Administration Of Resiniferatoxin Research Articles

Thoracic Dorsal Root Ganglion Application of Resiniferatoxin Reduces Myocardial Ischemia-Induced Ventricular Arrhythmias.

A myocardial ischemia/reperfusion (IR) injury activates the transient receptor potential vanilloid 1 (TRPV1) dorsal root ganglion (DRG) neurons. The activation of TRPV1 DRG neurons triggers the spinal dorsal horn and the sympathetic preganglionic neurons in the spinal intermediolateral column, which results in sympathoexcitation. In this study, we hypothesize that the selective epidural administration of resiniferatoxin (RTX) to DRGs may provide cardioprotection against ventricular arrhythmias by inhibiting afferent neurotransmission during IR injury. Yorkshire pigs (n = 21) were assigned to either the sham, IR, or IR + RTX group. A laminectomy and sternotomy were performed on the anesthetized animals to expose the left T2-T4 spinal dorsal root and the heart for IR intervention, respectively. RTX (50 μg) was administered to the DRGs in the IR + RTX group. The activation recovery interval (ARI) was measured as a surrogate for the action potential duration (APD). Arrhythmia risk was investigated by assessing the dispersion of repolarization (DOR), a marker of arrhythmogenicity, and measuring the arrhythmia score and the number of non-sustained ventricular tachycardias (VTs). TRPV1 and calcitonin gene-related peptide (CGRP) expressions in DRGs and CGRP expression in the spinal cord were assessed using immunohistochemistry. The RTX mitigated IR-induced ARI shortening (-105 ms ± 13 ms in IR vs. -65 ms ± 11 ms in IR + RTX, p = 0.028) and DOR augmentation (7093 ms2 ± 701 ms2 in IR vs. 3788 ms2 ± 1161 ms2 in IR + RTX, p = 0.020). The arrhythmia score and VT episodes during an IR were decreased by RTX (arrhythmia score: 8.01 ± 1.44 in IR vs. 3.70 ± 0.81 in IR + RTX, p = 0.037. number of VT episodes: 12.00 ± 3.29 in IR vs. 0.57 ± 0.3 in IR + RTX, p = 0.002). The CGRP expression in the DRGs and spinal cord was decreased by RTX (DRGs: 6.8% ± 1.3% in IR vs. 0.6% ± 0.2% in IR + RTX, p < 0.001. Spinal cord: 12.0% ± 2.6% in IR vs. 4.5% ± 0.8% in IR + RTX, p = 0.047). The administration of RTX locally to thoracic DRGs reduces ventricular arrhythmia in a porcine model of IR, likely by inhibiting spinal afferent hyperactivity in the cardio-spinal sympathetic pathways.

Influence of TRPV1 on Thermal Nociception in Rats with Temporomandibular Joint Persistent Inflammation Evaluated by the Operant Orofacial Pain Assessment Device (OPAD).

Temporomandibular joint (TMJ)-associated inflammation contributes to the pain reported by patients with temporomandibular disorders (TMD). It is common for patients diagnosed with TMD to report pain in the masticatory muscles and temporomandibular joints, headache, and jaw movement disturbances. Although TMD can have different origins, including trauma and malocclusion disorder, anxiety/depression substantially impacts the development and maintenance of TMD. In general, rodent studies on orofacial pain mechanisms involve the use of tests originally developed for other body regions, which were adapted to the orofacial area. To overcome limitations and expand knowledge in orofacial pain, our group validated and characterized an operant assessment paradigm in rats with both hot and cold stimuli as well mechanical stimuli. Nevertheless, persistent inflammation of the TMJ has not been evaluated with this operant orofacial pain assessment device (OPAD). We characterized the thermal orofacial sensitivity for cold, neutral, and hot stimuli during the development of TMD using the OPAD behavior test. In addition, we evaluated the role of transient receptor potential vanilloid 1 (TRPV1) expressing nociceptors in rats with persistent TMJ inflammation. The experiments were performed in male and female rats with TMJ inflammation induced by carrageenan (CARR). Additionally, resiniferatoxin (RTX) was administered into the TMJs prior CARR to lesion TRPV1-expressing neurons to evaluate the role of TRPV1-expressing neurons. We evidenced an increase in the number of facial contacts and changes in the number of reward licks per stimulus on neutral (37°C) and cold (21°C) temperatures. However, at the hot temperature (42°C), the inflammation did not induce changes in the OPAD test. The prior administration of RTX in the TMJ prevented the allodynia and thermal hyperalgesia induced by CARR. We showed that TRPV-expressing neurons are involved in the sensitivity to carrageenan-induced pain in male and female rats evaluated in the OPAD.

