Peripheral Inflammation In Rat Research Articles

Bilateral subdiaphragmatic vagotomy modulates the peripheral met‑enkephalin and striatal monoamine responses to peripheral inflammation in rat.

In the central nervous system, long‑term effects of a vagotomy include disturbance of monoaminergic activity of the limbic system. Since low vagal activity is observed in major depression and autism spectrum disorder, the study aimed to determine whether animals fully recovered after subdiaphragmatic vagotomy demonstrates neurochemical indicators of altered well‑being and social component of sickness behavior. Bilateral vagotomy or sham surgery was performed in adult rats. After one month of recovery, rats were challenged with lipopolysaccharide or vehicle to determine the role of central signaling upon sickness. Striatal monoamines and met‑enkephalin concentrations were evaluated using HPLC and RIA methods. We also defined a concentration of immune‑derived plasma met‑enkephalin to establish a long‑term effect of vagotomy on peripheral analgesic mechanisms. The data indicate that 30 days after vagotomy procedure, striatal dopaminergic, serotoninergic, and enkephalinergic neurochemistry was altered, both under physiological and inflammatory conditions. Vagotomy prevented inflammation‑induced increases of plasma met‑enkephalin - an opioid analgesic. Our data suggest that in a long perspective, vagotomized rats may be more sensitive to pain and social stimuli during peripheral inflammation.

Spinal PAR2 Activation Contributes to Hypersensitivity Induced by Peripheral Inflammation in Rats.

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.

Up-regulation of Cathepsin G in the Development of Chronic Postsurgical Pain: An Experimental and Clinical Genetic Study.

Proteases have been shown to modulate pain signaling in the spinal cord and may contribute to the development of chronic postsurgical pain. By using peripheral inflammation in rats as a chronic pain model, the authors identified the deregulation of proteases and their inhibitors as a hallmark of chronic pain development using a genome-wide screening approach. A microarray analysis was performed and identified spinal cathepsin G (CTSG) as the most up-regulated gene in rats with persistent hyperalgesia after intraplantar injection of complete Freund's adjuvant (n = 4). Further experiments were performed to elucidate the mechanisms of CTSG-induced hyperalgesia by intrathecally applying specific CTSG inhibitor (n = 10). The authors also evaluated the association between CTSG gene polymorphisms and the risk of chronic postsurgical pain in 1,152 surgical patients. CTSG blockade reduced heat hyperalgesia, accompanied by a reduction in neutrophil infiltration and interleukin 1β levels in the dorsal horns. In the gene association study, 246 patients (21.4%) reported chronic postsurgical pain at 12-month follow-up. Patients with AA genotypes at polymorphisms rs2070697 (AA-15.3%, GA-24.1%, and GG-22.3%) or rs2236742 (AA-6.4%, GA-20.4%, and GG-22.6%) in the CTSG gene had lower risk for chronic postsurgical pain compared with wild-types. The adjusted odds ratios were 0.67 (95% CI, 0.26 to 0.99) and 0.34 (95% CI, 0.21 to 0.98), respectively. This study demonstrated that CTSG is a pronociceptive mediator in both animal model and human study. CTSG represents a new target for pain control and a potential marker to predict patients who are prone to develop chronic pain after surgery.

Activation of spinal NF-κB/p65 contributes to peripheral inflammation and hyperalgesia in rat adjuvant-induced arthritis.

It is known that noxious stimuli from inflamed tissue may increase the excitability of spinal dorsal horn neurons (a process known as central sensitization), which can signal back and contribute to peripheral inflammation. However, the underlying mechanisms have yet to be fully defined. A number of recent studies have indicated that spinal NF-κB/p65 is involved in central sensitization, as well as pain-related behavior. Thus, the aim of this study was to determine whether NF-κB/p65 can facilitate a peripheral inflammatory response in rat adjuvant-induced arthritis (AIA). Lentiviral vectors encoding short hairpin RNAs that target NF-κB/p65 (LV-shNF-κB/p65) were constructed for gene silencing. The spines of rats with AIA were injected with LV-shNF-κB/p65 on day 3 or day 10 after treatment with Freund's complete adjuvant (CFA). During an observation period of 20 days, pain-related behavior, paw swelling, and joint histopathologic changes were evaluated. Moreover, the expression levels of spinal tumor necrosis factor α (TNFα), interleukin-1β (IL-1β), and cyclooxygenase 2 (COX-2) were assessed on day 14 after CFA treatment. The presence of peripheral inflammation in rats with AIA induced an increase in NF-κB/p65 expression in the spinal cord, mainly in the dorsal horn neurons and astrocytes. Delivery of LV-shNF-κB/p65 to the spinal cord knocked down the expression of NF-κB/p65 and significantly attenuated hyperalgesia, paw edema, and joint destruction. In addition, spinal delivery of LV-shNF-κB/p65 reduced the overexpression of spinal TNFα, IL-1β, and COX-2. These findings indicate that spinal NF-κB/p65 plays an important role in the initiation and development of both peripheral inflammation and hyperalgesia. Thus, inhibition of spinal NF-κB/p65 expression may provide a potential treatment to manage painful inflammatory disorders.

