Intradermal Injection Of Capsaicin Research Articles

A data science approach to the selection of most informative readouts of the human intradermal capsaicin pain model to assess pregabalin effects.

Persistent and, in particular, neuropathic pain is a major healthcare problem with still insufficient pharmacological treatment options. This triggered research activities aimed at finding analgesics with a novel mechanism of action. Results of these efforts will need to pass through the phases of drug development, in which experimental human pain models are established components e.g. implemented as chemical hyperalgesia induced by capsaicin. We aimed at ranking the various readouts of a human capsaicin-based pain model with respect to the most relevant information about the effects of a potential reference analgesic. In a placebo-controlled, randomized cross-over study, seven different pain-related readouts were acquired in 16 healthy individuals before and after oral administration of 300mg pregabalin. The sizes of the effect on pain induced by intradermal injection of capsaicin were quantified by calculating Cohen's d. While in four of the seven pain-related parameters, pregabalin provided a small effect judged by values of Cohen's d exceeding 0.2, an item categorization technique implemented as computed ABC analysis identified the pain intensities in the area of secondary hyperalgesia and of allodynia as the most suitable parameters to quantify the analgesic effects of pregabalin. Results of this study provide further support for the ability of the intradermal capsaicin pain model to show analgesic effects of pregabalin. Results can serve as a basis for the designs of studies where the inclusion of this particular pain model and pregabalin is planned.

Phoneutria toxin PnTx3-5 inhibits TRPV1 channel with antinociceptive action in an orofacial pain model

Capsaicin, an agonist of TRPV1, evokes intracellular [Ca2+] transients and glutamate release from perfused trigeminal ganglion. The spider toxin PnTx3-5, native or recombinant is more potent than the selective TRPV1 blocker SB-366791 with IC50 of 47 ± 0.18 nM, 45 ± 1.18 nM and 390 ± 5.1 nM in the same experimental conditions. PnTx3-5 is thus more potent than the selective TRPV1 blocker SB-366791. PnTx3-5 (40 nM) and SB-366791 (3 μM) also inhibited the capsaicin-induced increase in intracellular Ca2+ in HEK293 cells transfected with TRPV1 by 75 ± 16% and 84 ± 3.2%, respectively. In HEK293 cells transfected with TRPA1, cinnamaldehyde (30 μM) generated an increase in intracellular Ca2+ that was blocked by the TRPA1 antagonist HC-030031 (10 μM, 89% inhibition), but not by PnTx3-5 (40 nM), indicating selectivity of the toxin for TRPV1. In whole-cell patch-clamp experiments on HEK293 cells transfected with TRPV1, capsaicin (10 μM) generated inward currents that were blocked by SB-366791 and by both native and recombinant PnTx3-5 by 47 ± 1.4%; 54 ± 7.8% and 56 ± 9.0%, respectively. Intradermal injection of capsaicin into the rat left vibrissa induced nociceptive behavior that was blocked by pre-injection with either SB-366791 (3 nmol/site i.d., 83.3 ± 7.2% inhibition) or PnTx3-5 (100 fmol/site, 89 ± 8.4% inhibition). We conclude that both native and recombinant PnTx3-5 are potent TRPV1 receptor antagonists with antinociceptive action on pain behavior evoked by capsaicin.

Minocycline Inhibits the Enhanced Antidromic Stimulation-induced Sensitization of C-Fibers Following Intradermal Capsaicin Injection

Background: Our previous studies indicated that retrograde signaling initiating from the spinal cord was mediated by the plasticity of Dorsal Root Ganglion (DRG) neurons. Both retrograde signaling and neuronal plasticity contributed to neurogenic inflammation, which were modulated by the activity of Satellite Glial Cells (SGCs). Thus, we want to know whether retrograde signaling is involved in the hypersensitivity of nociceptive afferents, and whether this process is affected by the plasticity of DRG neurons and glia. Objective: The study aims to examine if retrograde signaling can induce hypersensitivity of primary afferent nociceptors and if hypersensitivity involves glial modulation. Methods: Antidromic Electrical Stimulation (ES) of dorsal roots was used to mimic retrograde signaling activity. C- primary nociceptive afferent activity was recorded for testing the effect of antidromic ES. In a separate group, intradermal capsaicin injection to the ipsilateral hindpaw was used to prime DRG nociceptive neurons. For the third group, a glial inhibitor, minocycline, was pre-administered to test glial modulation in this process. Results: Antidromic ES sensitized the responses of C-fibers to nociceptive mechanical stimuli. For rats subjected to intradermal capsaicin injection, C fibers experienced more drastic sensitization induced by antidromic ES, shown as a greater response and longer duration, implying that sensitization correlates with the activation of DRG neurons. Minocycline pretreatment significantly blocked the priming effect of capsaicin on C-fiber sensitization induced by antidromic ES, indicating the involvement of SGCs in DRG neuronal sensitization. Conclusion: Retrograde signaling may be one of the important mechanisms in neurogenic inflammation-mediated nociception, and this process is subjected to satellite glial modulation.

