Anandamide (AEA) is an important modulator of nociception in the spinal dorsal horn, acting presynaptically through Cannabinoid (CB1) and Transient receptor potential vanilloid (TRPV1) receptors. The role of AEA (1 μM, 10 μM, and 30 μM) application on the modulation of nociceptive synaptic transmission under control and inflammatory conditions was studied by recording miniature excitatory postsynaptic currents (mEPSCs) from neurons in spinal cord slices. Inhibition of the CB1 receptors by PF514273, TRPV1 by SB366791, and the fatty acid amide hydrolase (FAAH) by URB597 was used. Under naïve conditions, the AEA application did not affect the mEPSCs frequency (1.43±0.12 Hz) when all the recorded neurons were considered. The mEPSC frequency increased (180.0±39.2%) only when AEA (30 μM) was applied with PF514273 and URB597. Analysis showed that one sub-population of neurons had synaptic input inhibited (39.1% of neurons), the second excited (43.5%), whereas 8.7% showed a mixed effect and 8.7% did not respond to the AEA. With inflammation, the AEA effect was highly inhibitory (72.7%), while the excitation was negligible (9.1%), and 18.2% were not modulated. After inflammation, more neurons (45.0%) responded even to low AEA by mEPSC frequency increase with PF514273/URB597 present. AEA-induced dual (excitatory/inhibitory) effects at the 1st nociceptive synapse should be considered when developing analgesics targeting the endocannabinoid system. These findings contrast the clear inhibitory effects of the AEA precursor 20:4-NAPE application described previously and suggest that modulation of endogenous AEA production may be more favorable for analgesic treatments.

• MeSH
• amidohydrolasy MeSH
• analgetika farmakologie   MeSH
• benzamidy * MeSH
• endokanabinoidy * farmakologie   MeSH
• karbamáty * MeSH
• kyseliny arachidonové * MeSH
• lidé MeSH
• nocicepce * MeSH
• polynenasycené alkamidy farmakologie   MeSH
• zadní rohy míšní MeSH
• zánět farmakoterapie   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH

Activation of microglia in the spinal cord dorsal horn after peripheral nerve injury contributes to the development of pain hypersensitivity. How activated microglia selectively enhance the activity of spinal nociceptive circuits is not well understood. We discovered that after peripheral nerve injury, microglia degrade extracellular matrix structures, perineuronal nets (PNNs), in lamina I of the spinal cord dorsal horn. Lamina I PNNs selectively enwrap spinoparabrachial projection neurons, which integrate nociceptive information in the spinal cord and convey it to supraspinal brain regions to induce pain sensation. Degradation of PNNs by microglia enhances the activity of projection neurons and induces pain-related behaviors. Thus, nerve injury-induced degradation of PNNs is a mechanism by which microglia selectively augment the output of spinal nociceptive circuits and cause pain hypersensitivity.

• MeSH
• bolest * patologie   patofyziologie   MeSH
• extracelulární matrix patologie   MeSH
• hyperalgezie * etiologie   patologie   patofyziologie   MeSH
• krysa rodu rattus MeSH
• mikroglie * patologie   MeSH
• poranění periferního nervu * komplikace   patologie   MeSH
• potkani Sprague-Dawley MeSH
• zadní rohy míšní * patologie   patofyziologie   MeSH
• zvířata MeSH
• Check Tag
• krysa rodu rattus MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH

Second-order spinal cord excitatory neurons play a key role in spinal processing and transmission of pain signals to the brain. Exogenously induced change in developmentally imprinted excitatory neurotransmitter phenotypes of these neurons to inhibitory has not yet been achieved. Here, we use a subpial dorsal horn-targeted delivery of AAV (adeno-associated virus) vector(s) encoding GABA (gamma-aminobutyric acid) synthesizing-releasing inhibitory machinery in mice with neuropathic pain. Treated animals showed a progressive and complete reversal of neuropathic pain (tactile and brush-evoked pain behavior) that persisted for a minimum of 2.5 months post-treatment. The mechanism of this treatment effect results from the switch of excitatory to preferential inhibitory neurotransmitter phenotype in dorsal horn nociceptive neurons and a resulting increase in inhibitory activity in regional spinal circuitry after peripheral nociceptive stimulation. No detectable side effects (e.g., sedation, motor weakness, loss of normal sensation) were seen between 2 and 13 months post-treatment in naive adult mice, pigs, and non-human primates. The use of this treatment approach may represent a potent and safe treatment modality in patients suffering from spinal cord or peripheral nerve injury-induced neuropathic pain.

