resiniferatoxin and Small-Fiber-Neuropathy

Neurotrophic factors and their corresponding receptors play key roles in the maintenance of different phenotypic dorsal root ganglion (DRG) neurons, the axons of which degenerate in small fiber neuropathy, leading to various neuropathic manifestations. Mechanisms underlying positive and negative symptoms of small fiber neuropathy have not been systematically explored. This study investigated the molecular basis of these seemingly paradoxical neuropathic behaviors according to the profiles of TrkA and Ret with immunohistochemical and pharmacological interventions in a mouse model of resiniferatoxin (RTX)-induced small fiber neuropathy. Mice with RTX neuropathy exhibited thermal hypoalgesia and mechanical allodynia, reduced skin innervation, and altered DRG expression profiles with decreased TrkA(+) neurons and increased Ret(+) neurons. RTX neuropathy induced the expression of activating transcription factor 3 (ATF3), and ATF3(+) neurons were colocalized with Ret but not with TrkA (P<0.001). As a neuroprotectant, 4-Methylcatechol (4MC) promoted skin reinnervation partially with correlated reversal of the neuropathic behaviors and altered neurochemical expression. Gambogic amide, a selective TrkA agonist, normalized thermal hypoalgesia, and GW441756, a TrkA kinase inhibitor, induced thermal hypoalgesia, which was already reversed by 4MC. Mechanical allodynia was reversed by a Ret kinase inhibitor, AST487, which induced thermal hyperalgesia in naïve mice. The activation of Ret signaling by XIB4035 induced mechanical allodynia and thermal hypoalgesia in RTX neuropathy mice in which the neuropathic behaviors were previously normalized by 4MC. Distinct neurotrophic factor receptors, TrkA and Ret, accounted for negative and positive neuropathic behaviors in RTX-induced small fiber neuropathy, respectively: TrkA for thermal hypoalgesia and Ret for mechanical allodynia and thermal hypoalgesia.

Topics: Animals; Disease Models, Animal; Diterpenes; Mice; Pain Measurement; Proto-Oncogene Proteins c-ret; Quinolines; Receptor, trkA; Small Fiber Neuropathy; Xanthones

Patients with diabetes mellitus (DM) or those experiencing the neurotoxic effects of chemotherapeutic agents may develop sensation disorders due to degeneration and injury of small-diameter sensory neurons, referred to as small fiber neuropathy. Present animal models of small fiber neuropathy affect both large- and small-diameter sensory fibers and thus create a neuropathology too complex to properly assess the effects of injured small-diameter sensory fibers. Therefore, it is necessary to develop an experimental model of pure small fiber neuropathy to adequately examine these issues. This protocol describes an experimental model of small fiber neuropathy specifically affecting small-diameter sensory nerves with resiniferatoxin (RTX), an ultrapotent agonist of transient receptor potential vanilloid type 1 (TRPV1), through a single dose of intraperitoneal injection, referred to as RTX neuropathy. This RTX neuropathy showed pathological manifestations and behavioral abnormalities that mimic the clinical characteristics of patients with small fiber neuropathy, including intraepidermal nerve fiber (IENF) degeneration, specifically injury in small-diameter neurons, and induction of thermal hypoalgesia and mechanical allodynia. This protocol tested three doses of RTX (200, 50, and 10 µg/kg, respectively) and concluded that a critical dose of RTX (50 µg/kg) is required for the development of typical small fiber neuropathy manifestations, and prepared a modified immunostaining procedure to investigate IENF degeneration and neuronal soma injury. The modified procedure is fast, systematic, and economic. Behavioral evaluation of neuropathic pain is critical to reveal the function of small-diameter sensory nerves. The evaluation of mechanical thresholds in experimental rodents is particularly challenging and this protocol describes a customized metal mesh that is suitable for this type of assessment in rodents. In summary, RTX neuropathy is a new and easily established experimental model to evaluate the molecular significance and intervention underlying neuropathic pain for the development of therapeutic agents.

Topics: Animals; Disease Models, Animal; Diterpenes; Male; Mice; Mice, Inbred ICR; Nerve Fibers; Nociception; Small Fiber Neuropathy; TRPV Cation Channels