Chronic ablation of TRPV1-sensitive skeletal muscle afferents attenuates the muscle metaboreflex.

Exercise intolerance is a hallmark symptom of cardiovascular disease and likely occurs via enhanced activation of muscle metaboreflex-induced vasoconstriction of the heart and active skeletal muscle which, thereby limits cardiac output and peripheral blood flow. Muscle metaboreflex vasoconstrictor responses occur via activation of metabolite-sensitive afferent fibers located in ischemic active skeletal muscle, some of which express transient receptor potential vanilloid 1 (TRPV1) cation channels. Local cardiac and intrathecal administration of an ultrapotent noncompetitive, dominant negative agonist resiniferatoxin (RTX) can ablate these TRPV1-sensitive afferents. This technique has been used to attenuate cardiac sympathetic afferents and nociceptive pain. We investigated whether intrathecal administration (L4-L6) of RTX (2 µg/kg) could chronically attenuate subsequent muscle metaboreflex responses elicited by reductions in hindlimb blood flow during mild exercise (3.2 km/h) in chronically instrumented conscious canines. RTX significantly attenuated metaboreflex-induced increases in mean arterial pressure (27 ± 5.0 mmHg vs. 6 ± 8.2 mmHg), cardiac output (1.40 ± 0.2 L/min vs. 0.28 ± 0.1 L/min), and stroke work (2.27 ± 0.2 L·mmHg vs. 1.01 ± 0.2 L·mmHg). Effects were maintained until 78 ± 14 days post-RTX at which point the efficacy of RTX injection was tested by intra-arterial administration of capsaicin (20 µg/kg). A significant reduction in the mean arterial pressure response (+45.7 ± 6.5 mmHg pre-RTX vs. +19.7 ± 3.1 mmHg post-RTX) was observed. We conclude that intrathecal administration of RTX can chronically attenuate the muscle metaboreflex and could potentially alleviate enhanced sympatho-activation observed in cardiovascular disease states.

Chronic Ablation of Hindlimb Afferents Containing TRPV1 Receptors Attenuates Cardiovascular Responses to Muscle Metaboreflex Activation

Increases in sympathetic nerve activity during exercise occur, in part, due to activation of the muscle metaboreflex – a pressor reflex triggered via stimulation of type III and IV skeletal muscle afferents. Some of these afferents express capsaicin sensitive transient receptor potential vanilloid 1 (TRPV1) channels. Epicardial and intrathecal administration of resiniferatoxin (RTX), an ultra‐potent analog of capsaicin, attenuates responses to cardiac sympathetic afferent activation and provides relief of limb pain due to bone cancer. We investigated whether administration of intrathecal RTX (2μg/kg, L4–L6) would attenuate the muscle metaboreflex (induced by partial reductions in hindlimb blood flow during steady‐state mild exercise; 3.2 km/h). Experiments were performed using conscious, chronically instrumented canines before and 10–42 days after RTX. RTX markedly attenuated metaboreflex‐induced increases in mean arterial pressure (+27.0 ± 5.0 mmHg vs. +5.9 ± 8.2 mmHg) and cardiac output (+1.40 ± 0.20 l/min vs. +0.28 ± 0.12 l/min). Terminal experiments under isoflurane anesthesia performed 78 ± 14 days after RTX showed attenuated pressor responses to hindlimb intra‐arterial infusion of 20μg/kg capsaicin (+45.7 ± 6.5 mmHg pre RTX vs. +19.7 ± 3.1 mmHg post RTX). We conclude that intrathecal RTX chronically attenuates muscle metaboreflex pressor responses. Thus, RTX could be an effective treatment to reduce excessive sympatho‐activation often seen during exercise in patients with heart failure or other diseases when the metaboreflex is overstimulated due to underperfusion of the active skeletal muscle.Support or Funding InformationHL‐55473, HL‐126706, HL‐120822 and GM‐058905