Spinal matrix metalloproteinase 3 mediates inflammatory hyperalgesia via a tumor necrosis factor-dependent mechanism

Matrix metalloproteinases (MMPs) have been implicated in the modulation of synaptic plasticity, glial activation, and long-term potentiation in the CNS. Here we demonstrate for the first time a mechanism for the regulation of nociceptive processing by spinal MMP-3 during peripheral inflammation. We first determined by western blotting that the catalytic (active) form of MMP-3 (cMMP-3) is increased in lumbar spinal cord following peripheral inflammation in rats. The peripheral inflammation-induced thermal hyperalgesia and tactile hypersensitivity was transiently (2-3 h) attenuated by intrathecal (IT) pretreatment with either an MMP-3 inhibitor (NNGH), or a broad spectrum MMP inhibitor (GM6001). In addition, IT delivery of cMMP-3 evoked hypersensitivity, whereas the pro (enzymatically inactive) form of MMP-3 did not. This suggests a pro-algesic effect of spinal MMP-3 mediated by an enzymatic mechanism. This cMMP-3-induced hypersensitivity is concurrent with increased tumor necrosis factor (TNF) in the spinal cord. The hypersensitivity behavior is prevented by intrathecal etanercept (TNF blockade). Treatment with cMMP-3 resulted in an increase in TNF release from spinal primary microglial, but not astrocyte cultures. These findings thus present direct evidence implicating MMP-3 in the coordination of spinal nociceptive processing via a spinal TNF-dependent mechanism.

Specific Involvement of Atypical PKCξ/PKMξ in Spinal Persistent Nociceptive Processing following Peripheral Inflammation in Rat

BackgroundCentral sensitization requires the activation of various intracellular signalling pathways within spinal dorsal horn neurons, leading to a lowering of activation threshold and enhanced responsiveness of these cells. Such plasticity contributes to the manifestation of chronic pain states and displays a number of features of long-term potentiation (LTP), a ubiquitous neuronal mechanism of increased synaptic strength. Here we describe the role of a novel pathway involving atypical PKCζ/PKMζ in persistent spinal nociceptive processing, previously implicated in the maintenance of late-phase LTP.ResultsUsing both behavioral tests and in vivo electrophysiology in rats, we show that inhibition of this pathway, via spinal delivery of a myristoylated protein kinase C-ζ pseudo-substrate inhibitor, reduces both pain-related behaviors and the activity of deep dorsal horn wide dynamic range neurons (WDRs) following formalin administration. In addition, Complete Freund's Adjuvant (CFA)-induced mechanical and thermal hypersensitivity was also reduced by inhibition of PKCζ/PKMζ activity. Importantly, this inhibition did not affect acute pain or locomotor behavior in normal rats and interestingly, did not inhibited mechanical allodynia and hyperalgesia in neuropathic rats. Pain-related behaviors in both inflammatory models coincided with increased phosphorylation of PKCζ/PKMζ in dorsal horn neurons, specifically PKMζ phosphorylation in formalin rats. Finally, inhibition of PKCζ/PKMζ activity decreased the expression of Fos in response to formalin and CFA in both superficial and deep laminae of the dorsal horn.ConclusionsThese results suggest that PKCζ, especially PKMζ isoform, is a significant factor involved in spinal persistent nociceptive processing, specifically, the manifestation of chronic pain states following peripheral inflammation.

Comparison of a bupivacaine peripheral nerve block and systemic ketoprofen on peripheral inflammation and hyperalgesia in rats

