Substance P is released in the spinal cord in response to painful stimuli, but its role in nociceptive signaling remains unclear. When a conjugate of substance P and the ribosome-inactivating protein saporin was infused into the spinal cord, it was internalized and cytotoxic to lamina I spinal cord neurons that express the substance P receptor. This treatment left responses to mild noxious stimuli unchanged, but markedly attenuated responses to highly noxious stimuli and mechanical and thermal hyperalgesia. Thus, lamina I spinal cord neurons that express the substance P receptor play a pivotal role in the transmission of highly noxious stimuli and the maintenance of hyperalgesia.

Activation of tetrodotoxin-resistant sodium channels contributes to action potential electrogenesis in neurons. Antisense oligonucleotide studies directed against Nav1.8 have shown that this channel contributes to experimental inflammatory and neuropathic pain. We report here the discovery of A-803467, a sodium channel blocker that potently blocks tetrodotoxin-resistant currents (IC50 ‫؍‬ 140 nM) and the generation of spontaneous and electrically evoked action potentials in vitro in rat dorsal root ganglion neurons. In recombinant cell lines, A-803467 potently blocked human Nav1.8 (IC50 ‫؍‬ 8 nM) and was >100-fold selective vs. human Nav1.2, Nav1.3, Nav1.5, and Nav1.

The cancer-related event that is most disruptive to the cancer patient's quality of life is pain. To begin to define the mechanisms that give rise to cancer pain, we examined the neurochemical changes that occur in the spinal cord and associated dorsal root ganglia in a murine model of bone cancer. Twenty-one days after intramedullary injection of osteolytic sarcoma cells into the femur, there was extensive bone destruction and invasion of the tumor into the periosteum, similar to that found in patients with osteolytic bone cancer. In the spinal cord, ipsilateral to the cancerous bone, there was a massive astrocyte hypertrophy without neuronal loss, an expression of dynorphin and c-Fos protein in neurons in the deep laminae of the dorsal horn. Additionally, normally non-noxious palpation of the bone with cancer induced behaviors indicative of pain, the internalization of the substance P receptor, and c-Fos expression in lamina I neurons. The alterations in the neurochemistry of the spinal cord and the sensitization of primary afferents were positively correlated with the extent of bone destruction and the growth of the tumor. This "neurochemical signature" of bone cancer pain appears unique when compared to changes that occur in persistent inflammatory or neuropathic pain states. Understanding the mechanisms by which the cancer cells induce this neurochemical reorganization may provide insight into peripheral factors that drive spinal cord plasticity and in the development of more effective treatments for cancer pain.

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P2X3 and P2X2/3 receptors are highly localized on peripheral and central processes of sensory afferent nerves, and activation of these channels contributes to the pronociceptive effects of ATP. A-317491 is a novel non-nucleotide antagonist of P2X3 and P2X2/3 receptor activation. A-317491 potently blocked recombinant human and rat P2X3 and P2X2/3 receptor-mediated calcium flux (Ki ‫؍‬ 22-92 nM) and was highly selective (IC50 >10 M) over other P2 receptors and other neurotransmitter receptors, ion channels, and enzymes. A-317491 also blocked native P2X3 and P2X2/3 receptors in rat dorsal root ganglion neurons. Blockade of P2X3 containing channels was stereospecific because the R-enantiomer (A-317344) of A-317491 was significantly less active at P2X3 and P2X2/3 receptors. A-317491 dosedependently (ED50 ‫؍‬ 30 mol͞kg s.c.) reduced complete Freund's adjuvant-induced thermal hyperalgesia in the rat. A-317491 was most potent (ED50 ‫؍‬ 10 -15 mol͞kg s.c.) in attenuating both thermal hyperalgesia and mechanical allodynia after chronic nerve constriction injury. The R-enantiomer, A-317344, was inactive in these chronic pain models. Although active in chronic pain models, A-317491 was ineffective (ED 50 >100 mol͞kg s.c.) in reducing nociception in animal models of acute pain, postoperative pain, and visceral pain. The present data indicate that a potent and selective antagonist of P2X 3 and P2X2/3 receptors effectively reduces both nerve injury and chronic inflammatory nociception, but P2X 3 and P2X2/3 receptor activation may not be a major mediator of acute, acute inflammatory, or visceral pain.T he cloning and characterization of the P2X 3 receptor, a specific ATP-sensitive ligand-gated ion channel that is selectively localized on peripheral and central processes of sensory afferent neurons (1-3), has generated much interest in the role of this receptor in nociceptive signaling (4). The discovery of the P2X 3 receptor has provided a putative mechanism for previous reports that ATP, released from sensory nerves (5), produces fast excitatory potentials in dorsal root ganglion (DRG) neurons (6). These actions appear to be physiologically relevant because iontophoretic application of ATP to human skin elicits pain (7) and exogenously applied ATP enhances pain sensations in a human blister base model (8).The P2X 3 receptor is natively expressed as a functional homomer and as a heteromultimeric combination with the P2X 2 (P2X 2/3 ) receptor (1, 2, 9). Both P2X 3 -containing channels are expressed on a high proportion of isolectin IB4-positive neurons in DRG (3, 10). These receptors share similar pharmacological profiles (11), but differ in their acute desensitization kinetics (10, 12). Immunohistochemical studies have shown that P2X 3 receptor expression is up-regulated in DRG neurons and ipsilateral spinal cord after chronic constriction injury (CCI) of the sciatic nerve (13). Additionally, CCI results in a specific ectopic sensitivity to ATP that is not observed on contralateral (uninjured) nerves (14).Recently, the phenotyp...