Preliminary results from a phase 1b double-blind study to assess the safety, tolerability and efficacy of intra-articular administration of resiniferatoxin or placebo for the treatment of moderate to severe pain due to osteoarthritis of the knee

Purpose: Osteoarthritis (OA) is a degenerative disease marked by breakdown of joint cartilage and associated bony changes. A dominant symptom is pain, which can become severe to the point where the patient is a candidate for joint replacement surgery. Although a wide range of analgesics and therapeutics are available, pain frequently remains an intractable problem. Even after total knee replacement, chronic knee pain has been reported in up to a third of patients. Resiniferatoxin (RTX) is a potent TRPV1 calcium channel agonist.

Resiniferatoxin reduces ventricular arrhythmias in heart failure via selectively blunting cardiac sympathetic afferent projection into spinal cord in rats

Excessive sympathetic activity is associated with heart failure and ventricular arrhythmias, which regulated by enhanced cardiac sympathetic afferent reflex, which can be blunted by resiniferatoxin, a selective receptor agonist of transient vanilloid potential 1 (TRPV1) + primary sensory afferents. The present study is aimed to determine whether intrathecal resiniferatoxin application affect cardiac sympathetic tone and electrophysiology, furtherly create a new effective strategy to prevent lethal arrhythmias in chronic heart failure. Four weeks after coronary artery occlusion to induce heart failure in rats, RTX (2μg/10 μl) or vehicle was injected intrathecally into the T2/T3 interspace. Cardiac sympathetic nerve activities (CSNA) and cardiac electrophysiology were evaluated two weeks later. Intrathecal resiniferatoxin significantly and selectively abolished the afferent markers expression (TRPV1 and calcitonin gene-related peptide) in dorsal horn and reduced overactivated CSNA. Electrophysiological studies revealed that resiniferatoxin administration intrathecally significantly reversed the prolongation of action potential duration (APD) and APD alternan, reduced the inducibilities of ventricular arrhythmias. Moreover, the over-activated calcium handling related protein CaMKII and RyR2 in heart failure was reversed by resiniferatoxin administration. In conclusion, these results firstly demonstrate that central chemo-ablation of the TRPV1+ afferents in spinal cord prevent heart from ventricular arrhythmias in heart failure via selectively blunting cardiac sympathetic afferent projection into spinal cord, which suggest a novel promising therapeutic method for anti-arrhythmia in heart failure.

Effects of early intravesical administration of resiniferatoxin to spinal cord-injured rats in neurogenic detrusor overactivity.

To investigate if intravesical administration during spinal shock of resiniferatoxin (RTX), an ultrapotent desensitizing agonist of transient receptor potential vanilloid-1 (TRPV1), would silence TRPV1-expressing bladder afferents at an early stage of disease progression and modulate neurogenic detrusor overactivity (NDO) emergence. Rats submitted to largely incomplete spinal cord transection at T8/9 spinal segment were treated with intravesical RTX (50 nM) or its vehicle during spinal shock. Four weeks after spinal lesion, bladder-reflex activity was evaluated by cystometry under urethane anesthesia, after which the bladder, spinal cord, and dorsal root ganglia were collected and processed. We found improvements on bladder function several weeks after early intravesical RTX administration, including a marked decrease of intravesical pressures and amplitude of bladder contractions. Such strong long-lasting urodynamic effects resulted from the very potent desensitizing activity of RTX on peripheral terminals of sensory afferents, an effect restricted to the bladder. Our results support that an early intervention with RTX could potentially attenuate NDO development and ensuing urinary incontinence, with a dramatic impact on the quality of life of spinal cord injury patients.

Cardiac Afferent Denervation Abolishes Ganglionated Plexi and Sympathetic Responses to Apnea: Implications for Atrial Fibrillation.