Descending inhibition selectively counteracts the capsaicin-induced facilitation of dorsal horn neurons activated by joint nociceptive afferents.

Previous studies from our laboratory showed that in the anesthetized cat, the intradermal injection of capsaicin in the hindpaw facilitated the intraspinal field potentials (IFPs) evoked by stimulation of the intermediate and high-threshold myelinated fibers in the posterior articular nerve (PAN). The capsaicin-induced facilitation was significantly reduced 3-4h after the injection, despite the persistence of hindpaw inflammation. Although this effect was attributed to an incremented descending inhibition acting on the spinal pathways, it was not clear if it was set in operation once the capsaicin-induced effects exceeded a certain threshold, or if it was continuously operating to keep the increased neuronal activation within manageable limits. To evaluate the changes in descending inhibition, we now examined the effects of successive reversible spinal blocks on the amplitude of the PAN IFPs evoked at different times after the intradermal injection of capsaicin. We found that after capsaicin the PAN IFPs recorded in laminae III-V by activation of high-threshold nociceptive Aδ myelinated fibers increased gradually during successive reversible spinal blocks, while the IFPs evoked by intermediate and low threshold proprioceptive Aβ afferents were only slightly affected. It is concluded that during the development of the central sensitization produced by capsaicin, there is a gradual increase of descending inhibition that tends to limit the nociceptive-induced facilitation, mainly by acting on the neuronal populations receiving inputs from the capsaicin-activated afferents without significantly affecting the information on joint angle transmitted by the low threshold afferents.

Supraspinal modulation of neuronal synchronization by nociceptive stimulation induces an enduring reorganization of dorsal horn neuronal connectivity

The state of central sensitization induced by the intradermic injection of capsaicin leads to structured (non-random) changes in functional connectivity between dorsal horn neuronal populations distributed along the spinal lumbar segments in anaesthetized cats. The capsaicin-induced changes in neuronal connectivity and the concurrent increase in secondary hyperalgesia are transiently reversed by the systemic administration of small doses of lidocaine, a clinically effective procedure to treat neuropathic pain. The effects of both capsaicin and lidocaine are greatly attenuated in spinalized preparations, showing that supraspinal influences play a significant role in the shaping of nociceptive-induced changes in dorsal horn functional neuronal connectivity. We conclude that changes in functional connectivity between segmental populations of dorsal horn neurones induced by capsaicin and lidocaine result from a cooperative adaptive interaction between supraspinal and spinal neuronal networks, a process that may have a relevant role in the pathogenesis of chronic pain and analgesia. Despite a profusion of information on the molecular and cellular mechanisms involved in the central sensitization produced by intense nociceptive stimulation, the changes in the patterns of functional connectivity between spinal neurones associated with the development of secondary hyperalgesia and allodynia remain largely unknown. Here we show that the state of central sensitization produced by the intradermal injection of capsaicin is associated with structured transformations in neuronal synchronization that lead to an enduring reorganization of the functional connectivity within a segmentally distributed ensemble of dorsal horn neurones. These changes are transiently reversed by the systemic administration of small doses of lidocaine, a clinically effective procedure to treat neuropathic pain. Lidocaine also reduces the capsaicin-induced facilitation of the spinal responses evoked by weak mechanical stimulation of the skin in the region of secondary but not primary hyperalgesia. The effects of both intradermic capsaicin and systemic lidocaine on the segmental correlation and coherence between ongoing cord dorsum potentials and on the responses evoked by tactile stimulation in the region of secondary hyperalgesia are greatly attenuated in spinalized preparations, showing that supraspinal influences are involved in the reorganization of the nociceptive-induced structured patterns of dorsal horn neuronal connectivity. We conclude that the structured reorganization of the functional connectivity between the dorsal horn neurones induced by capsaicin nociceptive stimulation results from cooperative interactions between supraspinal and spinal networks, a process that may have a relevant role in the shaping of the spinal state in the pathogenesis of chronic pain and analgesia.

