury and regeneration. However, translation of these preclinical findings may be greatly improved by direct validation in human tissues. We have developed an approach to extract and culture human sensory neurons in collaboration with a local organ procurement organization (OPO). Here we describe the surgical procedure for extraction of human dorsal root ganglia (hDRG) and the necessary modifications to existing culture techniques to prepare viable adult human sensory neurons for functional studies. Dissociated sensory neurons can be maintained in culture for >10 d, and they are amenable to electrophysiological recording, calcium imaging and viral gene transfer. The entire process of extraction and culturing can be completed in <7 h, and it can be performed by trained graduate students. This approach can be applied at any institution with access to organ donors consenting to tissue donation for research, and is an invaluable resource for improving translational research.

Inflammation and nerve injury minimally affect mouse voluntary behaviors proposed as indicators of pain

Highlights • Inflammation suppressed wheel running and locomotion, and impaired gait in mice.• Nerve injury gave rise to gait deficits that are likely motor-, not pain-related.• Changes in wheel running or gait were unrelated to the degree of hypersensitivity.• Neither inflammation nor nerve injury altered social interactions or anxiety-like behavior.

Metabotropic Glutamate Receptor 2/3 (mGluR2/3) Activation Suppresses TRPV1 Sensitization in Mouse, But Not Human, Sensory Neurons

Abstract The use of human tissue to validate putative analgesic targets identified in rodents is a promising strategy for improving the historically poor translational record of preclinical pain research. We recently demonstrated that in mouse and human sensory neurons, agonists for metabotropic glutamate receptors 2 and 3 (mGluR2/3) reduce membrane hyperexcitability produced by the inflammatory mediator prostaglandin E2 (PGE2). Previous rodent studies indicate that mGluR2/3 can also reduce peripheral sensitization by suppressing inflammation-induced sensitization of TRPV1. Whether this observation similarly translates to human sensory neurons has not yet been tested. We found that activation of mGluR2/3 with the agonist APDC suppressed PGE2-induced sensitization of TRPV1 in mouse, but not human, sensory neurons. We also evaluated sensory neuron expression of the gene transcripts for mGluR2 (Grm2), mGluR3 (Grm3), and TRPV1 (Trpv1). The majority of Trpv1 + mouse and human sensory neurons expressed Grm2 and/or Grm3, and in both mice and humans, Grm2 was expressed in a greater percentage of sensory neurons than Grm3. Although we demonstrated a functional difference in the modulation of TRPV1 sensitization by mGluR2/3 activation between mouse and human, there were no species differences in the gene transcript colocalization of mGluR2 or mGluR3 with TRPV1 that might explain this functional difference. Taken together with our previous work, these results suggest that mGluR2/3 activation suppresses only some aspects of human sensory neuron sensitization caused by PGE2. These differences have implications for potential healthy human voluntary studies or clinical trials evaluating the analgesic efficacy of mGluR2/3 agonists or positive allosteric modulators.

Macrophage-to-sensory neuron crosstalk mediated by Angiotensin II type-2 receptor elicits neuropathic pain

Peripheral nerve damage initiates a complex series of cellular and structural processes that culminate in chronic neuropathic pain. Our study defines local angiotensin signaling via activation of the Angiotensin II (Ang II) type-2 receptor (AT2R) on macrophages as the critical trigger of neuropathic pain. An AT2R-selective antagonist attenuates neuropathic, but not inflammatory pain hypersensitivity in mice, and requires the cell damage-sensing ion channel transient receptor potential family-A member-1 (TRPA1). Mechanical and cold pain hypersensitivity that are characteristic of neuropathic conditions can be attenuated by chemogenetic depletion of peripheral macrophages and AT2R-null hematopoietic cell transplantation. Our findings show no AT2R expression in mouse or human sensory neurons, rather AT2R expression and activation in macrophages triggers production of reactive oxygen/nitrogen species, which trans-activate TRPA1 on sensory neurons. Our study defines the precise neuro-immune crosstalk underlying nociceptor sensitization at the site of nerve injury. This form of cell-to-cell signaling represents a critical peripheral mechanism for chronic neuropathic pain, and therefore identifies multiple analgesic targets.