Background The autonomic nervous system response to apnea and its mechanistic connection to atrial fibrillation (AF) are unclear. We hypothesize that sensory neurons within the ganglionated plexi (GP) play a role. We aimed to delineate the autonomic response to apnea and to test the effects of ablation of cardiac sensory neurons with resiniferatoxin (RTX), a neurotoxic TRPV1 (transient receptor potential vanilloid 1) agonist. Methods Sixteen dogs were anesthetized and ventilated. Apnea was induced by stopping ventilation until oxygen saturations decreased to 80%. Nerve recordings from bilateral vagal nerves, left stellate ganglion, and anterior right GP were obtained before and during apnea, before and after RTX injection in the anterior right GP (protocol 1, n=7). Atrial effective refractory period and AF inducibility on single extrastimulation were assessed before and during apnea, and before and after intrapericardial RTX administration (protocol 2, n=9). GPs underwent immunohistochemical staining for TRPV1. Results Apnea increased anterior right GP activity, followed by clustered crescendo vagal bursts synchronized with heart rate and blood pressure oscillations. On further oxygen desaturation, a tonic increase in stellate ganglion activity and blood pressure ensued. Apnea-induced effective refractory period shortening from 110.20±31.3 ms to 90.6±29.1 ms ( P<0.001), and AF induction in 9/9 dogs versus 0/9 at baseline. After RTX administration, increases in GP and stellate ganglion activity and blood pressure during apnea were abolished, effective refractory period increased to 126.7±26.9 ms ( P=0.0001), and AF was not induced. Vagal bursts remained unchanged. GP cells showed cytoplasmic microvacuolization and apoptosis. Conclusions Apnea increases GP activity, followed by vagal bursts and tonic stellate ganglion firing. RTX decreases sympathetic and GP nerve activity, abolishes apnea's electrophysiological response, and AF inducibility. Sensory neurons play a role in apnea-induced AF.

A sensitive bioanalytical method for quantitative determination of resiniferatoxin in rat plasma using ultra-high performance liquid chromatography coupled to tandem mass spectrometry and its application in pharmacokinetic study

Resiniferatoxin (RTX) is a daphnane diterpene isolated from the latex of Euphorbia resinifera O. Berg, a potent activator of transient receptor potential vanilloid 1 (TrpV1), with a potency 103-105 times greater than pure capsaicin. Intravenous administration of RTX at very low concentration improves urodynamic parameters in patients with neurogenic detrusor overactivity and also reduces bladder pain in patients. Herein, a simple, rapid, selective and sensitive method for determination of RTX with silydianin as an internal standard was developed using ultra-high performance liquid chromatography coupled to tandem mass spectrometry with electrospray ionization source (UHPLC-ESI-MS/MS) in multiple reaction monitoring mode. The mass spectrometer was operated in positive electrospray ionization ((+) ESI) mode. Multiple reaction monitoring (MRM) mode was performed with ion pairs of m/z: 629.23→283.2 for RTX and 483.24→153.1 for IS. The limit of detection achieved in this method for RTX was 0.05 ng/mL and had good linearity in calibration range of 0.2–50 ng/mL ( r2 = 0.99). Precision and accuracy values were found to be < 15% (within acceptable limit), extraction recovery (≥ 88.2%), matrix effect (≥ 89.7%) and stability were in accordance with the bioanalytical guidelines. The sensitivity of this bio-analytical method supported the successful pharmacokinetic evaluation of RTX on rat plasma (2.5 μg/kg dose; i.v.) and has demonstrated pharmacokinetic parameters Vd and AUC0-∞ as 191.0 ± 71.31 mL/kg and 981.6 ± 137.40 min*ng/mL, respectively. The clearance was found to be 2.6 ± 0.38 mL/min/kg and half-life was 53.6 ± 23.51 min. This efficient, rapid and reliable method promises the quantification at low concentration of RTX, allowing determination of the pharmacokinetic profile, which is essential in future drug delivery and clinical application.

