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The absence of the leukotriene B4 receptor BLT1 attenuates peripheral inflammation and spinal nociceptive processing following intraplantar formalin injury.

Asahara M, Ito N, Yokomizo T, Nakamura M, Shimizu T, Yamada Y - Mol Pain (2015)

Bottom Line: Leukotriene B4 (LTB4) is a potent lipid mediator of inflammation, and its biological effects are mediated primarily through the high affinity LTB4 receptor BLT1.Edema formation and neutrophil infiltration in the paw were significantly decreased in BLT1 knockout mice compared with wild-type mice.Our data suggest that LTB4-BLT1 axis contributes not only to the peripheral inflammation but also to the neuronal activation in the spinal cord induced by intraplantar formalin injections.

View Article: PubMed Central - PubMed

Affiliation: Department of Anesthesiology, Faculty of Medicine, The University of Tokyo, Tokyo, Japan. asaharam-ane@h.u-tokyo.ac.jp.

ABSTRACT

Background: Leukotriene B4 (LTB4) is a potent lipid mediator of inflammation, and its biological effects are mediated primarily through the high affinity LTB4 receptor BLT1. Although numerous studies have reported that LTB4-BLT1 signaling is involved in inflammatory diseases, the role of BLT1 signaling in pain remains undefined. To clarify the role of LTB4-BLT1 signaling in acute inflammatory pain induced by tissue injury, we performed pain behavioral analysis and assessment of local inflammation induced by peripheral formalin injections in BLT1 knockout mice. We examined the phosphorylation of cAMP response element-binding protein (CREB) in the spinal cord both in wild-type and BLT1 knockout mice because phosphorylation of CREB in spinal cord neurons is important for nociceptive sensitization following peripheral injury. We also examined the effect of a BLT1 antagonist on formalin-induced pain responses in mice.

Results: BLT1 knockout mice exhibited markedly attenuated nociceptive responses induced by intraplantar formalin injections. Edema formation and neutrophil infiltration in the paw were significantly decreased in BLT1 knockout mice compared with wild-type mice. Phosphorylation of CREB in the spinal cord after the intraplantar formalin injection was decreased in BLT1 knockout mice. In addition, mice pretreated with a BLT1 antagonist showed reduced nociception and attenuated CREB phosphorylation in the spinal cord after the formalin injection.

Conclusions: Our data suggest that LTB4-BLT1 axis contributes not only to the peripheral inflammation but also to the neuronal activation in the spinal cord induced by intraplantar formalin injections. Thus, LTB4-BLT1 signaling is a potential target for therapeutic intervention of acute and persistent pain induced by tissue injury.

No MeSH data available.


Related in: MedlinePlus

TRPV1 and CGRP expression profiles of nociceptive neurons are unaffected by BLT1 receptor knockout (BLT1KO). (A) Transient receptor potential vanilloid 1(TRPV1)- and (B) calcitonin-gene related peptide (CGRP)-positive neurons in the L4 dorsal root ganglion (DRG). (C) Representative immunofluorescence images show the localization of TRPV1 (green) and CGRP (red) in L4 DRG neurons (magnification, 200×). Scale bar represents 50 μm. (D, E) Representative immunofluorescence images showing the localization of TRPV1 (green), CGRP (green) and NeuN (red) in the lumbar spinal cord (magnification, 100×). Scale bars represent 100 μm. (F) Quantitative analysis of TRPV1-immunopositive density. β-Actin and TRPA1 mRNA levels in the DRG (G) and spinal cord (H) analyzed by quantitative RT-PCR. (n = 5–7, a Kruskal-Wallis with Dunn’s multiple comparison test ). Data was expressed as Ct value.
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Fig2: TRPV1 and CGRP expression profiles of nociceptive neurons are unaffected by BLT1 receptor knockout (BLT1KO). (A) Transient receptor potential vanilloid 1(TRPV1)- and (B) calcitonin-gene related peptide (CGRP)-positive neurons in the L4 dorsal root ganglion (DRG). (C) Representative immunofluorescence images show the localization of TRPV1 (green) and CGRP (red) in L4 DRG neurons (magnification, 200×). Scale bar represents 50 μm. (D, E) Representative immunofluorescence images showing the localization of TRPV1 (green), CGRP (green) and NeuN (red) in the lumbar spinal cord (magnification, 100×). Scale bars represent 100 μm. (F) Quantitative analysis of TRPV1-immunopositive density. β-Actin and TRPA1 mRNA levels in the DRG (G) and spinal cord (H) analyzed by quantitative RT-PCR. (n = 5–7, a Kruskal-Wallis with Dunn’s multiple comparison test ). Data was expressed as Ct value.

Mentions: To evaluate the difference in the properties of nociceptive neurons between BLT1WT and BLT1KO mice, we analyzed the expression of the transient receptor potential vanilloid 1 (TRPV1) receptor as a marker of noxious heat sensor and CGRP as a marker of peptidergic nociceptive neurons in naïve DRG neurons using immunohistochemistry. As shown in Figure 2AB and C no significant differences were observed in the percent of the TRPV1-positive neurons and CGRP-positive neurons between BLT1WT and BLT1KO mice (39.0 ± 4.8% vs. 38.8 ± 6.3% for TRPV1 and 43.2 ± 3.2% vs. 36.2 ± 3.2% for CGRP). In addition, immunoreactivity for TRPV1 and CGRP in the dorsal horn was observed in the superficial lamina for both WT and BLT1KO mice (Figure 2D and E). No significant differences were observed in integral densities of TRPV1 immunoreactivity between genotypes (Figure 2F). Because no good antibodies are currently available for transient receptor potential ankyrin 1 (TRPA1), we confirmed the expression of TRPA1 in the lumbar DRG and spinal cord using quantitative RT-PCR analysis. Since several reports indicated the function of spinal TRPA1 [24-27], we analyzed and compared the expression of TRPA1 in spinal cord. There were no significant differences between the genotypes (Figure 2G, and H). Ct values of spinal cord (30.4±0.4 in WT, 31.8±1.1 in BLT1KO) were higher than Ct values of DRG (23.7±0.5 in WT, 25.6±0.3 in BLT1 KO).Figure 2


The absence of the leukotriene B4 receptor BLT1 attenuates peripheral inflammation and spinal nociceptive processing following intraplantar formalin injury.

Asahara M, Ito N, Yokomizo T, Nakamura M, Shimizu T, Yamada Y - Mol Pain (2015)

TRPV1 and CGRP expression profiles of nociceptive neurons are unaffected by BLT1 receptor knockout (BLT1KO). (A) Transient receptor potential vanilloid 1(TRPV1)- and (B) calcitonin-gene related peptide (CGRP)-positive neurons in the L4 dorsal root ganglion (DRG). (C) Representative immunofluorescence images show the localization of TRPV1 (green) and CGRP (red) in L4 DRG neurons (magnification, 200×). Scale bar represents 50 μm. (D, E) Representative immunofluorescence images showing the localization of TRPV1 (green), CGRP (green) and NeuN (red) in the lumbar spinal cord (magnification, 100×). Scale bars represent 100 μm. (F) Quantitative analysis of TRPV1-immunopositive density. β-Actin and TRPA1 mRNA levels in the DRG (G) and spinal cord (H) analyzed by quantitative RT-PCR. (n = 5–7, a Kruskal-Wallis with Dunn’s multiple comparison test ). Data was expressed as Ct value.
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Fig2: TRPV1 and CGRP expression profiles of nociceptive neurons are unaffected by BLT1 receptor knockout (BLT1KO). (A) Transient receptor potential vanilloid 1(TRPV1)- and (B) calcitonin-gene related peptide (CGRP)-positive neurons in the L4 dorsal root ganglion (DRG). (C) Representative immunofluorescence images show the localization of TRPV1 (green) and CGRP (red) in L4 DRG neurons (magnification, 200×). Scale bar represents 50 μm. (D, E) Representative immunofluorescence images showing the localization of TRPV1 (green), CGRP (green) and NeuN (red) in the lumbar spinal cord (magnification, 100×). Scale bars represent 100 μm. (F) Quantitative analysis of TRPV1-immunopositive density. β-Actin and TRPA1 mRNA levels in the DRG (G) and spinal cord (H) analyzed by quantitative RT-PCR. (n = 5–7, a Kruskal-Wallis with Dunn’s multiple comparison test ). Data was expressed as Ct value.
Mentions: To evaluate the difference in the properties of nociceptive neurons between BLT1WT and BLT1KO mice, we analyzed the expression of the transient receptor potential vanilloid 1 (TRPV1) receptor as a marker of noxious heat sensor and CGRP as a marker of peptidergic nociceptive neurons in naïve DRG neurons using immunohistochemistry. As shown in Figure 2AB and C no significant differences were observed in the percent of the TRPV1-positive neurons and CGRP-positive neurons between BLT1WT and BLT1KO mice (39.0 ± 4.8% vs. 38.8 ± 6.3% for TRPV1 and 43.2 ± 3.2% vs. 36.2 ± 3.2% for CGRP). In addition, immunoreactivity for TRPV1 and CGRP in the dorsal horn was observed in the superficial lamina for both WT and BLT1KO mice (Figure 2D and E). No significant differences were observed in integral densities of TRPV1 immunoreactivity between genotypes (Figure 2F). Because no good antibodies are currently available for transient receptor potential ankyrin 1 (TRPA1), we confirmed the expression of TRPA1 in the lumbar DRG and spinal cord using quantitative RT-PCR analysis. Since several reports indicated the function of spinal TRPA1 [24-27], we analyzed and compared the expression of TRPA1 in spinal cord. There were no significant differences between the genotypes (Figure 2G, and H). Ct values of spinal cord (30.4±0.4 in WT, 31.8±1.1 in BLT1KO) were higher than Ct values of DRG (23.7±0.5 in WT, 25.6±0.3 in BLT1 KO).Figure 2

Bottom Line: Leukotriene B4 (LTB4) is a potent lipid mediator of inflammation, and its biological effects are mediated primarily through the high affinity LTB4 receptor BLT1.Edema formation and neutrophil infiltration in the paw were significantly decreased in BLT1 knockout mice compared with wild-type mice.Our data suggest that LTB4-BLT1 axis contributes not only to the peripheral inflammation but also to the neuronal activation in the spinal cord induced by intraplantar formalin injections.

View Article: PubMed Central - PubMed

Affiliation: Department of Anesthesiology, Faculty of Medicine, The University of Tokyo, Tokyo, Japan. asaharam-ane@h.u-tokyo.ac.jp.

ABSTRACT

Background: Leukotriene B4 (LTB4) is a potent lipid mediator of inflammation, and its biological effects are mediated primarily through the high affinity LTB4 receptor BLT1. Although numerous studies have reported that LTB4-BLT1 signaling is involved in inflammatory diseases, the role of BLT1 signaling in pain remains undefined. To clarify the role of LTB4-BLT1 signaling in acute inflammatory pain induced by tissue injury, we performed pain behavioral analysis and assessment of local inflammation induced by peripheral formalin injections in BLT1 knockout mice. We examined the phosphorylation of cAMP response element-binding protein (CREB) in the spinal cord both in wild-type and BLT1 knockout mice because phosphorylation of CREB in spinal cord neurons is important for nociceptive sensitization following peripheral injury. We also examined the effect of a BLT1 antagonist on formalin-induced pain responses in mice.

Results: BLT1 knockout mice exhibited markedly attenuated nociceptive responses induced by intraplantar formalin injections. Edema formation and neutrophil infiltration in the paw were significantly decreased in BLT1 knockout mice compared with wild-type mice. Phosphorylation of CREB in the spinal cord after the intraplantar formalin injection was decreased in BLT1 knockout mice. In addition, mice pretreated with a BLT1 antagonist showed reduced nociception and attenuated CREB phosphorylation in the spinal cord after the formalin injection.

Conclusions: Our data suggest that LTB4-BLT1 axis contributes not only to the peripheral inflammation but also to the neuronal activation in the spinal cord induced by intraplantar formalin injections. Thus, LTB4-BLT1 signaling is a potential target for therapeutic intervention of acute and persistent pain induced by tissue injury.

No MeSH data available.


Related in: MedlinePlus