ATP-sensitive P2X 7 receptors are localized on cells of immunological origin including glial cells in the central nervous system. Activation of P2X 7 receptors leads to rapid changes in intracellular calcium concentrations, release of the proinflammatory cytokine interleukin-1␤ (IL-1␤), and following prolonged agonist exposure, cytolytic plasma membrane pore formation. P2X 7 knockout mice show reduced inflammation as well as decreased nociceptive sensitivity following peripheral nerve injury. A-740003 (N-(1-{[(cyanoimino)(5-quinolinylamino) methyl] amino}-2,2-dimethylpropyl)-2-(3,4-dimethoxyphenyl)acetamide) is a novel competitive antagonist of P2X 7 receptors (IC 50 values ϭ 40 nM for human and 18 nM for rat) as measured by agonist-stimulated changes in intracellular calcium concentrations. A-740003 showed weak or no activity (IC 50 Ͼ 10 M) at other P2 receptors and an array of other neurotransmitter and peptide receptors, ion channels, reuptake sites, and enzymes.A-740003 potently blocked agonist-evoked IL-1␤ release (IC 50 ϭ 156 nM) and pore formation (IC 50 ϭ 92 nM) in differentiated human THP-1 cells. Systemic administration of A-740003 produced dose-dependent antinociception in a spinal nerve ligation model (ED 50 ϭ 19 mg/kg i.p.) in the rat. A-740003 also attenuated tactile allodynia in two other models of neuropathic pain, chronic constriction injury of the sciatic nerve and vincristine-induced neuropathy. In addition, A-740003 effectively reduced thermal hyperalgesia observed following intraplantar administration of carrageenan or complete Freund's adjuvant (ED 50 ϭ 38 -54 mg/kg i.p.). A-740003 was ineffective in attenuating acute thermal nociception in normal rats and did not alter motor performance at analgesic doses. These data demonstrate that selective blockade of P2X 7 receptors in vivo produces significant antinociception in animal models of neuropathic and inflammatory pain.

Substance P receptor (SPR)-expressing spinal neurons were ablated with the selective cytotoxin substance P-saporin. Loss of these neurons resulted in a reduction of thermal hyperalgesia and mechanical allodynia associated with persistent neuropathic and inflammatory pain states. This loss appeared to be permanent. Responses to mildly painful stimuli and morphine analgesia were unaffected by this treatment. These results identify a target for treating persistent pain and suggest that the small population of SPR-expressing neurons in the dorsal horn of the spinal cord plays a pivotal role in the generation and maintenance of chronic neuropathic and inflammatory pain.

Bone cancer pain is common among cancer patients and can have a devastating effect on their quality of life. A chief problem in designing new therapies for bone cancer pain is that it is unclear what mechanisms drive this distinct pain condition. Here we show that osteoprotegerin, a secreted 'decoy' receptor that inhibits osteoclast activity, also blocks behaviors indicative of pain in mice with bone cancer. A substantial part of the actions of osteoprotegerin seems to result from inhibition of tumor-induced bone destruction that in turn inhibits the neurochemical changes in the spinal cord that are thought to be involved in the generation and maintenance of cancer pain. These results demonstrate that excessive tumor-induced bone destruction is involved in the generation of bone cancer pain and that osteoprotegerin may provide an effective treatment for this common human condition.