(129) - Pain input after spinal cord injury (SCI) expands the region of secondary injury

Due to their traumatic etiology, spinal cord injury patients present with injuries to other parts of the body. These associated injuries can be a source of pain input through the damaged spinal cord. Animal research has shown that activating nociceptive fibers soon after injury induces maladaptive plasticity (Grau et al., 2004). In those studies, male rats received six minutes of variable electrical stimulation to the tail at an intensity that engaged C-fibers. Acutely, shocked animals show increases in pro-inflammatory cytokines and cell death (Turtle et at., 2017). Chronically, a single exposure caused the lesion to expand, locomotor deficits, and increased mortality (Grau et al., 2004). Recent work suggests that the loss of tissue at the site of injury is related to a breakdown in the blood spinal cord barrier (BSCB) and infiltration of blood (hemorrhage) at the site of injury. The present study explored the generality of these effects by examining whether chemically activating unmyelinated C fibers, by applying the irritant capsaicin to one hind paw, would have the same effect. Male rats underwent a contusion injury at T12. Twenty-four hours later, subjects received an intradermal injection of capsaicin (3%) or vehicle to one hind paw. Tissue was collected at three and 24 hours after injection and assessed for hemorrhage. Capsaicin treatment induced an acute disruption in locomotor performance. Cellular assays showed that pain input increased the infiltration of blood into the area of injury. We next examined whether these effects generalize to female rats. Here too capsaicin induced an acute disruption in locomotor performance and increased hemorrhage. Further, capsaicin treatment disrupted long-term locomotor recovery in male rats. These findings extend the generality of our results and suggest that pain input (polytrauma) after SCI impairs recovery because it expands the region of secondary injury through a breakdown of the BSCB.

Differential effect of Incobotulinumtoxin A on pain, neurogenic flare and hyperalgesia in human surrogate models of neurogenic pain.

The effectiveness of Botulinum-neurotoxin A (BoNT/A) to treat pain in human pain models is very divergent. This study was conducted to clarify if the pain models or the route of BoNT/A application might be responsible for these divergent findings. Sixteen healthy subjects (8 males, mean age 27±5years) were included in a first set of experiments consisting of three visits: (1) Visit: Quantitative sensory testing (QST) was performed before and after intradermal capsaicin injection (CAPS, 15μg) on one thigh and electrical current stimulation (ES, 1Hz) on the contralateral thigh. During stimulation pain and the neurogenic flare response (laser-Doppler imaging) were assessed. (2) Four weeks later, BoNT/A (Xeomin® , 25MU) was injected intracutaneously on both sides. (3) Seven days later, the area of BoNT/A application was determined by the iodine-starch staining and the procedure of the (1) visit was exactly repeated. In consequence of these results, 8 healthy subjects (4 males, mean age 26±3years) were included into a second set of experiments. The experimental setting was exactly the same with the exception that stimulation frequency of ES was increased to 4Hz and BoNT/A was injected subcutaneously into the thigh, which was stimulated by capsaicin. BoNT/A reduced the 1Hz ES flare size (p<0.001) and pain ratings (p<0.01), but had no effect on 4Hz ES and capsaicin-induced pain, hyperalgesia, or flare size, regardless of the depth of BoNT/A injection (i.c./s.c). Moreover, i.c. BoNT/A injection significantly increased warm detection and heat pain thresholds in naive skin (WDT, Δ 2.2°C, p<0.001; HPT Δ 1.8°C, p<0.005). BoNT/A has a moderate inhibitory effect on peptidergic and thermal C-fibers in healthy human skin. The study demonstrates that BoNT/A (Incobotulinumtoxin A) has differential effects in human pain models: It reduces the neurogenic flare and had a moderate analgesic effects in low frequency but not high frequency current stimulation of cutaneous afferent fibers at C-fiber strength; BoNT/A had no effect in capsaicin-induced (CAPS) neurogenic flare or pain, or on hyperalgesia to mechanical or heat stimuli in both pain models. Intracutaneous BoNT/A increases warm and heat pain thresholds on naïve skin.

Effect of Intravenous Ethanol on Capsaicin-Induced Hyperalgesia in Human Subjects.

