Descending facilitatory pain pathways mediate ongoing pain and tactile hypersensitivity in a rat model of trigeminal neuropathic pain.

The Chronic Constriction Injury of the Infraorbital Nerve (CCI-ION) is a well-established model to study facial sensory changes related to trigeminal neuropathic pain. CCI-ION induces heat hypersensitivity that resolves within 2-3 weeks and a delayed mechanical hypersensitivity that emerges during the second week post-injury. The role of descending facilitatory pain pathways from the rostro ventromedial medulla (RVM) in mediating the heat and tactile hypersensitivity was examined. CCI-ION induced heat hypersensitivity observed 5days post-surgery was reversed by systemic, but not RVM lidocaine. CCI-ION-induced tactile hypersensitivity observed 15days post-surgery was reversed by systemic lidocaine and attenuated by RVM lidocaine. CCI-ION-induced spontaneous pain was determined using conditioned place preference (CPP) to pain relief at each time-point. At day 5 post-CCI-ION, neither systemic nor RVM lidocaine induced CPP. However, at 15days post-CCI-ION, CPP was observed to the chamber paired with RVM lidocaine, but not systemic lidocaine. These data indicate that CCI-ION induced heat hypersensitivity is not dependent on descending facilitatory pain pathways 5-days post-injury whereas descending facilitatory pain pathways mediate tactile allodynia and spontaneous pain 15days post-CCI-ION. This suggests that CCI-ION induces early peripheral sensitization followed by development of central sensitization that mediates spontaneous pain and contributes to mechanical hypersensitivity.

Potential role for ET-2 acting through ETA receptors in experimental colitis in mice.

OBJECTIVE AND DESIGN: This study attempted to clarify the roles of endothelins and mechanisms associated with ETA/ETB receptors in mouse models of colitis. MATERIALS AND METHODS: Colitis was induced by intracolonic administration of 2,4,6-trinitrobenzene sulfonic acid (TNBS, 1.5 mg/animal) or dextran sulfate sodium (DSS, 3%). After colitis establishment, mice received Atrasentan (ETA receptor antagonist, 10 mg/kg), A-192621 (ETB receptor antagonist, 20 mg/kg) or Dexamethasone (1 mg/kg) and several inflammatory parameters were assessed, as well as mRNA levels for ET-1, ET-2 and ET receptors. RESULTS: Atrasentan treatment ameliorates TNBS- and DSS-induced colitis. In the TNBS model was observed reduction in macroscopic and microscopic score, colon weight, neutrophil influx, IL-1ß, MIP-2 and keratinocyte chemoattractant (KC) levels, inhibition of adhesion molecules expression and restoration of IL-10 levels. However, A192621 treatment did not modify any parameter. ET-1 and ET-2 mRNA was decreased 24 h, but ET-2 mRNA was markedly increased at 48 h after TNBS. ET-2 was able to potentiate LPS-induced KC production in vitro. ETA and ETB receptors mRNA were increased at 24, 48 and 72 h after colitis induction. CONCLUSIONS: Atrasentan treatment was effective in reducing the severity of colitis in DSS- and TNBS-treated mice, suggesting that ETA receptors might be a potential target for inflammatory bowel diseases.

Exacerbation of DSS-induced colitis in mice lacking kinin B(1) receptors through compensatory up-regulation of kinin B(2) receptors: the role of tight junctions and intestinal homeostasis.

BACKGROUND AND PURPOSE: Kinins are pro-inflammatory peptides that are released during tissue injury, including that caused by inflammatory bowel disease. Herein, we assessed the role and underlying mechanisms through which the absence of kinin B(1) receptors exacerbates the development of dextran sulfate sodium (DSS)-induced colitis in mice. EXPERIMENTAL APPROACH: B(1) and B(2) receptor antagonists and B(1) receptor knockout mice (B1(-/-) ) were used to assess the involvement of B(1) and B(2) receptor signalling in a DSS-colitis. B(1) receptor, B(2) receptor, occludin and claudin-4 expression, cytokine levels and cell permeability were evaluated in colon from wild-type (WT) and B1(-/-) mice. KEY RESULTS: DSS-induced colitis was significantly exacerbated in B1(-/-) compared with WT mice. IL-1ß, IFN-γ, keratinocyte-derived chemokine and macrophage inflammatory protein-2 were markedly increased in the colon from DSS-treated B1(-/-) compared with DSS-treated WT mice. Treatment of WT mice with a selective B(1) receptor antagonist, DALBK or SSR240612, had no effect on DSS-induced colitis. Of note, B(2) receptor mRNA expression was significantly up-regulated in colonic tissue from the B1(-/-) mice after DSS administration. Moreover, treatment with a selective B(2) receptor antagonist prevented the exacerbation of colitis in B1(-/-) mice following DSS administration. The water- or DSS-treated B1(-/-) mice showed a decrease in occludin gene expression, which was partially prevented by the B(2) receptor antagonist. CONCLUSIONS AND IMPLICATIONS: A loss of B(1) receptors markedly exacerbates the severity of DSS-induced colitis in mice. The increased susceptibility of B1(-/-) may be associated with compensatory overexpression of B(2) receptors, which, in turn, modulates tight junction expression.

Inhibitor of PI3Kγ ameliorates TNBS-induced colitis in mice by affecting the functional activity of CD4+CD25+FoxP3+ regulatory T cells.

BACKGROUND AND PURPOSE: Phosphoinositide 3-kinase-γ (PI3Kγ) is implicated in many pathophysiological conditions, and recent evidence has suggested its involvement in colitis. In the present study, we investigated the effects of AS605240, a relatively selective PI3Kγ inhibitor, in experimental colitis and its underlying mechanisms. EXPERIMENTAL APPROACH: Acute colitis was induced in mice by treatment with trinitrobenzene sulphonic acid (TNBS), and the effect of AS605240 on colonic injury was assessed. Pro-inflammatory mediators and cytokines were measured by immunohistochemistry, elisa, real time-polymerase chain reaction and flow cytometry. KEY RESULTS: Oral administration of AS605240 significantly attenuated TNBS-induced acute colitis and diminished the expression of matrix metalloproteinase-9 and vascular endothelial growth factor. The colonic levels and expression of IL-1ß, CXCL-1/KC, MIP-2 and TNF-α were also reduced following therapeutic treatment with AS605240. Moreover, AS605240 reduced MIP-2 levels in a culture of neutrophils stimulated with lipopolysaccharide. The mechanisms underlying these actions of AS605240 are related to nuclear factor-κ (NF-κB) inhibition. Importantly, the PI3Kγ inhibitor also up-regulated IL-10, CD25 and FoxP3 expression. In addition, a significant increase in CD25 and FoxP3 expression was found in isolated lamina propria CD4+ T cells of AS605240-treated mice. The effect of AS605240 on Treg induction was further confirmed by showing that concomitant in vivo blockade of IL-10R significantly attenuated its therapeutic activity. CONCLUSIONS AND IMPLICATIONS: These results suggest that AS605240 protects mice against TNBS-induced colitis by inhibiting multiple inflammatory components through the NF-κB pathway while simultaneously inducing an increase in the functional activity of CD4+CD25+ Treg. Thus, AS605240 may offer a promising new therapeutic strategy for the treatment of inflammatory bowel diseases.

Intraplantar PGE2 causes nociceptive behaviour and mechanical allodynia: the role of prostanoid E receptors and protein kinases.

BACKGROUND AND PURPOSE: Receptor subtypes involved in PGE(2)-induced nociception are still controversial. The present study investigated the prostanoid E receptor (EP) subtypes and the protein kinase (PK) pathways involved in the nociception induced by PGE(2) injection in the mouse paw. EXPERIMENTAL APPROACH: Paw-licking and mechanical allodynia were measured in vivo and protein kinase activation ex vivo by Western blots of extracts of paw skin. KEY RESULTS: Intraplantar (i.pl.) injection of PGE(2) into the mouse paw caused nociceptive behaviour of short duration with mean ED(50) of 1.43 nmol. PGE(2) produced a longer-lasting mechanical allodynia, with an ED(50) of 0.05 nmol. Intraplantar injection of antagonists at EP(3) or EP(4), but not at EP(1) or EP(2) receptors inhibited PGE(2)-induced paw-licking. Paw-licking caused by PGE(2) was blocked by an inhibitor of PKA but only partially decreased by inhibition of the extracellular-regulated kinase (ERK). Selective inhibitors of PKC, c-Jun N-terminal kinase (JNK) or p38, all failed to affect PGE(2)-induced paw-licking. An EP(3) antagonist inhibited PGE(2)-induced mechanical allodynia. However, inhibitors of PKA, PKC or ERK, but not p38 or JNK, also partially inhibited PGE(2)-induced mechanical allodynia. Western blot analyses confirmed that i.pl. injection of PGE(2) activated PKA, PKCalpha, and mitogen activated kinases (MAPKs) in the paw. Co-treatment with EP(3) or EP(4) receptor antagonists reduced PGE(2)-induced PKA and ERK, but not PKCalpha activation. CONCLUSIONS AND IMPLICATIONS: The present results indicate that the nociceptive behaviour and mechanical allodynia caused by i.pl. PGE(2) are mediated through activation of distinct EP receptors and PK-dependent mechanisms.