Peripheral initiators of muscle pain are virtually unknown, but likely key to development of chronic pain after muscle insult. The current study tested the hypothesis that ASIC3 in muscle is necessary for development of cutaneous mechanical, but not heat hyperalgesia induced by muscle inflammation. Using mechanical and heat stimuli, we assessed behavioral responses in ASIC3−/− and ASIC3+/+ mice after induction of carrageenan muscle inflammation. ASIC3−/−mice did not develop cutaneous mechanical hyperalgesia after muscle inflammation when compared to ASIC3+/ + mice; heat hyperalgesia developed similarly between groups. We then tested if the phenotype could be rescued in ASIC3−/− mice by using a recombinant herpes virus vector to express ASIC3 in skin (where testing occurred) or muscle (where inflammation occurred). Infection of mouse DRG neurons with ASIC3-encoding virus resulted in functional expression of ASICs. Injection of ASIC3-encoding virus into muscle or skin of ASIC3−/− mice resulted in ASIC3 mRNA in DRG and protein expression in DRG and the peripheral injection site. Injection of ASIC3-encoding virus into muscle, but not skin, resulted in development of mechanical hyperalgesia similar to that observed in ASIC3+/+ mice. Thus, ASIC3 in primary afferent fibers innervating muscle is critical to development of hyperalgesia that results from muscle insult.

To test the utility of gene therapeutic approaches for the treatment of pain, a recombinant herpes simplex virus, type 1, has been engineered to contain the cDNA for an opioid peptide precursor, human preproenkephalin, under control of the human cytomegalovirus promoter. This virus and a similar recombinant containing the Escherichia coli lacZ gene were applied to the abraded skin of the dorsal hindpaw of mice. After infection, the presence of beta-galactosidase in neuronal cell bodies of the relevant spinal ganglia (lacZ-containing virus) and of human proenkephalin (preproenkephalin-encoding virus) in the central terminals of these neurons indicated appropriate gene delivery and expression. Baseline foot withdrawal responses to noxious radiant heat mediated by Adelta and C fibers were similar in animals infected with proenkephalin-encoding and beta-galactosidase-encoding viruses. Sensitization of the foot withdrawal response after application of capsaicin (C fibers) or dimethyl sulfoxide (Adelta fibers) observed in control animals was reduced or eliminated in animals infected with the proenkephalin-encoding virus for at least 7 weeks postinfection. Hence, preproenkephalin cDNA delivery selectively blocked hyperalgesia without disrupting baseline sensory neurotransmission. This blockade of sensitization was reversed by administration of the opioid antagonist naloxone, apparently acting in the spinal cord. The results demonstrate that the function of sensory neurons can be selectively altered by viral delivery of a transgene. Because hyperalgesic mechanisms may be important in establishing and maintaining neuropathic and other chronic pain states, this approach may be useful for treatment of chronic pain and hyperalgesia in humans.

Behavioral nociceptive responses evoked by relatively high rates of noxious radiant skin heating appear to be mediated by A delta nociceptor activation, whereas responses evoked by low rates of skin heating appear to be mediated by the activation of C-fiber nociceptors. This hypothesis was confirmed by the results of single unit recordings of A delta and C nociceptive afferent fibers isolated from the saphenous nerves of pentobarbital anesthetized rats. Heating the hind paw skin of the rat at a relatively high rate of 6.5 degrees C/sec activated A delta units within 2 sec after the onset of the stimulus. This response latency is similar to the 2.5 sec latency of the foot withdrawal response to a similar stimulus. In contrast, C-fibers were only slightly activated at a longer latency of 5-6 sec. Conversely, heating the hind paw skin at a relatively low rate of 0.9 degrees C/sec activated C-fibers, but evoked only a few action potentials in A delta nociceptors. C-fibers began firing at a rate less than 1 Hz between 8 and 10 sec after the onset of heating and fired at a mean rate of 1.5 Hz between 10 and 12 sec, which corresponds to the latency of the foot withdrawal response. Topical application of capsaicin to the hind paw skin decreased the latency of C-fiber responses from control values of 8-12 sec to approximately 4 sec after topical capsaicin treatment. The mean latency of the foot withdrawal response to skin heating at the low rate is also reduced from control values of 12-14 sec to 4-5 sec after capsaicin treatment. In contrast, capsaicin treatment did not significantly affect the responses of A delta nociceptors. These results support the conclusion that nociceptive foot withdrawal responses to a low rate of skin heating are mediated predominantly by the activation of C-fiber nociceptors. These results provide direct evidence that, under the conditions of these experiments, nociceptive foot withdrawal responses evoked by high rates of skin heating are primarily mediated by A delta nociceptors, and foot withdrawal responses evoked by low rates of skin heating are primarily mediated by C-fiber nociceptors.

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Several lines of evidence suggest that different classes of nociceptive afferents mediate the responses produced by different rates of noxious skin heating. More specifically, low skin heating rates evoke nociceptive responses that appear to be mediated by the activation of capsaicin-sensitive C-fiber nociceptors, whereas high skin heating rates appear to produce responses mediated by the activation of other nociceptors. This hypothesis was examined by both electrophysiological and behavioral experiments. This report describes the results of experiments designed to determine whether pharmacologic treatments that selectively alter the activity of C-fiber nociceptive afferents also produce selective effects on foot withdrawal responses to either high or low rates of noxious foot heating. The results of these experiments demonstrate that: (1) topical application of a low concentration of capsaicin, which sensitizes C-fiber nociceptors, selectively decreased the latency of responses to low heating rates; (2) topical application of a high concentration of capsaicin, that desensitizes C-fiber nociceptors, selectively increased the latency of responses to low heating rates; (3) low doses of systemic morphine, which selectively attenuate nociception produced by the activation of C-fiber nociceptors, selectively increased response latencies for low skin heating rates. These results support the conclusion that foot withdrawal responses evoked by low skin heating rates are mediated by the activation of capsaicin-sensitive C-fiber nociceptors and foot withdrawal responses evoked by high skin heating rates are mediated by the activation of other nociceptors. This conclusion is supported by the results of the accompanying electrophysiological study which provides direct evidence that low rates of skin heating preferentially activate C-fiber nociceptors while high rates of skin heating preferentially activate A delta nociceptors.

The tissue directly surrounding incisions in mouse hind paws undergoes large changes in the content of specific cytokines in addition to demonstrating edema and nociceptive sensitization. By blocking the receptor for one component of the complement system, C5a, all of these changes can be reduced. Complement receptor inhibitors may constitute a novel group of compounds useful in reducing the pain and swelling of surgical incisions.

Background: Inflammation and nociceptive sensitization are hallmarks of tissue surrounding surgical incisions. Recent studies demonstrate that several cytokines may participate in the enhancement of nociception near these wounds. Since opioids like morphine interact with neutrophils and other immunocytes, it is possible that morphine exerts some of its antinociceptive action after surgical incision by altering the vigor of the inflammatory response. On the other hand, keratinocytes also express opioid receptors and have the capacity to produce cytokines after injury. Our studies were directed towards determining if opioids alter cytokine production near incisions and to identify cell populations responsible for producing these cytokines.

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