The objective of this study was to assess ethanol's (EtOH's) effects on capsaicin-induced hyperalgesia in healthy participants. Specifically, we investigated the change in area of capsaicin-induced hyperalgesia following 3 interventions: intravenous EtOH at 2 targeted breath alcohol concentrations (BrAC), or placebo. Eighteen participants participated in 3 test days in a randomized order. Each test day, participants received an intradermal capsaicin injection on the volar surface of the forearm, followed by either infusion of high concentration EtOH (targeted BrAC=0.100g/dl), low concentration EtOH (targeted BrAC=0.040g/dl), or placebo. The area of hyperalgesia was determined by von Frey technique at 2 time points, prior to EtOH infusion, and again when target BrAC was reached. The primary outcome was the percent change in the area of capsaicin-induced hyperalgesia. Additional outcome measures included the visual analogue scale of mood states (VAS), which was administered at each time point. There was a marked 30% reduction in the area of capsaicin-induced hyperalgesia with infusion of a high concentration of EtOH (p<0.05). Low concentration EtOH produced a 10% reduction in hyperalgesia area, although this finding did not reach significance. Further, participants reported significant feelings of euphoria and drowsiness at high concentrations of EtOH (p<0.05), as measured by the VAS. In a human model examining pain phenomena related to central sensitization, this study is the first to demonstrate that capsaicin-induced hyperalgesia is markedly attenuated by EtOH. The capsaicin experimental pain paradigm employed provides a novel approach to evaluate EtOH's effects on pain processing. The antihyperalgesic effects of EtOH observed have important clinical implications for the converging fields of substance abuse and pain medicine and may inform why patients with chronic pain often report alcohol use as a form of self-medication.

Protein Kinase C γ Interneurons Mediate C-fiber-induced Orofacial Secondary Static Mechanical Allodynia, but Not C-fiber-induced Nociceptive Behavior.

Tissue injury enhances pain sensitivity both at the site of tissue damage and in surrounding uninjured skin (secondary hyperalgesia). Secondary hyperalgesia encompasses several pain symptoms including pain to innocuous punctate stimuli or static mechanical allodynia. How injury-induced barrage from C-fiber nociceptors produces secondary static mechanical allodynia has not been elucidated. Combining behavioral, immunohistochemical, and Western blot analysis, the authors investigated the cell and molecular mechanisms underlying the secondary static mechanical allodynia in the rat medullary dorsal horn (MDH) using the capsaicin model (n = 4 to 5 per group). Intradermal injection of capsaicin (25 μg) into the vibrissa pad produces a spontaneous pain and a secondary static mechanical allodynia. This allodynia is associated with the activation of a neuronal network encompassing lamina I-outer lamina III, including interneurons expressing the γ isoform of protein kinase C (PKCγ) within inner lamina II (IIi) of MDH. PKCγ is concomitantly phosphorylated (+351.4 ± 79.2%, mean ± SD; P = 0.0003). Mechanical allodynia and innocuous punctate stimulus-evoked laminae I to III neuronal activation can be replicated after intracisternally applied γ-aminobutyric acid receptor type A (GABAA) antagonist (bicuculline: 0.05 μg) or reactive oxygen species (ROS) donor (tert-butyl hydroperoxide: 50 to 250 ng). Conversely, intracisternal PKCγ antagonist, GABAA receptor agonist, or ROS scavenger prevent capsaicin-induced static mechanical allodynia and neuronal activation. Sensitization of lamina IIi PKCγ interneurons is required for the manifestation of secondary static mechanical allodynia but not for spontaneous pain. Such sensitization is driven by ROS and GABAAergic disinhibition. ROS released during intense C-fiber nociceptor activation might produce a GABAAergic disinhibition of PKCγ interneurons. Innocuous punctate inputs carried by Aδ low-threshold mechanoreceptors onto PKCγ interneurons can then gain access to the pain transmission circuitry of superficial MDH, producing pain.

Impact of Genetic Reduction of NMNAT2 on Chemotherapy-Induced Losses in Cell Viability In Vitro and Peripheral Neuropathy In Vivo.