• MeSH
• buňky zadních rohů míšních MeSH
• mícha MeSH
• myši MeSH
• neuralgie * etiologie   terapie   MeSH
• nociceptory * MeSH
• prasata MeSH
• technika přenosu genů MeSH
• zadní rohy míšní MeSH
• zvířata MeSH
• Check Tag
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Research Support, N.I.H., Extramural MeSH

BACKGROUND: Opioid analgesics remain widely used for pain treatment despite the related serious side effects. Some of those, such as opioid tolerance and opioid-induced hyperalgesia may be at least partially due to modulation of opioid receptors (OR) function at nociceptive synapses in the spinal cord dorsal horn. It was suggested that increased release of different chemokines under pathological conditions may play a role in this process. The goal of this study was to investigate the crosstalk between the μOR, transient receptor potential vanilloid 1 (TRPV1) receptor and C-C motif ligand 2 (CCL2) chemokine and the involvement of spinal microglia in the modulation of opioid analgesia. METHODS: Patch-clamp recordings of miniature excitatory postsynaptic currents (mEPSCs) and dorsal root evoked currents (eEPSC) in spinal cord slices superficial dorsal horn neurons were used to evaluate the effect of μOR agonist [D-Ala2, N-Me-Phe4, Gly5-ol]-enkephalin (DAMGO), CCL2, TRPV1 antagonist SB366791 and minocycline. Paw withdrawal test to thermal stimuli was combined with intrathecal (i.t.) delivery of CCL2 and DAMGO to investigate the modulation in vivo. RESULTS: Application of DAMGO induced a rapid decrease of mEPSC frequency and eEPSC amplitude, followed by a delayed increase of the eESPC amplitude, which was prevented by SB366791. Chemokine CCL2 treatment significantly diminished all the DAMGO-induced changes. Minocycline treatment prevented the CCL2 effects on the DAMGO-induced eEPSC depression, while mEPSC changes were unaffected. In behavioral experiments, i.t. injection of CCL2 completely blocked DAMGO-induced thermal hypoalgesia and intraperitoneal pre-treatment with minocycline prevented the CCL2 effect. CONCLUSIONS: Our results indicate that opioid-induced inhibition of the excitatory synaptic transmission could be severely attenuated by increased CCL2 levels most likely through a microglia activation-dependent mechanism. Delayed potentiation of neurotransmission after μOR activation is dependent on TRPV1 receptors activation. Targeting CCL2 and its receptors and TRPV1 receptors in combination with opioid therapy could significantly improve the analgesic properties of opioids, especially during pathological states.

• MeSH
• anilidy farmakologie   MeSH
• chemokin CCL2 farmakologie   MeSH
• cinnamáty farmakologie   MeSH
• enkefalin, Ala(2)-MePhe(4)-Gly(5)- farmakologie   MeSH
• excitační postsynaptické potenciály účinky léků   MeSH
• krysa rodu rattus MeSH
• mícha účinky léků   MeSH
• miniaturní postsynaptické potenciály účinky léků   MeSH
• nervový přenos účinky léků   MeSH
• neurony účinky léků   MeSH
• nocicepce účinky léků   MeSH
• opioidní analgetika farmakologie   MeSH
• potkani Wistar MeSH
• zadní rohy míšní účinky léků   MeSH
• zvířata MeSH
• Check Tag
• krysa rodu rattus MeSH
• mužské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH

The mechanisms of inflammatory pain need to be identified in order to find new superior treatments. Protease-activated receptors 2 (PAR2) and transient receptor potential vanilloid 1 (TRPV1) are highly co-expressed in dorsal root ganglion neurons and implicated in pain development. Here, we examined the role of spinal PAR2 in hyperalgesia and the modulation of synaptic transmission in carrageenan-induced peripheral inflammation, using intrathecal (i.t.) treatment in the behavioral experiments and recordings of spontaneous, miniature and dorsal root stimulation-evoked excitatory postsynaptic currents (sEPSCs, mEPSCs and eEPSCs) in spinal cord slices. Intrathecal PAR2-activating peptide (AP) administration aggravated the carrageenan-induced thermal hyperalgesia, and this was prevented by a TRPV1 antagonist (SB 366791) and staurosporine i.t. pretreatment. Additionally, the frequency of the mEPSC and sEPSC and the amplitude of the eEPSC recorded from the superficial dorsal horn neurons were enhanced after acute PAR2 AP application, while prevented with SB 366791 or staurosporine pretreatment. PAR2 antagonist application reduced the thermal hyperalgesia and decreased the frequency of mEPSC and sEPSC and the amplitude of eEPSC. Our findings highlight the contribution of spinal PAR2 activation to carrageenan-induced hyperalgesia and the importance of dorsal horn PAR2 and TRPV1 receptor interactions in the modulation of nociceptive synaptic transmission.

• MeSH
• anilidy farmakologie   MeSH
• buňky zadních rohů míšních účinky léků   metabolismus   fyziologie   MeSH
• cinnamáty farmakologie   MeSH
• excitační postsynaptické potenciály MeSH
• hyperalgezie etiologie   metabolismus   patofyziologie   MeSH
• karagenan farmakologie   toxicita   MeSH
• kationtové kanály TRPV antagonisté a inhibitory   metabolismus   MeSH
• krysa rodu rattus MeSH
• miniaturní postsynaptické potenciály MeSH
• nocicepce MeSH
• potkani Wistar MeSH
• receptor PAR-2 metabolismus   MeSH
• staurosporin farmakologie   MeSH
• zvířata MeSH
• Check Tag
• krysa rodu rattus MeSH
• mužské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH

This study investigated the effects of wheel-running using the upper limbs following immobilization after inducing arthritis in the knees of rats. Forty male Wistar rats (aged 8 weeks) divided into four groups randomly: arthritis (AR), immobilization after arthritis (Im), wheel-running exercise with the upper limbs following immobilization after arthritis induction (Im+Ex) and sham arthritis induction (Con). The knee joints of the Im and Im+Ex groups were immobilized with a cast for 4 weeks. In the Im+Ex group, wheel-running exercise was administered for 60 min/day (5 times/week). The swelling and the pressure pain threshold (PPT) of the knee joint were evaluated for observing the condition of inflammatory symptoms in affected area, and the paw withdraw response (PWR) was evaluated for observing the condition of secondary hyperalgesia in distant area. Especially, in order to evaluate histological inflammation in the knee joint, the number of macrophage (CD68-positive cells) in the synovium was examined. The expression of calcitonin gene-related peptide (CGRP) in the spinal dorsal horn (L2-3 and L4-5) was examined to evaluate central sensitization. The Im+Ex group showed a significantly better recovery than the Im group in the swelling, PPTs, and PWRs. Additionally, CGRP expression of the spinal dorsal horn (L2-3 and L4-5) in the Im+Ex group was significantly decreased compared with the Im group. According to the results, upper limb exercise can decrease pain in the affected area, reduce hyperalgesia in distant areas, and suppress the central sensitization in the spinal dorsal horn by triggering exercise-induced hypoalgesia (EIH).

• MeSH
• artritida etiologie   patologie   rehabilitace   MeSH
• bolest patologie   prevence a kontrola   MeSH
• horní končetina fyziologie   MeSH
• imobilizace metody   MeSH
• kolenní kloub patofyziologie   MeSH
• kondiční příprava zvířat metody   MeSH
• krysa rodu rattus MeSH
• modely nemocí na zvířatech MeSH
• potkani Wistar MeSH
• zadní rohy míšní patologie   MeSH
• zánět patologie   prevence a kontrola   MeSH
• zvířata MeSH
• Check Tag
• krysa rodu rattus MeSH
• mužské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH

• MeSH
• lidé MeSH
• mozková kůra fyziologie   MeSH
• nervové dráhy anatomie a histologie   fyziologie   MeSH
• nervový přenos fyziologie   MeSH
• nocicepce * fyziologie   MeSH
• nociceptory fyziologie   MeSH
• periferní nervy anatomie a histologie   fyziologie   MeSH
• porodní bolesti * MeSH
• signální transdukce fyziologie   MeSH
• těhotenství MeSH
• zadní rohy míšní fyziologie   MeSH
• Check Tag
• lidé MeSH
• těhotenství MeSH
• ženské pohlaví MeSH

Glial cells activated by peripheral nerve injury contribute to the induction and maintenance of neuropathic pain by releasing neuromodulating cytokines and chemokines. We investigated the activation of microglia and astrocytes as well as the cellular distribution of the chemokine CCL2 and its receptor CCR2 in the trigeminal subnucleus caudalis (TSC) ipsilateral and contralateral to infraorbital nerve ligature (IONL). The left infraorbital nerve was ligated under aseptic conditions, and sham controls were operated without nerve ligature. Tactile hypersensitivity was significantly increased bilaterally in vibrissal pads of both sham- and IONL-operated animals from day 1 to 7 and tended to normalize in sham controls surviving for 14 days. Activated microglial cells significantly increased bilaterally in the TSC of both sham- and IONL-operated animals with a marked but gradual increase in the ipsilateral TSC from 1 to 7 days followed by a decrease by day 14. In contrast, robust activation of astrocytes was found bilaterally in the TSC of IONL-operated rats from 3 to 14 days with a transient activation in the ipsilateral TSC of sham-operated animals. Cellular distribution of CCL2 varied with survival time. CCL2 immunofluorescence was detected in neurons within 3 days and in astrocytes at later time points. In contrast, CCR2 was found only in astrocytes at all time points with CCR2 intensity being dominant in the ipsilateral TSC. In summary, our results reveal bilateral activation of microglial cells and astrocytes as well as changes in the cellular distribution of CCL2 and its receptor CCR2 in the TSC during the development and maintenance of orofacial neuropathic pain.

• MeSH
• chemokin CCL2 metabolismus   MeSH
• krysa rodu rattus MeSH
• modely nemocí na zvířatech MeSH
• neuralgie metabolismus   MeSH
• neuroglie metabolismus   MeSH
• potkani Wistar MeSH
• receptory CCR2 metabolismus   MeSH
• substantia gelatinosa metabolismus   MeSH
• zvířata MeSH
• Check Tag
• krysa rodu rattus MeSH
• mužské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH

Paclitaxel chemotherapy treatment often leads to neuropathic pain resistant to available analgesic treatments. Recently spinal Toll-like receptor 4 (TLR4) and the transient receptor potential cation channel subfamily V member 1 (TRPV1) were identified to be involved in the pro-nociceptive effect of paclitaxel. The aim of this study was to investigate the role of phosphatidylinositol 3-kinase (PI3K) and serine/threonine kinases in this process, with the use of their antagonists (wortmannin, LY-294002, and staurosporine). The single paclitaxel administration (8 mg/kg i.p.) in mice induced robust mechanical allodynia measured as a reduced threshold to von Frey filament stimulation and generated reduced tachyphylaxis of capsaicin-evoked responses, recorded as changes in mEPSC frequency in patch-clamp recordings of dorsal horn neurons activity in vitro, for up to eight days. Paclitaxel application also induced increased Akt kinase phosphorylation in rat DRG neurons. All these paclitaxel-induced changes were prevented by the wortmannin in vivo pretreatment. Acute co-application of wortmannin or LY-294002 with paclitaxel in spinal cord slices also attenuated the paclitaxel effect on capsaicin-evoked responses. Staurosporine was effective in the acute in vitro experiments and on the first day after the paclitaxel treatment in vivo, but in contrast to wortmannin, it did not have a significant impact later. Our data suggest that the inhibition of PI3K signaling may help alleviate pathological pain syndromes in the paclitaxel-induced neuropathy.