Focal selective chemo-ablation of spinal cardiac afferent nerve by resiniferatoxin protects the heart from pressure overload-induced hypertrophy

Resiniferatoxin (RTX), a selective transient receptor potential vanilloid 1 (TRPV1) receptor agonist, can eliminate TRPV1+ primary sensory afferents and blunt cardiac sympathetic afferent reflex for a relatively long period. The present study determined the effects of intrathecal RTX administration on transverse aortic constriction (TAC)-induced cardiac dysfunction and cardiac remodeling in rats. Five days before TAC, RTX (2 μg/10 μl) was injected intrathecally into the T2/T3 interspace of rats. Cardiac sympathetic nerve activities (CSNAs) and cardiac structure and function were determined eight weeks after TAC. Intrathecal RTX administration abolished TRPV1 expression in the dorsal horn and reduced over-activated CSNA in the TAC rat model. Hemodynamic analysis revealed that RTX reduced left ventricular end-diastolic pressure, indicating the improvement of cardiac compliance. Histologic analysis, real-time reverse transcription-polymerase chain reaction, and Western blots showed that RTX prevented TAC-induced cardiac hypertrophy, cardiac fibrosis, and cardiac apoptosis and reduced the expression of apoptotic proteins and myocardial mRNAs. In conclusion, these results demonstrate that focal chemo-ablation of TRPV1+ afferents in the spinal cord protects the heart from pressure overload-induced cardiac remodeling and cardiac dysfunction, which suggest a novel promising therapeutic method for cardiac hypertrophy and diastolic dysfunction.

Stimulating Frequency-dependent Analgesic Effect of Electroacupuncture via Down-regulating Expression of VEGF in Dorsal Horns of Lumbar Spinal Cord in Post Herpetic Neuralgia Rats

To observe the effect of different frequencies of electroacupuncture (EA) stimulation on pain threshold (PT) and expression of vascular endothelial growth factor (VEGF) in dorsal horns (DHs) of the lumbar spinal cord in resiniferatoxin (RTX)-induced post-herpetic neuralgia (PHN) rats, so as to reveal its mechanism in alleviating PHN. Male SD rats were randomized into control, model, 2 Hz-EA, 15 Hz-EA, 100 Hz-EA and sham EA groups (n=16 in each). The PHN model was induced by a single intraperitoneal injection of RTX (250 μg/kg), and rats of the control group received intraperitoneal injection of the same dose of vehicle (10% Tween 80, 10% alcohol and 0.9% NaCl). Rats of EA treatment groups received EA stimulation (2 Hz, 15 Hz or 100 Hz, 1 mA) at the left "Huantiao" (GB 30) and "Yanglingquan" (GB 34) for 30 min, once every other day for 35 days, starting from 1 week after RTX injection. For sham control, acupuncture needles were inserted ipsilaterally into GB 30 and GB 34 for 30 min without electrical stimulation or manual needle manipulation. The mechanical allodynia was quantified with Von Frey filaments. The expression of mRNA and protein of VEGF in the DHs of lumbar spinal cord 4-6 segments (sampled under light microscope) was detected by quantitative polymerase chain reaction (qPCR) and Western blot, respectively. A single RTX injection gradually induced tactile allodynia (significant reduction of the mechanical PT) within 3 weeks relevant to the control group (P<0.01). EA applied to GB 30 and GB 34 at 2 Hz and 15 Hz, but not 100 Hz, significantly decreased the tactile allodynia after the treatment (2 Hz from 2 weeks on and 15 Hz from 3 weeks on) in RTX-treated rats (P<0.05). RTX administration increased the mRNA and protein expression of VEGF in the lumbar spinal cord compared with the control group (P<0. 05). Moreover, 2 Hz, but not 15 Hz and 100 Hz EA significantly reduced VEGF mRNA and protein expression(P<0.05). The expression of both VEGF mRNA and protein was negatively correlated with mechanical PT in RTX-induced PHN rats. EA at 2 Hz can significantly reduce VEGF expression in the lumbar spinal cord DHs of PHN rats, which is possibly in part related to its effect in alleviating the mechanical allodynia. Our study suggests that 2 Hz EA is the best stimulation frequency for relieving PHN.