Nicotinamide mononucleotide adenylyl transferases (NMNATs) are essential neuronal maintenance factors postulated to preserve neuronal function and protect against axonal degeneration in various neurodegenerative disease states. We used in vitro and in vivo approaches to assess the impact of NMNAT2 reduction on cellular and physiological functions induced by treatment with a vinca alkaloid (vincristine) and a taxane-based (paclitaxel) chemotherapeutic agent. NMNAT2 null (NMNAT2-/-) mutant mice die at birth and cannot be used to probe functions of NMNAT2 in adult animals. Nonetheless, primary cortical cultures derived from NMNAT2-/- embryos showed reduced cell viability in response to either vincristine or paclitaxel treatment whereas those derived from NMNAT2 heterozygous (NMNAT2+/-) mice were preferentially sensitive to vincristine-induced degeneration. Adult NMNAT2+/- mice, which survive to adulthood, exhibited a 50% reduction of NMNAT2 protein levels in dorsal root ganglia relative to wildtype (WT) mice with no change in levels of other NMNAT isoforms (NMNAT1 or NMNAT3), NMNAT enzyme activity (i.e. NAD/NADH levels) or microtubule associated protein-2 (MAP2) or neurofilament protein levels. We therefore compared the impact of NMNAT2 knockdown on the development and maintenance of chemotherapy-induced peripheral neuropathy induced by vincristine and paclitaxel treatment using NMNAT2+/- and WT mice. NMNAT2+/- did not differ from WT mice in either the development or maintenance of either mechanical or cold allodynia induced by either vincristine or paclitaxel treatment. Intradermal injection of capsaicin, the pungent ingredient in hot chili peppers, produced equivalent hypersensitivity in NMNAT2+/- and WT mice receiving vehicle in lieu of paclitaxel. Capsaicin-evoked hypersensitivity was enhanced by prior paclitaxel treatment but did not differ in either NMNAT2+/- or WT mice. Thus, capsaicin failed to unmask differences in nociceptive behaviors in either paclitaxel-treated or paclitaxel-untreated NMNAT2+/- and WT mice. Moreover, no differences in motor behavior were detected between genotypes in the rotarod test. Our studies do not preclude the possibility that complete knockout of NMNAT2 in a conditional knockout animal could unmask a role for NMNAT2 in protection against detrimental effects of chemotherapeutic treatment.

Characterizing pinprick-evoked brain potentials before and after experimentally induced secondary hyperalgesia.

Secondary hyperalgesia is believed to be a key feature of "central sensitization" and is characterized by enhanced pain to mechanical nociceptive stimuli. The aim of the present study was to characterize, using EEG, the effects of pinprick stimulation intensity on the magnitude of pinprick-elicited brain potentials [event-related potentials (ERPs)] before and after secondary hyperalgesia induced by intradermal capsaicin in humans. Pinprick-elicited ERPs and pinprick-evoked pain ratings were recorded in 19 healthy volunteers, with mechanical pinprick stimuli of varying intensities (0.25-mm probe applied with a force extending between 16 and 512 mN). The recordings were performed before (T0) and 30 min after (T1) intradermal capsaicin injection. The contralateral noninjected arm served as control. ERPs elicited by stimulation of untreated skin were characterized by 1) an early-latency negative-positive complex peaking between 120 and 250 ms after stimulus onset (N120-P240) and maximal at the vertex and 2) a long-lasting positive wave peaking 400-600 ms after stimulus onset and maximal more posterior (P500), which was correlated to perceived pinprick pain. After capsaicin injection, pinprick stimuli were perceived as more intense in the area of secondary hyperalgesia and this effect was stronger for lower compared with higher stimulus intensities. In addition, there was an enhancement of the P500 elicited by stimuli of intermediate intensity, which was significant for 64 mN. The other components of the ERPs were unaffected by capsaicin. Our results suggest that the increase in P500 magnitude after capsaicin is mediated by facilitated mechanical nociceptive pathways.

Minocycline inhibits neurogenic inflammation by blocking the effects of tumor necrosis factor-α.

It has been well established that neurogenic inflammation is one of the major pathological processes underlying inflammatory pain, but there are few effective anti-inflammatory drugs to alleviate such pain. The present study shows that minocycline, a widely used glial activation inhibitor, is effective in reducing neurogenic inflammation. Patch-clamp recordings showed that small sized dorsal root ganglion (DRG) neurons were dramatically excited following intradermal capsaicin injection in the rat hind paw, evidenced by decreased rheobase and membrane threshold. Pretreatment with minocycline (30mg/kg for 1day, intraperitoneal injection) blocked the increased neuronal excitability. Western blot and immunostaining of DRG revealed the activation of satellite glial cells (SGCs) following capsaicin injection. The up-regulation of glial fibrillary acidic protein (GFAP) was significantly inhibited by minocycline pre-administration. Measurement of tumor necrosis factor α (TNF-α) and its receptor, TNF-α receptor 1 (TNFR1), showed that minocycline mainly blocked the up-regulation of TNF-α in SGCs and TNFR1s in neurons following capsaicin injection. The pivotal role of TNF-α in neurogenic inflammation was further supported by the findings that incubation DRG with TNF-α mimicked the increased excitability of DRG neurons induced by capsaicin injection, and that TNF-α application enhanced cutaneous vasodilation in the hind paws induced by antidromic electrical stimulation of dorsal roots. Based on these results, we propose that minocycline is a potential therapeutic drug that can reduce neuronal excitability and neurogenic inflammation by working on SGCs to inhibit the expression of TNF-α.

