OBJECTIVE The authors’ laboratory has previously demonstrated beneficial effects of noninvasive low intensity focused ultrasound (liFUS), targeted at the dorsal root ganglion (DRG), for reducing allodynia in rodent neuropathic pain models. However, in rats the DRG is 5 mm below the skin when approached laterally, while in humans the DRG is typically 5–8 cm deep. Here, using a modified liFUS probe, the authors demonstrated the feasibility of using external liFUS for modulation of antinociceptive responses in neuropathic swine. METHODS Two cohorts of swine underwent a common peroneal nerve injury (CPNI) to induce neuropathic pain. In the first cohort, pigs (14 kg) were iteratively tested to determine treatment parameters. liFUS penetration to the L5 DRG was verified by using a thermocouple to monitor tissue temperature changes and by measuring nerve conduction velocity (NCV) at the corresponding common peroneal nerve (CPN). Pain behaviors were monitored before and after treatment. DRG was evaluated for tissue damage postmortem. Based on data from the first cohort, a treatment algorithm was developed, parameter predictions were verified, and neuropathic pain was significantly modified in a second cohort of larger swine (20 kg). RESULTS The authors performed a dose-response curve analysis in 14-kg CPNI swine. Specifically, after confirming that the liFUS probe could reach 5 cm in ex vivo tissue experiments, the authors tested liFUS in 14-kg CPNI swine. The mean ± SEM DRG depth was 3.79 ± 0.09 cm in this initial cohort. The parameters were determined and then extrapolated to larger animals (20 kg), and predictions were verified. Tissue temperature elevations at the treatment site did not exceed 2°C, and the expected increases in the CPN NCV were observed. liFUS treatment eliminated pain guarding in all animals for the duration of follow-up (up to 1 month) and improved allodynia for 5 days postprocedure. No evidence of histological damage was seen using Fluoro-Jade and H&E staining. CONCLUSIONS The results demonstrate that a 5-cm depth can be reached with external liFUS and alters pain behavior and allodynia in a large-animal model of neuropathic pain.

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OBJECTIVE To date, muscular and bone pain have been studied in domestic swine models, but the only neuropathic pain model described in swine is a mixed neuritis model. Common peroneal nerve injury (CPNI) neuropathic pain models have been utilized in both mice and rats. METHODS The authors developed a swine surgical CPNI model of neuropathic pain. Behavioral outcomes were validated with von Frey filament testing, thermal sensitivity assessments, and social and motor scoring. Demyelination of the nerve was confirmed through standard histological assessment. The contralateral nerve served as the control. RESULTS CPNI induced mechanical and thermal allodynia (p < 0.001 [n = 10] and p < 0.05 [n = 4], respectively) and increased pain behavior, i.e., guarding of the painful leg (n = 12). Myelin protein zero (P0) staining revealed demyelination of the ligated nerve upstream of the ligation site. CONCLUSIONS In a neuropathic pain model in domestic swine, the authors demonstrated that CPNI induces demyelination of the common peroneal nerve, which the authors hypothesize is responsible for the resulting allodynic pain behavior. As the anatomical features of domestic swine resemble those of humans more closely than previously used rat and mouse models, utilizing this swine model, which is to the authors’ knowledge the first of its kind, will aid in the translation of experimental treatments to clinical trials.

INTRODUCTION Various rodent models have long been employed to study treatments for chronic pain. However, these studies have resulted in unsuccessful phase-I and II human trials that have failed to result in viable options for patients. Though limitations of animal models are no doubt one issue, lack of objective markers corresponding with pain relief play a role. Our lab has shown significant pain relief in a common peroneal nerve injury (CPNI) rat model following administration of external pulsed low-intensity focused ultrasound (liFUS), thus, establishing external liFUS as a promising technique for treatment of neuropathic pain. Current knowledge of liFUS effects are limited to observable behavioral changes, and little is known of the mechanism of action. To successfully translate this device into the clinic, we examine molecular changes in the inflammatory cascade. METHODS Male rats underwent CPNI to induce neuropathic pain. External liFUS treatment was performed on the L5 dorsal root ganglion (DRG) in the neuropathic model, which was determined from responses to Von Frey fibers (VFF). 24 h post liFUS treatment, L5 DRGs were obtained from 4 distinct cohorts: rats that underwent CPNI with liFUS, CPNI with sham liFUS, sham CPNI with liFUS, and sham CPNI sham liFUS (n = 4 for each group). Using a membrane-based sandwich immunoassay (Proteome Profiler Rat Cytokine Array Kit from R&D System), we assessed relative abundances of 6 anti-inflammatory cytokines and 16 pro-inflammatory cytokines. RESULTS CPNI resulted in an 82.5% decrease of tumor necrosis factor alpha (TNFa) and a 61.8% increase of macrophage inflammatory protein 1-alpha (MIP-1a). liFUS led to a 60% decrease in MIP-1a and a 40% increase in TNFa. Other changes in cytokines were not affected by CPNI or liFUS. CONCLUSION liFUS resulted in similar changes in TNFa and MIP-1a, as compared to spinal cord stimulation and other medical treatments for pain syndromes. Further work will examine inflammatory responses over time and in female rats.