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Prostaglandin metabolite induces inhibition of TRPA1 and channel-dependent nociception.

Weng Y, Batista-Schepman PA, Barabas ME, Harris EQ, Dinsmore TB, Kossyreva EA, Foshage AM, Wang MH, Schwab MJ, Wang VM, Stucky CL, Story GM - Mol Pain (2012)

Bottom Line: Ca2+ imaging studies of DRG neurons demonstrated that 15d-PGJ2 pre-exposure reduced the magnitude and number of neuronal responses to AITC, but not CAP.AITC responses were not reduced when neurons were pre-exposed to 15d-PGJ2 combined with HC-030031 (TRPA1 antagonist), demonstrating that inhibitory effects of 15d-PGJ2 depend on TRPA1 activation.Collectively, our studies support that TRPA1 agonists may be useful as pain therapeutics.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Anesthesiology, Washington University Pain Center, St. Louis, MO 63110, USA.

ABSTRACT

Background: The Transient Receptor Potential (TRP) ion channel TRPA1 is a key player in pain pathways. Irritant chemicals activate ion channel TRPA1 via covalent modification of N-terminal cysteines. We and others have shown that 15-Deoxy-Δ12, 14-prostaglandin J2 (15d-PGJ2) similarly activates TRPA1 and causes channel-dependent nociception. Paradoxically, 15d-PGJ2 can also be anti-nociceptive in several pain models. Here we hypothesized that activation and subsequent desensitization of TRPA1 in dorsal root ganglion (DRG) neurons underlies the anti-nociceptive property of 15d-PGJ2. To investigate this, we utilized a battery of behavioral assays and intracellular Ca2+ imaging in DRG neurons to test if pre-treatment with 15d-PGJ2 inhibited TRPA1 to subsequent stimulation.

Results: Intraplantar pre-injection of 15d-PGJ2, in contrast to mustard oil (AITC), attenuated acute nocifensive responses to subsequent injections of 15d-PGJ2 and AITC, but not capsaicin (CAP). Intraplantar 15d-PGJ2-administered after the induction of inflammation-reduced mechanical hypersensitivity in the Complete Freund's Adjuvant (CFA) model for up to 2 h post-injection. The 15d-PGJ2-mediated reduction in mechanical hypersensitivity is dependent on TRPA1, as this effect was absent in TRPA1 knockout mice. Ca2+ imaging studies of DRG neurons demonstrated that 15d-PGJ2 pre-exposure reduced the magnitude and number of neuronal responses to AITC, but not CAP. AITC responses were not reduced when neurons were pre-exposed to 15d-PGJ2 combined with HC-030031 (TRPA1 antagonist), demonstrating that inhibitory effects of 15d-PGJ2 depend on TRPA1 activation. Single daily doses of 15d-PGJ2, administered during the course of 4 days in the CFA model, effectively reversed mechanical hypersensitivity without apparent tolerance or toxicity.

Conclusions: Taken together, our data support the hypothesis that 15d-PGJ2 induces activation followed by persistent inhibition of TRPA1 channels in DRG sensory neurons in vitro and in vivo. Moreover, we demonstrate novel evidence that 15d-PGJ2 is analgesic in mouse models of pain via a TRPA1-dependent mechanism. Collectively, our studies support that TRPA1 agonists may be useful as pain therapeutics.