Effects of Intradermal Capsaicin Injections in Women With Primary and Secondary Vestibulodynia [42

Effects of Intradermal Capsaicin Injections in Women With Primary and Secondary Vestibulodynia [42

A randomized, double-blind, crossover study to evaluate the depth response relationship of intradermal capsaicin-induced pain and hyperalgesia in healthy adult volunteers.

The purpose of this study was to evaluate pain and hyperalgesia in response to different depths of intradermal (ID) capsaicin injections in healthy volunteers. Double-blind, cross-over study. Clinical Research Laboratory. Fifteen healthy male subjects received ID capsaicin injections into the volar aspect of each forearm at depths of 1 mm, 3 mm, 5 mm, and 7 mm. After injection, spontaneous pain, elicited pain, flare response, heat thresholds, and area of hyperalgesia were measured at various time points. Spontaneous pain, elicited pain (pinprick, stroking, and hot pain), hyperalgesia area, and allodynia area. No significant difference was found between any depths in spontaneous pain, elicited pain (pinprick, stroking, hot pain), hyperalgesia area, or allodynia area. A significant difference was found in the change in heat threshold between 5 mm and 1 mm, 7 mm and 1 mm, 5 mm and 3 mm, 7 mm and 3 mm depths. A significant difference was found in flare area between 5 mm and 3 mm depths. A significant difference was found in systolic blood pressure area under the curve (AUC) between 7 mm and 1 mm depths, and for both systolic and diastolic pressures for 5 mm and 1 mm depths, and 5 mm and 3 mm depths. A significant difference was found in pulse AUC between 5 mm and 1 mm depths and 5 mm and 3 mm depths. Injection of capsaicin at different depths in the skin had different effects on heart rate and blood pressure but no effect on pain. These results may have implications on the pharmacology and analgesic predictive value of the model of ID capsaicin.

Identifying the integrated neural networks involved in capsaicin-induced pain using fMRI in awake TRPV1 knockout and wild-type rats.

In the present study, we used functional MRI in awake rats to investigate the pain response that accompanies intradermal injection of capsaicin into the hindpaw. To this end, we used BOLD imaging together with a 3D segmented, annotated rat atlas and computational analysis to identify the integrated neural circuits involved in capsaicin-induced pain. The specificity of the pain response to capsaicin was tested in a transgenic model that contains a biallelic deletion of the gene encoding for the transient receptor potential cation channel subfamily V member 1 (TRPV1). Capsaicin is an exogenous ligand for the TRPV1 receptor, and in wild-type rats, activated the putative pain neural circuit. In addition, capsaicin-treated wild-type rats exhibited activation in brain regions comprising the Papez circuit and habenular system, systems that play important roles in the integration of emotional information, and learning and memory of aversive information, respectively. As expected, capsaicin administration to TRPV1-KO rats failed to elicit the robust BOLD activation pattern observed in wild-type controls. However, the intradermal injection of formalin elicited a significant activation of the putative pain pathway as represented by such areas as the anterior cingulate, somatosensory cortex, parabrachial nucleus, and periaqueductal gray. Notably, comparison of neural responses to capsaicin in wild-type vs. knock-out rats uncovered evidence that capsaicin may function in an antinociceptive capacity independent of TRPV1 signaling. Our data suggest that neuroimaging of pain in awake, conscious animals has the potential to inform the neurobiological basis of full and integrated perceptions of pain.