580 - Prevention of neurogenic detrusor overactivity following spinal cord injury: Effects of early administration of resiniferatoxin and botulinum toxin A

580 - Prevention of neurogenic detrusor overactivity following spinal cord injury: Effects of early administration of resiniferatoxin and botulinum toxin A

Supraspinal-selective TRPV1 desensitization induced by intracerebroventricular treatment with resiniferatoxin

The transient receptor potential vanilloid type 1 (TRPV1) is a thermosensitive cation channel that triggers heat pain in the periphery. Long-term desensitization of TRPV1, which can be induced by excess amounts of agonists, has been a method for investigating the physiological relevance of TRPV1-containing neuronal circuits, and desensitization induced by various routes of administration, including systemic, intrathecal and intraganglionic, has been demonstrated in rodents. In the present study, we examined the effect of intracerebroventricular (i.c.v.) treatment with an ultrapotent TRPV1 agonist, resiniferatoxin (RTX), on nociception and the analgesic effect of acetaminophen, which is known to mediate the activation of central TRPV1. I.c.v. administration of RTX a week before the test did not affect the licking/biting response to intraplantar injection of RTX (RTX test), suggesting that such i.c.v. treatment spares the function of TRPV1 at the hindpaw. Mice that had been i.c.v.-administered RTX also exhibited normal nociceptive responses in the formalin test and the tail pressure test, but acetaminophen failed to induce analgesia in those mice in any of the tests. These results suggest that i.c.v. administration of RTX leads to brain-selective TRPV1 desensitization in mice.

Pharmacological cardiac sympathetic afferent denervation in pacing‐induced heart failure

Chronic sympathoexcitation is a hallmark of congestive heart failure. Enhanced input from the cardiac sympathetic afferent reflex (CSAR) appears to be a major contributor to augmented central sympathetic outflow in heart failure (HF) in rats (Myocardial ischemia model. Wang et al., Hypertension, 2014) and dogs (Rapid ventricular pacing model. Wang and Zucker, Am. J. Physiol., 1996). Transient receptor potential vanilloid 1 (TRPV1) expressing cardiac afferent fibers are responsible for CSAR activation, since systemic (Zahner et al., J. Physiol., 2003) and epicardial (Wang et al., Hypertension, 2014) administration of resiniferatoxin (RTX), an ultra‐potent analog of capsaicin that degenerates TRPV1‐expressing neurons, can almost completely abolish CSAR activation in rats. We tested whether epicardial application of RTX degenerates cardiac sympathetic TRPV1‐expressing neurons therefore reducing CSAR reactivity, in canines with pacing‐induced HF. In control studies, stimulation of left ventricular afferents containing TRPV1 via topical application of capsaicin (2 mg/ml) caused a pressor response and tachycardia (Δ=+15 mmHg, +44bpm). These responses were abolished by acute topical application of RTX (50 μg/mL). Experiments were repeated 9 weeks following RTX treatment and the subsequent induction of HF via rapid ventricular pacing. RTX pretreatment abolished the responses to capsaicin, whereas CSAR was preserved in HF if no RTX pretreatment was performed. TRPV1 expressing neuronal degeneration was evaluated through immunofluorescence histology (IFH) in left ventricle tissue samples, with triple labeling for DAPI (nuclei marker), PGP9.5 (nerve terminal marker,) and TRPV1 receptors. RTX treated tissue samples showed considerably lower TRPV1 positive staining when compared to non‐treated samples. The present study validates RTX as a potential therapeutic method to block excessive CSAR activation in pacing‐induced heart failure dogs, therefore expanding pharmacotherapy options for heart failureSupport or Funding InformationNIH HL55473 and HL126706

(162) Pre-surgical intraplantar resiniferatoxin decreases post-operative pain in rats with plantar incision