Local anaesthetics via a nerve block can attenuate inflammation. However, their effects have never been compared with the anti-inflammatory effects of systemic NSAIDs. The aim of this study was to compare the effects of bupivacaine via sciatic block with a systemic NSAID on oedema and hyperalgesia and on indices of systemic inflammation as measured by cytokines and prostaglandin E2 production in a model of peripheral inflammation in rats. Rats (n=56) randomly received three injections: a left subcutaneous hind paw injection (carrageenan or physiological saline); a left sciatic block (bupivacaine or physiological saline); and a systemic injection (ketoprofen or physiological saline). Six hours later, local oedema, thermal and mechanical hyperalgesia as well as cerebrospinal fluid prostaglandin E2 concentration and tumour necrosis factor-alpha and interleukin 1beta-stimulated production in whole blood cultures were measured. A sciatic block with bupivacaine as well as a systemic injection of NSAID significantly decreased the oedema and the thermal and mechanical hyperalgesia induced by carrageenan. Oedema was decreased by approximately 12%, and thermal and mechanical thresholds were increased by three-fold to fourfold and 1.5-2-fold, respectively. They both inhibited the increased production of prostaglandin E2 in cerebrospinal fluid and tumour necrosis factor-alpha and interleukin 1beta in stimulated whole blood induced by carrageenan. There was no further benefit of the combination of a sciatic block and systemic NSAID. In the current study, a bupivacaine block alone achieved the same anti-inflammatory effect as systemic NSAID or as when the same block is combined with a NSAID.

Adjuvant effect of caffeine on acetylsalicylic acid anti-nociception: Prostaglandin E2 synthesis determination in carrageenan-induced peripheral inflammation in rat

In the present study, we report a synergistic interaction between acetylsalicylic acid (ASA) and caffeine (CAF) on the inhibition of nociception in a model of peripheral inflammation in rat; on the contrary no interaction have been found on anti-inflammatory effects and peripheral prostaglandin E2 (PGE-2) synthesis inhibition. Acute inflammation was induced by the subplantar injection of carrageenan into the right hind paw, and the effects of the drugs were evaluated from 0 to 5 h. Nociception was assessed using the Randall & Selitto test, and the inflammatory response by plethismometry. Oral administration of ASA (10–400 mg/kg) induced dose-related anti-nociceptive and anti-inflammatory effects. On the other hand, oral CAF administration (5–50 mg/kg) did not show a dose-related inhibitory effect, neither on the inhibition of nociception nor on the inflammatory response. To analyze a possible interaction between both drugs a dose–response curve to ASA plus a fixed dose of CAF (5 mg/kg) was obtained 3 h after the injection of carrageenan, when the inflammatory pain peaked. A fixed dose of CAF was able to improve the anti-nociceptive, but not the anti-inflammatory, effects of ASA. We also assessed, by enzyme immunoassay, the effects of the combination on peripheral PGE-2 levels: CAF did not alter the inhibitory effect of ASA on PGE-2 synthesis. Our results corroborate the well-known clinical effects of combining ASA and CAF; on the other hand, we rule out that prostaglandin synthesis inhibition at peripheral sites would be the mechanism responsible of the adjuvant anti-nociceptive effect of CAF.

O.344 Comparison of the post-extraction bleeding in patients with and without suspension of aspirin 100 mg daily

In the present study, we report a synergistic interaction between acetylsalicylic acid (ASA) and caffeine (CAF) on the inhibition of nociception in a model of peripheral inflammation in rat; on the contrary no interaction have been found on anti-inflammatory effects and peripheral prostaglandin E2 (PGE-2) synthesis inhibition. Acute inflammation was induced by the subplantar injection of carrageenan into the right hind paw, and the effects of the drugs were evaluated from 0 to 5 h. Nociception was assessed using the Randall & Selitto test, and the inflammatory response by plethismometry. Oral administration of ASA (10–400 mg/kg) induced dose-related anti-nociceptive and anti-inflammatory effects. On the other hand, oral CAF administration (5–50 mg/kg) did not show a dose-related inhibitory effect, neither on the inhibition of nociception nor on the inflammatory response. To analyze a possible interaction between both drugs a dose–response curve to ASA plus a fixed dose of CAF (5 mg/kg) was obtained 3 h after the injection of carrageenan, when the inflammatory pain peaked. A fixed dose of CAF was able to improve the anti-nociceptive, but not the anti-inflammatory, effects of ASA. We also assessed, by enzyme immunoassay, the effects of the combination on peripheral PGE-2 levels: CAF did not alter the inhibitory effect of ASA on PGE-2 synthesis. Our results corroborate the well-known clinical effects of combining ASA and CAF; on the other hand, we rule out that prostaglandin synthesis inhibition at peripheral sites would be the mechanism responsible of the adjuvant anti-nociceptive effect of CAF.