Intradermal glutamate and capsaicin injections: Intra‐ and interindividual variability of provoked hyperalgesia and allodynia

Intradermal injections of glutamate and capsaicin are attractive to use in human experimental pain models because hyperalgesia and allodynia mimic isolated aspects of clinical pain disorders. The aim of the present study was to investigate the reproducibility of these models. Twenty healthy male volunteers (mean age 24years; range 18-38years) received intradermal injections of glutamate and capsaicin in the volar forearm. Magnitudes of secondary pinprick hyperalgesia and brush-evoked allodynia were investigated using von Frey filaments (gauges 10, 15, 60 and 100g) and brush strokes. Areas of secondary hyperalgesia and allodynia were quantified immediately after injection and after 15, 30 and 60min. Two identical experiments separated by at least 7days were performed. Reproducibility across and within volunteers (inter- and intra-individual variation, respectively) was assessed using intraclass correlation coefficient (ICC) and coefficient of variation (CV). Secondary pinprick hyperalgesia was observed as a marked increase in the visual analogue scale (VAS) response to von Frey gauges 60 and 100g (P<0.001) after glutamate injection. For capsaicin, secondary pinprick hyperalgesia was detected with all von Frey gauges (P<0.001). Glutamate evoked reproducible VAS response to all von Frey gauges (ICC >0.60) and brush strokes (ICC >0.83). Capsaicin injection was reproducible for secondary hyperalgesia (ICC >0.70) and allodynia (ICC >0.71). Intra-individual variability was generally lower for the VAS response to von Frey and brush compared with areas of secondary hyperalgesia and allodynia. In conclusion, glutamate and capsaicin yield reproducible hyperalgesic and allodynic responses, and the present model is well suited for basic research, as well as for assessing the modulation of central phenomena.

Cortical theta is increased while thalamocortical coherence is decreased in rat models of acute and chronic pain

Thalamocortical oscillations are critical for sensory perception. Although pain is known to disrupt synchrony in thalamocortical oscillations, evidence in the literature is controversial. Thalamocortical coherence has been reported to be increased in patients with neurogenic pain but decreased in a rat model of central pain. Moreover, theta (4 to 8Hz) oscillations in primary somatosensory (S1) cortex are speculated to predict pain in humans. To date, the link between pain and network oscillations in animal models has been understudied. Thus, we tested the hypothesis that pain disrupts thalamocortical coherence and S1 theta power in two rat models of pain. We recorded electrocorticography (ECoG) waveforms over S1 and local field potentials (LFP) within ventral posterolateral thalamus in freely behaving rats under spontaneous (stimulus-independent) pain conditions. Rats received intradermal capsaicin injection (Cap) in the hindpaw, followed hours later by chronic constriction injury (CCI) of the sciatic nerve lasting several days. Our results show that pain decreases coherence between LFP and ECoG waveforms in the 2- to 30-Hz range, and increases ECoG power in the theta range. These changes are short-lasting after Cap and longer-lasting after CCI. These data might be particularly relevant to preclinical correlates of spontaneous pain-like behavior, with potential implications to clinical biomarkers of ongoing pain.

Antiedematogenic effects of the polar fractions of Persea cordata Mez. (Lauraceae) on microvascular extravasation in rat skin

Ethnopharmacological relevancePersea cordata Mez. (Lauraceae) is a medicinal plant used in veterinary ethnopharmacology, which is a popular medicine used as an anti-inflammatory and healing agent, mainly on animal skin diseases, characterized by cutaneous open wounds, in South Brazil. Aim of this studyThe purpose of this study was to investigate a possible antiedematogenic effect of ethyl acetate (EtAc) and butanol (BuOH) polar fractions of Persea cordata on Evans blue dye leakage induced by pro-inflammatory agents in rat skin. Materials and methodsMale Wistar rats (180–200g, n=5–6) were pretreated with a single intraperitoneal administration of EtAc or BuOH (1 to 600mgkg−1) fractions followed by intravenous Evans blue dye injection (1%, 30mgkg−1, i.v.), 60min before the injection of phlogistic agents. Animals received intradermal injections (0.05ml) of carrageenan (CAR, 300µg/site), 48/80 compound (C4880, 10µg/site), histamine (HIS, 0.3µg/site), serotonin (5-HT, 0.01µg/site), dextran (DEX, 200µg/site), bradykinin (BK, 0.003µg/site), capsaicin (CPS, 400µg/site), substance P (SP, 0.003µg/site) or prostaglandin E2 (PGE2 10nmol/site) and they were submitted to euthanasia after 60min. Skin samples were obtained in the extravasation sites of Evans blue dye. Skin fragments were soaked in formamide at 37°C (during 24h) for Evans blue extraction. The amount of dye leakage in the tissue fragment was determined by a spectrophotometer (620nm). ResultsIn a very similar manner in terms of potency and efficacy, systemic administration of EtAc and BuOH fractions caused dose-dependent inhibition of vascular Evans blue dye leakage induced by phlogistic agents in the rat skin. The results obtained (ID50 values in mgkg−1 and maximal inhibition in %) with EtAc fraction, as follows were: CAR (34.42 and 63.0), 4880 (8.52 and 59.1), HIS (21.22 and 66.8), 5-HT (32.99 and 73.4), DEX (41.74 and 67.0), BK (34.03 and 68.0), CPS (100.7 and 77), SP (2.1 and 78.9) and PGE2 (133 and 71.0). BuOH fraction significantly inhibited CAR (25.9 and 70)-, 4880 (36.8 and 66)-, HIS (17.6 and 77)-, 5-HT (32.8 and 56)-, DEX (89.6 and 75)-, BK (28.0 and 66)-, CPS (136.37 and 71)-, SP (5.6 and 78)- and PGE2 (109.64 and 56)-induced VE, respectively. ConclusionSystemic administration of Persea cordata polar fractions exerts a non-specific inhibitory effect on microvascular leakage induced by pro-inflammatory agents in rat skin, probably to interfering with different biological systems involved in the development of the inflammatory process, reinforcing the popular use of Persea cordata as an anti-inflammatory and healing agent for skin.