Post-operative pain is a major cause of morbidity and suffering. Central sensitization during surgical intervention has been implicated in the development of chronic pain this may, in part, be due to inadequate peri-operative and post-operative analgesia. Neurons expressing TRPV1 (transient receptor potential cation channel subfamily V) have been implicated in nociception in a variety of pain models. The present study investigated therapeutic and molecular aspects of perioperative ablation of TRPV1-containing peripheral nerve terminals with intraplantar RTX (resiniferatoxin) in a surgical incision pain model. Furthermore, as RTX has algesic actions before institution of analgesia, the effect of lidocaine administered before RTX on post-operative hyperalgesia was investigated. Sprague-Dawley rats were administered RTX and/or lidocaine in the perioperative window prior to intraplantar incision in a 2x2 factorial design. Evoked pain to mechanical and thermal stimulation was assessed, as was spontaneous pain with measurement of guarding and home cage monitoring. RNA was extracted from spinal cord dorsal horn on post-operative day 2 for transcriptomic analysis. Plantar incision caused thermal hyperalgesia, but intraplantar RTX attenuated responses to noxious infrared diode laser thermal A-delta or C-fiber stimulation during the 4 days of the study (p<0.05). Mechanical allodynia, measured by Von Frey stimulation, was pronounced over the first 2 post-operative days, and was also attenuated by pre-emptive RTX. Guarding of the wound was most pronounced at 4 hours and 24 hours pre-emptive RTX attenuated this measure of spontaneous pain (assessed by guarding score, p<0.05). Mechanical pain sensation, measured by pinprick, was preserved after RTX treatment, as was motoric function (measured by time spent on rotarod). Lidocaine pre-treatment had no effect on any of the behavioral assays. Transcriptomic analysis for differential gene expression was also undertaken. Peri-operative administration of RTX, delivered in a clinically relevant manner, was efficacious in decreasing post-operative pain.

Intrathecal Resiniferatoxin Modulates TRPV1 in DRG Neurons and Reduces TNF-Induced Pain-Related Behavior.

Transient receptor potential vanilloid-1 (TRPV1) is a nonselective cation channel, predominantly expressed in sensory neurons. TRPV1 is known to play an important role in the pathogenesis of inflammatory and neuropathic pain states. Previous studies suggest interactions between tumor necrosis factor- (TNF-) alpha and TRPV1, resulting in a modulation of ion channel function and protein expression in sensory neurons. We examined the effect of intrathecal administration of the ultrapotent TRPV1 agonist resiniferatoxin (RTX) on TNF-induced pain-associated behavior of rats using von Frey and hot plate behavioral testing. Intrathecal injection of TNF induces mechanical allodynia (2 and 20 ng/kg) and thermal hyperalgesia (200 ng) 24 h after administration. The additional intrathecal administration of RTX (1.9 μg/kg) alleviates TNF-induced mechanical allodynia and thermal hyperalgesia 24 h after injection. In addition, TNF increases the TRPV1 protein level and number of TRPV1-expressing neurons. Both effects could be abolished by the administration of RTX. These results suggest that the involvement of TRPV1 in TNF-induced pain offers new TRPV1-based experimental therapeutic approaches and demonstrates the analgesic potential of RTX in inflammatory pain diseases.

Intraluminal Administration of Resiniferatoxin Protects against Clostridium difficile Toxin A-Induced Colitis.

Clostridium difficile toxin A is a colonic inflammatory agent that acts partially by activation of TRPV1 (transient receptor potential vanilloid type 1). Resiniferatoxin (RTX) is an excitotoxin that activates TRPV1 at low concentrations and defunctionalizes TRPV1 at high concentrations. RTX at various doses was injected intraluminally into isolated ileal segments in anesthetized rats. After 3 hours, the treated segments were removed and inflammation was assessed. This acute treatment with RTX resulted in biphasic responses: (1) an increase in inflammation similar to that caused by toxin A and capsaicin at low doses of up to 100 ng RTX and (2) no inflammatory effect of RTX at higher doses (1–100 μg), consistent with a defunctionalizing or neurotoxic effect of RTX at high doses. Separately, anesthetized rats were treated with RTX enemas and one or four weeks later were challenged with toxin A. Toxin A-induced colitis was significantly inhibited one week after an RTX enema, and this effect was RTX dose dependent. When tested four weeks after administration of the RTX enema, protection against toxin A colitis was lost. In conclusion, an RTX enema protects against toxin A-induced colitis in rats for at least one week but less than four weeks.

The effect of intra-articular vanilloid receptor agonists on pain behavior measures in a murine model of acute monoarthritis.