The anti-inflammatory effects of 2 Hz electroacupuncture with different intensities on acute carrageenan-induced inflammation in the rat paw

The aim of this study was to investigate the anti-inflammatory effects of low frequency electroacupuncture (EA) with different intensities in carrageenan-injected rat paw. Bilateral 2 Hz EA stimulation with 0.5, 1 and 3 mA were delivered at those acupoints corresponding to Zusanli and Sanyinjiao in man via needles for 30 min. We first examined the degrees of edema and hyperalgesia using measurements of paw swelling and hot plate latency over 180 min, at 30 min intervals, after the carrageenan injection. The edema and thermal sensitivities of the hind paw induced by a carrageenan-injection were strongly inhibited by EA stimulation, especially at low intensity. At 3 h after carrageenan-injection, we investigated whether the effects of a 2 Hz EA on a carrageenan-induced edema and hyperalgesia are associated with peripheral cyclooxygenase (COX) mRNA induction and prostaglandin E2 (PGE2) synthesis. Although the induction of COX-2 mRNA was inhibited by 2 Hz EA at various intensities, there was no significant change. However, the synthesis of PGE2 induced by a carrageenan injection was significantly attenuated in rat paw with low intensity EA treatment. The data indicate that low frequency EA treatment with relatively low intensity might be a useful therapy for mitigation of inflammatory edema and hyperalgesia through the regulation of COX-2 expression and PGE2 production in a model of peripheral inflammation in rats.

Asian scorpion BmK venom induces plasma extravasation and thermal hyperalgesia in the rat

In the present study, the effects of scorpion Buthus martensi Karsch (BmK) venom on plasma extravasation and paw withdrawal latency (PWL) to radiant heat have been investigated in rats. BmK venom (20–200 μg/kg) by subcutaneous injection under the surface of the rat hindpaw causes dose-dependant increased plasma extravasation that could be partially inhibited by intraperitoneal (i.p.) injected morphine (6 mg/kg). Peak plasma extravasation was reached at 10 min and persisted for 60 min at a dose of 200 μg/kg. BmK venom induced cutaneous hyperalgesia as indicated by decreased PWL to radiant heat in the ipsilateral paw following subcutaneous injection of 20 μg/kg BmK venom without effect on PWL of the contralateral hindpaw. Meanwhile, it was found that i.p. morphine injection could inhibit this decreased ipsilateral PWL. The results thus suggest that BmK venom could induce peripheral inflammation in rat by subcutaneous injection, and may prove a valuable animal model for investigating the pathophysiology of a number of inflammatory diseases and identifying potential anti-inflammatory and analgesic drugs.

Role of endogenous interleukin-1 receptor antagonist in regulating fever induced by localised inflammation in the rat.

1. Interleukin (IL)-1 is a mediator of host defence responses to inflammation and injury, including fever, but its sites of synthesis and action have not been fully elucidated. The actions of IL-1 are antagonised by IL-1 receptor antagonist (IL-1ra). The present study tested the hypothesis that IL-1 and IL-1ra are produced locally at sites of peripheral inflammation in rats, and that endogenous IL-1ra acts to limit the fever resulting from the inflammation. 2. Injection of lipopolysaccharide (LPS; 100 microg kg-1) into a subcutaneous air pouch (I.PO.) of rats induced a significant increase in body temperature. Virtually all (approximately 85 %) of the injected LPS was recovered from the pouch between 1 and 8 h (when the experiment was terminated) after injection of LPS, but LPS was undetectable (< 50 pg ml-1) in plasma at any time. Concentrations of immunoreactive IL-1alpha and IL-1beta were increased significantly in the pouch at 1, 2, 3, 5 and 8 h after injection of LPS, corresponding with the rise in body temperature and the fever peak. The appearance of IL-1ra was delayed until 2 h. Thereafter, the concentrations of IL-1beta and IL-1ra increased in parallel with the development of fever, while the concentrations of IL-1alpha remained constant. IL-1ra, but not IL-1alpha or IL-1bet, was detected in significant quantities in the plasma of LPS-injected animals. 3. Treatment of rats with an anti-IL-1ra serum (2 ml, I.PO.) at the time of injection of LPS (10 or 100 microg kg-1, I.PO.) abolished the appearance of IL-1ra in the circulation. Although neutralisation of endogenous IL-1ra did not affect the maximum body temperature reached after injection of submaximum (10 microg kg-1, I.PO.) or maximum (100 microg kg-1, I.PO.) doses of LPS, the duration of the fever was significantly prolonged, and was associated with a 3- to 4-fold increase in immunoreactive IL-1beta concentrations in the pouch fluid, but not in the plasma, at the 8 h time point. 4. These data show that effects of local (I.PO.) injection of LPS are not due to its action in the circulation or at distant sites (such as at the blood-brain barrier). These data also show that locally produced IL-1ra, in response to injection (I.PO.) of LPS, inhibits the production and/or action of locally produced IL-1beta. The ability of IL-1ra to limit the duration, rather than the magnitude of the fever, is consistent with its delayed production, relative to IL-IL-1ra, therefore, appears to play a key role in the resolution of fever induced by localised inflammatory responses.