Bilobalide, a unique constituent of Ginkgo biloba, inhibits inflammatory pain in rats

Standardized Ginkgo biloba extract EGb 761 has been shown to inhibit inflammatory hyperalgesia in rats; however, the mechanism of action is not known. This study set out to investigate the anti-inflammatory and analgesic potential of bilobalide, a unique G. biloba constituent, in three well-characterized models of acute inflammatory pain. The effect of oral, intraplantar or intrathecal administration of bilobalide or drug-vehicle (0.25% agar; 10% ethanol in H2O) on responses to noxious thermal and mechanical stimulation of the hindpaw, and paw oedema were assessed in adult male Wistar rats before and after intradermal hindpaw injection of carrageenan (3%; 50 μl) or capsaicin (10 μg; 50 μl) or after hindpaw incision (n=6-8/group). Oral administration of bilobalide (10-30 mg/kg) significantly inhibited thermal hyperalgesia in response to carrageenan, capsaicin and paw incision, independent of dose, with an efficacy similar to that of diclofenac. In the carrageenan model, mechanical hypersensitivity and paw oedema were also significantly reduced after treatment with bilobalide (10-30 mg/kg). Intrathecal administration of bilobalide (0.5-1 μg) inhibited carrageenan-induced thermal hyperalgesia, but had no effect on mechanical hypersensitivity or paw oedema (application≥2 μg induced adverse effects, precluding testing of higher doses). Intraplantar administration of bilobalide (30-100 μg) had no effect. These data show that bilobalide is a potent anti-inflammatory and antihyperalgesic agent, the therapeutic effects of which are mediated in part through a central site of action, and may account for the therapeutic action of the whole extract G. biloba.

Pinprick-evoked brain potentials: a novel tool to assess central sensitization of nociceptive pathways in humans

Although hyperalgesia to mechanical stimuli is a frequent sign in patients with inflammation or neuropathic pain, there is to date no objective electrophysiological measure for its evaluation in the clinical routine. Here we describe a technique for recording the electroencephalographic (EEG) responses elicited by mechanical stimulation with a flat-tip probe (diameter 0.25 mm, force 128 mN). Such probes activate Aδ nociceptors and are widely used to assess the presence of secondary hyperalgesia, a psychophysical correlate of sensitization in the nociceptive system. The corresponding pinprick-evoked potentials (PEPs) were recorded in 10 subjects during stimulation of the right and left hand dorsum before and after intradermal injection of capsaicin into the right hand and in 1 patient with a selective lesion of the right spinothalamic tract. PEPs in response to stimulation of normal skin were characterized by a vertex negative-positive (NP) complex, with N/P latencies and amplitudes of 111/245 ms and 3.5/11 μV, respectively. All subjects developed a robust capsaicin-induced increase in the pain elicited by pinprick stimulation of the secondary hyperalgesic area (+91.5%, P < 0.005). Such stimulation also resulted in a significant increase of the N-wave amplitude (+92.9%, P < 0.005), but not of the P wave (+6.6%, P = 0.61). In the patient, PEPs during stimulation of the hypoalgesic side were reduced. These results indicate that PEPs 1) reflect cortical activities triggered by somatosensory input transmitted in Aδ primary sensory afferents and spinothalamic projection neurons, 2) allow quantification of experimentally induced secondary mechanical hyperalgesia, and 3) have the potential to become a diagnostic tool to substantiate mechanical hyperalgesia in patients with presumed central sensitization.