The neurochemical effects of adenosine signaling in small-fiber neuropathy leading to neuropathic pain are yet to be explored in a direct manner. This study examined this system at the level of ligand (through the ectonucleotidase activity of prostatic acid phosphatase [PAP]) and adenosine A1 receptors (A1Rs) in resiniferatoxin (RTX) neuropathy, a peripheral neurodegenerative disorder that specifically affects nociceptive nerves expressing transient receptor potential vanilloid type 1 (TRPV1). We conducted immunohistochemistry on dorsal root ganglion (DRG) neurons, high-performance liquid chromatography for functional assays, and pharmacological interventions to alter PAP and A1Rs in mice with RTX neuropathy. In DRG of RTX neuropathy, PAP(+) neurons were reduced compared with vehicle-treated mice (P = 0.002). Functionally, PAP ectonucleotidase activity was consequently reduced (ie, the content of adenosine in DRG, P = 0.012). PAP(+) neuronal density was correlated with the degree of mechanical allodynia, which was reversed by intrathecal (i.t.) lumbar puncture injection of recombinant PAP with a dose-dependent effect. Furthermore, A1Rs were downregulated (P = 0.002), and this downregulation was colocalized with the TRPV1 receptor (31.0% ± 2.8%). Mechanical allodynia was attenuated in a dose-dependent response by i.t. injection of the A1R ligand, adenosine; however, no analgesia was evident when an exogenous adenosine was blocked by A1R antagonist. This study demonstrated dual mechanisms of neuropathic pain in TRPV1-induced neuropathy, involving a reduced adenosine system at both the ligand (adenosine) and receptor (A1Rs) levels.

Topics: Adenosine; Animals; Diterpenes; Down-Regulation; Ganglia, Spinal; Mice; Neuralgia; Neurons; Neurotoxins; Pain Measurement; Receptor, Adenosine A1; Small Fiber Neuropathy

Topics: Animals; Disease Models, Animal; Diterpenes; Male; Mice; Pain Measurement; Proto-Oncogene Proteins c-ret; Receptors, Tumor Necrosis Factor, Type I; Signal Transduction; Small Fiber Neuropathy; Tumor Necrosis Factor-alpha

Sensory defects associated with small-fiber neuropathy (SFN) can lead to profound disabilities. The relationship between the sensory nervous system and modulation of the renin-angiotensin system (RAS) has been described and focused on pain and neurodegeneration in several animal models. We have recently developed an experimental model of functional sensory neuropathy showing thermal hypoalgesia and neuropeptide depletion without nerve fiber degeneration. Here, we aimed to determine whether the modulation of angiotensin II (Ang II) activity could prevent sensory neuropathy induced by RTX. Control and RTX mice received ramipril, an Ang II converting enzyme (ACE) inhibitor, (0.5 mg/kg/day) or candesartan, an Ang II type 1 receptor (AT1R) blocker (0.5 mg/kg/day), one day before vehicle or RTX administration, and each day for the next seven days. Ramipril did not have a beneficial effect in RTX mice, whereas candesartan prevented thermal hypoalgesia and reduced neuropeptide depletion in intraepidermal nerve fibers and dorsal root ganglion neurons. The preventive effect of candesartan was not observed in mice deficient for the Ang II type 2 receptor (AT2R) and was counteracted in wild type mice by EMA200, an AT2R antagonist (3 mg/kg/day). Thus, candesartan may promote AT2R activation by blocking AT1R and increasing Ang II production and enhance its mechanisms of neuroprotection in our RTX model. Our finding that candesartan prevents nociception deficits and neuropeptide depletion encourages the evaluation of its therapeutic potential in patients presenting SFN, particularly those who experience chemotherapy-induced SFN.

Topics: Angiotensin II; Angiotensin II Type 1 Receptor Blockers; Angiotensin II Type 2 Receptor Blockers; Animals; Benzimidazoles; Biphenyl Compounds; Calcitonin Gene-Related Peptide; Diterpenes; Ganglia, Spinal; Imidazoles; Male; Mice; Mice, Knockout; Neuroprotective Agents; Pyridines; Ramipril; Receptor, Angiotensin, Type 2; Sensory Receptor Cells; Small Fiber Neuropathy; Substance P; Tetrazoles

Damage to the peripheral nervous system influences wound healing and, after a deep burn, imperfect cutaneous nerve regeneration occurs. A third-degree burn model was developed in rats combined with the use of resiniferatoxin (RTX), known to promote sensory neuropathy.. Rats were injected intraperitoneally either with RTX or vehicle. A mechanical sensory assay and the hot plate thermal sensory test were performed. The structural integrity of the sciatic nerve was assessed using transmission electron microcopy. After RTX injection, third-degree thermal burns were performed. Wound closure was monitored and samples were collected for histological analysis, immunohistochemistry and immunoblotting for neuronal markers.. RTX promoted both mechanical and thermal hypoalgesia. This transient RTX-mediated sensory deficit occurred without damaging the integrity of nerve fibers and induced a significant depletion of neuropeptides in both neuronal bodies and intraepidermal nerve fibers. Although wound closure rates were similar in both groups, the kinetic of granulation tissue remodeling was delayed in the RTX group compared with control group. A significant reduction of the peripherin expression in the RTX group was observed indicating impaired axonal regrowth of small fibers within the wound.. Our study confirms the important roles of innervation during skin healing and the defect of nerve regeneration after burn.

Topics: Animals; Burns; Diterpenes; Ganglia, Spinal; Granulation Tissue; Immunoblotting; Immunohistochemistry; Male; Microscopy, Electron, Transmission; Nerve Regeneration; Neuronal Outgrowth; Nociception; Rats; Rats, Sprague-Dawley; Sciatic Nerve; Small Fiber Neuropathy; Wound Healing