Arthritis is the most common cause of disability in the US, and the primary manifestation of arthritis is joint pain that leads to progressive physical limitation, disability, morbidity, and increased health care utilization. Capsaicin (CAP) is a vanilloid agonist that causes substance P depletion by interacting with vanilloid receptor transient receptor potential V1 on small unmyelinated C fibers. It has been used topically for analgesia in osteoarthritis with variable success. Resiniferatoxin (RTX) is an ultra potent CAP analog. The aim of this study was to measure the analgesic effects of intra-articular (IA) administration of CAP and RTX in experimental acute inflammatory arthritis in mice. Evoked pain score (EPS) and a dynamic weight bearing (DWB) device were used to measure nociceptive behaviors in a murine model of acute inflammatory monoarthritis. A total of 56 C57B16 male mice underwent EPS and DWB testing – 24 nonarthritic controls and 32 mice with carrageenan-induced arthritis. The effects of pretreatment with 0.1% CAP, 0.0003% RTX, or 0.001% RTX were measured. Nociception was reproducibly demonstrated by increased EPS and reduced DWB measures in the affected limb of arthritic mice. Pretreatment with 0.001% RTX resulted in statistically significant improvement in EPS and DWB measures when compared with those observed in carrageenan-induced arthritis animals. Pretreatment with IA 0.0003% RTX and IA 0.01% CAP resulted in improvement in some but not all of these measures. The remaining 24 mice underwent evaluation following treatment with 0.1% CAP, 0.0003% RTX, or 0.001% RTX, and the results obtained were similar to that of naïve, nonarthritic mice.

TRPV1 is crucial for proinflammatory STAT3 signaling and thermoregulation-associated pathways in the brain during inflammation.

Transient receptor potential vanilloid receptor 1 (TRPV1) is a non-selective cation channel that is stimulated by heat (>43 °C), mechanical/osmotic stimuli, and low pH. The importance of TRPV1 in inflammatory responses has been demonstrated, whereas its participation in brains remains unclear. In the present study, the intracerebroventricular (icv) administration of the TRPV1 agonist resiniferatoxin (RTX) induced the activation of signal transducer and activator of transcription 3 (STAT3) in circumventricular organs (CVOs) and thermoregulation-associated brain regions with a similar patttern to the peripheral and icv administration of lipopolysaccharide (LPS). With the peripheral and icv LPS stimuli, STAT3 activation was significantly lower in Trpv1−/− mice than in Trpv1+/+ mice. The icv administration of RTX induced transient hypothermia, whereas that of the TRPV1 antagonist capsazepine enhanced the magnitude and period of LPS-induced hyperthermia. These results indicate that TRPV1 is important for activating proinflammatory STAT3 signaling and thermoregulation-associated brain pathways in the brain.

Changes in Spinal Cord Following Inflammatory and Neuropathic Pain and the Effectiveness of Resiniferatoxin

Peripheral inflammation or nerve injury results in changes in the spinal cord, initiating a process of central sensitization. Although nociceptive Transient Receptor Potential (TRP) channels have been studied extensively, the role of these channels expressed at the central terminals in the spinal cord is not fully understood. Here, we studied the expression and function of TRPV1 channels at the spinal cord following induction of inflammatory pain by Complete Freund's Adjuvant (CFA) and neuropathic pain by Chronic Constriction Injury (CCI). Rats treated with CFA or subjected to CCI developed long-term thermal and mechanical hypersensitivity. Peripheral inflammation or injury induced an inflammatory response at the levels of spinal cord, which included activation of glia and increased levels of proinflammatory mediators. As a result, expression of TRPV1 was significantly increased and the associated function of TRPV1-mediated CGRP release was also significantly increased. Single intrathecal administration of resiniferatoxin (RTX), an ultrapotent TRPV1 agonist, selectively reversed inflammatory thermal hypersensitivity and the associated changes in TRPV1 expression and function without affecting mechanical hypersensitivity. In summary, peripheral nerve activity triggers an inflammatory response at the spinal dorsal horn, which results in enhanced expression and function of TRPV1 channels. Targeting TRPV1 expressed in the central terminals is a viable strategy to alleviate certain modalities of pain.