• MeSH
• buňky zadních rohů míšních účinky léků   metabolismus   MeSH
• excitační postsynaptické potenciály účinky léků   MeSH
• fosfatidylinositol-3-kinasy metabolismus   MeSH
• hyperalgezie chemicky indukované   farmakoterapie   metabolismus   MeSH
• inhibitory proteinkinas farmakologie   MeSH
• kapsaicin farmakologie   MeSH
• kationtové kanály TRP MeSH
• kationtové kanály TRPV metabolismus   MeSH
• krysa rodu rattus MeSH
• lipopolysacharidy farmakologie   MeSH
• mícha účinky léků   metabolismus   MeSH
• myši inbrední C57BL MeSH
• myši MeSH
• neuralgie chemicky indukované   farmakoterapie   metabolismus   MeSH
• onkogenní protein v-akt metabolismus   MeSH
• paclitaxel toxicita   MeSH
• peptidové fragmenty imunologie   metabolismus   MeSH
• potkani Wistar MeSH
• protein-serin-threoninkinasy antagonisté a inhibitory   metabolismus   MeSH
• proteinkinasa C imunologie   metabolismus   MeSH
• signální transdukce účinky léků   MeSH
• toll-like receptor 4 metabolismus   MeSH
• zvířata MeSH
• Check Tag
• krysa rodu rattus MeSH
• mužské pohlaví MeSH
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

BACKGROUND AND PURPOSE: Endocannabinoids play an important role in modulating spinal nociceptive signalling, crucial for the development of pain. The cannabinoid CB1 receptor and the TRPV1 cation channel are both activated by the endocannabinoid anandamide, a product of biosynthesis from the endogenous lipid precursor N-arachidonoylphosphatidylethanolamine (20:4-NAPE). Here, we report CB1 receptor- and TRPV1-mediated effects of 20:4-NAPE on spinal synaptic transmission in control and inflammatory conditions. EXPERIMENTAL APPROACH: Spontaneous (sEPSCs) and dorsal root stimulation-evoked (eEPSCs) excitatory postsynaptic currents from superficial dorsal horn neurons in rat spinal cord slices were assessed. Peripheral inflammation was induced by carrageenan. Anandamide concentration was assessed by mass spectrometry. KEY RESULTS: Application of 20:4-NAPE increased anandamide concentration in vitro. 20:4-NAPE (20 μM) decreased sEPSCs frequency and eEPSCs amplitude in control and inflammatory conditions. The inhibitory effect of 20:4-NAPE was sensitive to CB1 receptor antagonist PF514273 (0.2 μM) in both conditions, but to the TRPV1 antagonist SB366791 (10 μM) only after inflammation. After inflammation, 20:4-NAPE increased sEPSCs frequency in the presence of PF514273 and this increase was blocked by SB366791. CONCLUSIONS AND IMPLICATIONS: While 20:4-NAPE treatment inhibited the excitatory synaptic transmission in both naive and inflammatory conditions, peripheral inflammation altered the underlying mechanisms. Our data indicate that 20:4-NAPE application induced mainly CB1 receptor-mediated inhibitory effects in naive animals while TRPV1-mediated mechanisms were also involved after inflammation. Increasing anandamide levels for analgesic purposes by applying substrate for its local synthesis may be more effective than systemic anandamide application or inhibition of its degradation. LINKED ARTICLES: This article is part of a themed section on Recent Advances in Targeting Ion Channels to Treat Chronic Pain. To view the other articles in this section visit http://onlinelibrary.wiley.com/doi/10.1111/bph.v175.12/issuetoc.

• MeSH
• buňky zadních rohů míšních účinky léků   metabolismus   MeSH
• fosfatidylethanolaminy chemická syntéza   chemie   farmakologie   MeSH
• hmotnostní spektrometrie MeSH
• karagenan MeSH
• krysa rodu rattus MeSH
• mícha účinky léků   metabolismus   MeSH
• nervový přenos účinky léků   MeSH
• potkani Wistar MeSH
• vztah mezi dávkou a účinkem léčiva MeSH
• vztahy mezi strukturou a aktivitou MeSH
• zánět chemicky indukované   metabolismus   MeSH
• zvířata MeSH
• Check Tag
• krysa rodu rattus MeSH
• mužské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH