Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The .gov means it’s official.
Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site.

Https

The site is secure.
The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely.

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2012 Aug 8:9:189.
doi: 10.1186/1742-2094-9-189.

CC chemokine ligand 2 upregulates the current density and expression of TRPV1 channels and Nav1.8 sodium channels in dorsal root ganglion neurons

Der-Jang Kao  1 , Allen H Li, Jin-Chung Chen, Ro-Sun Luo, Ying-Ling Chen, Juu-Chin Lu, Hung-Li Wang
Affiliations

CC chemokine ligand 2 upregulates the current density and expression of TRPV1 channels and Nav1.8 sodium channels in dorsal root ganglion neurons

Der-Jang Kao et al. J Neuroinflammation. .

Abstract

Background: Inflammation or nerve injury-induced upregulation and release of chemokine CC chemokine ligand 2 (CCL2) within the dorsal root ganglion (DRG) is believed to enhance the activity of DRG nociceptive neurons and cause hyperalgesia. Transient receptor potential vanilloid receptor 1 (TRPV1) and tetrodotoxin (TTX)-resistant Na(v)1.8 sodium channels play an essential role in regulating the excitability and pain transmission of DRG nociceptive neurons. We therefore tested the hypothesis that CCL2 causes peripheral sensitization of nociceptive DRG neurons by upregulating the function and expression of TRPV1 and Nav1.8 channels.

Methods: DRG neuronal culture was prepared from 3-week-old Sprague-Dawley rats and incubated with various concentrations of CCL2 for 24 to 36 hours. Whole-cell voltage-clamp recordings were performed to record TRPV1 agonist capsaicin-evoked inward currents or TTX-insensitive Na(+) currents from control or CCL2-treated small DRG sensory neurons. The CCL2 effect on the mRNA expression of TRPV1 or Na(v)1.8 was measured by real-time quantitative RT-PCR assay.

Results: Pretreatment of CCL2 for 24 to 36 hours dose-dependently (EC(50) value = 0.6 ± 0.05 nM) increased the density of capsaicin-induced currents in small putative DRG nociceptive neurons. TRPV1 mRNA expression was greatly upregulated in DRG neurons preincubated with 5 nM CCL2. Pretreating small DRG sensory neurons with CCL2 also increased the density of TTX-resistant Na(+) currents with a concentration-dependent manner (EC(50) value = 0.7 ± 0.06 nM). The Na(v)1.8 mRNA level was significantly increased in DRG neurons pretreated with CCL2. In contrast, CCL2 preincubation failed to affect the mRNA level of TTX-resistant Nav1.9. In the presence of the specific phosphatidylinositol-3 kinase (PI3K) inhibitor LY294002 or Akt inhibitor IV, CCL2 pretreatment failed to increase the current density of capsaicin-evoked inward currents or TTX-insensitive Na(+) currents and the mRNA level of TRPV1 or Na(v)1.8.

Conclusions: Our results showed that CCL2 increased the function and mRNA level of TRPV1 channels and Na(v)1.8 sodium channels in small DRG sensory neurons via activating the PI3K/Akt signaling pathway. These findings suggest that following tissue inflammation or peripheral nerve injury, upregulation and release of CCL2 within the DRG could facilitate pain transmission mediated by nociceptive DRG neurons and could induce hyperalgesia by upregulating the expression and function of TRPV1 and Na(v)1.8 channels in DRG nociceptive neurons.

PubMed Disclaimer

Figures

Figure 1
Figure 1
CCL2 pretreatment augments capsaicin-evoked inward cationic currents in small-diameter dorsal root ganglion neurons. (A) Compared with capsaicin (0.3 μM)-induced inward cationic current in a control small dorsal root ganglion (DRG) sensory neuron, the amplitude of capsaicin-evoked inward current was greatly increased in a small-diameter DRG neuron preincubated with 5 nM chemokine CC chemokine ligand 2 (CCL2) for 24 hours. Holding potential (VH) = −60 mV. (B) CCL2 pretreatment increased the density of capsaicin (0.3 μM)-induced inward currents with a concentration-dependent manner. Each point shows the mean ± standard error value of 10 neurons.
Figure 2
Figure 2
CCL2 increases density of capsaicin-evoked inward currents by upregulating TRPV1 mRNA expression in sensory neurons. (A) The concentration–response curve for the density of capsaicin-induced inward currents was obtained from small-diameter dorsal root ganglion (DRG) neurons in the absence and presence of chemokine CC chemokine ligand 2 (CCL2; 5 nM) pretreatment. Note that CCL2 preincubation increased the density of capsaicin currents without significantly affecting the EC50 value. Holding potential (VH) = −60 mV. Each point represents the mean ± standard error (SE) value of 12 neurons. (B) Quantitative RT-PCR assays showed that pretreating cultured DRG neurons with 5 nM CCL2 for 24 to 36 hours greatly increased the mRNA level of transient receptor potential vanilloid receptor 1 (TRPV1). Each bar shows the mean ± SE value of five experiments. *P <0.01.
Figure 3
Figure 3
CCL2 augments capsaicin-evoked inward currents and increases TRPV1 mRNA in neurons via activating phosphatidylinositol-3 kinase. (A) Following co-treating cultured dorsal root ganglion (DRG) neurons with chemokine CC chemokine ligand 2 (CCL2) (5 nM) and phosphatidylinositol-3 kinase (PI3K) inhibitor LY294002 (10 μM) for 24 to 36 hours, CCL2 failed to significantly enhance the amplitude of capsaicin (0.3 μM)-evoked inward current in a small-diameter DRG neuron. In the presence of ERK 1/2 inhibitor U0126 (20 μM), CCL2 still greatly augmented the magnitude of capsaicin currents in a small DRG sensory neuron. Holding potential (VH) = −60 mV. (B) Pretreating small-diameter DRG neurons with 5 nM CCL2 significantly increased the density of capsaicin-evoked inward currents. LY294002 (10 μM) almost completely blocked CCL2 enhancement of capsaicin currents, and U0126 (20 μM) failed to affect CCL2 potentiation of capsaicin currents. Each bar shows the mean ± standard error (SE) value of 10 to 13 neurons. (C) RT-PCR assays showed that CCL2 (5 nM) pretreatment significantly increased the transient receptor potential vanilloid receptor 1 (TRPV1) mRNA level of cultured DRG neurons. In the presence of 10 μM LY294002, CCL2 pretreatment of DRG sensory neurons for 24 to 36 hours failed to upregulate TRPV1 mRNA expression. Each bar represents the mean ± SE value of five experiments. *P <0.01 compared with control neurons.
Figure 4
Figure 4
CCL2 enhances capsaicin currents and upregulates TRPV1 mRNA expression in neurons via activating Akt. (A) In the presence of Akt inhibitor IV (1 μM), chemokine CC chemokine ligand 2 (CCL2) (5 nM) pretreatment did not augment the magnitude of capsaicin (0.3 μM)-evoked inward current. Holding potential (VH) = −60 mV. (B) Akt inhibitor IV significantly blocked CCL2 upregulation of density of capsaicin currents. Each bar shows the mean ± standard error (SE) value of 10 neurons. (C) CCL2 pretreatment of dorsal root ganglion (DRG) neurons for 24 to 36 hours failed to increase transient receptor potential vanilloid receptor 1 (TRPV1) mRNA level in the presence of 1 μM Akt inhibitor IV. Each bar represents the mean ± SE value of five experiments. *P <0.01 compared with control neurons.
Figure 5
Figure 5
CCL2 increases the density of tetrodotoxin-resistant sodium currents in small dorsal root ganglion sensory neurons. (A) In the presence of 0.5 μM tetrodotoxin (TTX), the holding potential (VH) of small dorsal root ganglion (DRG) sensory neurons was held at −80 mV, and depolarizing steps (50 milliseconds) were applied from −50 mV to 50 mV with an increment of 10 mV. The I–V (current–voltage) curve of TTX-insensitive Na+ currents was then obtained from control or chemokine CC chemokine ligand 2 (CCL2; 5 nM)-pretreated small-diameter DRG neurons. Each point represents the mean ± standard error (SE) value of 10 neurons. (B) Traces of TTX-resistant sodium currents were evoked from a VH of −80 mV to step potentials ranging from −20 mV to 20 mV. Compared with a control small DRG sensory neuron, the magnitude of TTX-insensitive Na+ currents was greatly increased in a CCL2-pretreated small-diameter DRG neuron. (C) CCL2 pretreatment increased the density of TTX-resistant sodium currents recorded at −10 mV in a dose-dependent manner. Each point shows the mean ± SE value of eight neurons. (D) Pretreating cultured DRG neurons with 5 nM CCL2 for 24 to 36 hours significantly upregulated mRNA expression of Nav1.8 without affecting the Nav1.9 mRNA level. Each bar represents the mean ± SE value of five experiments. *P <0.01 compared with control neurons.
Figure 6
Figure 6
CCL2 increases tetrodotoxin-resistant Na +current amplitude and Nav1.8 mRNA through activating the phosphatidylinositol-3 kinase/Akt pathway. (A) Tetrodotoxin (TTX)-insensitive sodium currents were evoked from a holding potential (VH) of −80 mV to step potentials ranging from −20 mV to 20 mV. In the presence of ERK 1/2 inhibitor U0126 (20 μM), chemokine CC chemokine ligand 2 (CCL2) preincubation greatly increased the magnitude of TTX-resistant Na+ currents in a small dorsal root ganglion (DRG) sensory neuron. Pretreating small-diameter DRG neurons with 5 nM CCL2 did not augment the amplitude of TTX-resistant sodium currents in the presence of phosphatidylinositol-3 kinase (PI3K) inhibitor LY294002 (10 μM) or Akt inhibitor IV (1 μM). (B) Pretreating small-diameter DRG neurons with 5 nM CCL2 increased the density of TTX-insensitive sodium Na+ currents recorded at −20 mV. PI3K inhibitor LY294002 or Akt inhibitor IV almost completely inhibited CCL2 enhancement of TTX-resistant Na+ currents. Each bar shows the mean ± standard error (SE) value of 10 neurons. (C) Pretreating DRG neurons with 5 nM CCL2 for 24 to 36 hours significantly increased the mRNA level of Nav1.8. PI3K inhibitor LY294002 or Akt inhibitor IV significantly inhibited CCL2 upregulation of Nav1.8 mRNA expression. Each bar represents the mean ± SE value of five experiments. *P <0.01 compared with control neurons.
See this image and copyright information in PMC

Similar articles

See all similar articles

Cited by

See all "Cited by" articles

References

    1. Abbadie C, Bhangoo S, De Koninck Y, Malcangio M, Melik-Parsadaniantz S, White FA. Chemokines and pain mechanisms. Brain Res Rev. 2009;60:125–135. doi: 10.1016/j.brainresrev.2008.12.002. - DOI - PMC - PubMed
    1. Miller RJ, Jung H, Bhangoo SK, White FA. Cytokine and chemokine regulation of sensory neuron function. Handb Exp Pharmacol. 2009;194:417–449. doi: 10.1007/978-3-540-79090-7_12. - DOI - PMC - PubMed
    1. Gao YJ, Ji RR. Chemokines, neuronal–glial interactions, and central processing of neuropathic pain. Pharmacol Ther. 2010;126:56–68. doi: 10.1016/j.pharmthera.2010.01.002. - DOI - PMC - PubMed
    1. Ren K, Dubner R. Interactions between the immune and nervous systems in pain. Nat Med. 2010;16:1267–1276. doi: 10.1038/nm.2234. - DOI - PMC - PubMed
    1. White FA, Wilson N. Chemokines as pain mediators and modulators. Curr Opin Anaesthesiol. 2008;21:580–585. doi: 10.1097/ACO.0b013e32830eb69d. - DOI - PMC - PubMed

Publication types

LinkOut - more resources