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TRPA1-dependent 15d-PGJ2-mediated inhibition of AITC in vitro and mechanical hypersensivity in vivo. (A) In the presence of a specific TRPA1 antagonist, HC-030031 (100 μM), 100 μM 15d-PGJ2 did not block 50 μM AITC responses. (B) We also examined the reversibility of AITC-response inhibition in DRG neurons by 15d-PGJ2. 100 μM 15d-PGJ2 inhibited subsequent responses to 50 μM AITC for up to 16 min with continuous washout. Traces depict representative cells from 1 or more experiments in which >100 neurons were analyzed per experimental condition. (C) 15d-PGJ2 (10 μL ipl.) attenuates CFA-induced mechanical hypersensitivity in WT but not TRPA1 −/− mice. Vehicle or 15 mM 15d-PGJ2 was injected into the hindpaw of TRPA1 or WT littermates (n = 8 per group) 1 h prior to von Frey measurements. **p < 0.01 in comparing WT mice injected with 15d-PGJ2 vs. WT mice injected with vehicle. #p < 0.05, ##p < 0.01 in comparing WT mice injected with 15d-PGJ2 vs. TRPA1 KO mice injected with 15d-PGJ2. Values expressed as mean ± SEM. Data were analyzed using RMANOVA with Bonferonni post-hoc comparisons.
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Figure 7: TRPA1-dependent 15d-PGJ2-mediated inhibition of AITC in vitro and mechanical hypersensivity in vivo. (A) In the presence of a specific TRPA1 antagonist, HC-030031 (100 μM), 100 μM 15d-PGJ2 did not block 50 μM AITC responses. (B) We also examined the reversibility of AITC-response inhibition in DRG neurons by 15d-PGJ2. 100 μM 15d-PGJ2 inhibited subsequent responses to 50 μM AITC for up to 16 min with continuous washout. Traces depict representative cells from 1 or more experiments in which >100 neurons were analyzed per experimental condition. (C) 15d-PGJ2 (10 μL ipl.) attenuates CFA-induced mechanical hypersensitivity in WT but not TRPA1 −/− mice. Vehicle or 15 mM 15d-PGJ2 was injected into the hindpaw of TRPA1 or WT littermates (n = 8 per group) 1 h prior to von Frey measurements. **p < 0.01 in comparing WT mice injected with 15d-PGJ2 vs. WT mice injected with vehicle. #p < 0.05, ##p < 0.01 in comparing WT mice injected with 15d-PGJ2 vs. TRPA1 KO mice injected with 15d-PGJ2. Values expressed as mean ± SEM. Data were analyzed using RMANOVA with Bonferonni post-hoc comparisons.

Mentions: To address the possibility that 15d-PGJ2 inhibits DRG neuron responses to AITC via a mechanism separate from TRPA1 activation, we performed co-application studies in DRG using HC-030031, a selective TRPA1 antagonist. After a 4 min exposure to 100 μM 15d-PGJ2 in the presence of antagonist, 50 μM AITC responses of DRG neurons were equivalent to those after vehicle exposure (Figure 7A and data not shown). These data support that the inhibitory effect of 15d-PGJ2 pre-application on DRG neuron responses to AITC is mediated via TRPA1. We also examined the reversibility of desensitization of TRPA1 in DRG neurons by 15d-PGJ2. As shown in Figure 7B, 100 μM 15d-PGJ2 inhibited subsequent responses to 50 μM AITC for up to 16 min (4–16 min intervals tested) with continuous washout, while 50 mM KCl responses were unaltered, indicating that general neuronal viability remained intact.


Prostaglandin metabolite induces inhibition of TRPA1 and channel-dependent nociception.

Weng Y, Batista-Schepman PA, Barabas ME, Harris EQ, Dinsmore TB, Kossyreva EA, Foshage AM, Wang MH, Schwab MJ, Wang VM, Stucky CL, Story GM - Mol Pain (2012)

TRPA1-dependent 15d-PGJ2-mediated inhibition of AITC in vitro and mechanical hypersensivity in vivo. (A) In the presence of a specific TRPA1 antagonist, HC-030031 (100 μM), 100 μM 15d-PGJ2 did not block 50 μM AITC responses. (B) We also examined the reversibility of AITC-response inhibition in DRG neurons by 15d-PGJ2. 100 μM 15d-PGJ2 inhibited subsequent responses to 50 μM AITC for up to 16 min with continuous washout. Traces depict representative cells from 1 or more experiments in which >100 neurons were analyzed per experimental condition. (C) 15d-PGJ2 (10 μL ipl.) attenuates CFA-induced mechanical hypersensitivity in WT but not TRPA1 −/− mice. Vehicle or 15 mM 15d-PGJ2 was injected into the hindpaw of TRPA1 or WT littermates (n = 8 per group) 1 h prior to von Frey measurements. **p < 0.01 in comparing WT mice injected with 15d-PGJ2 vs. WT mice injected with vehicle. #p < 0.05, ##p < 0.01 in comparing WT mice injected with 15d-PGJ2 vs. TRPA1 KO mice injected with 15d-PGJ2. Values expressed as mean ± SEM. Data were analyzed using RMANOVA with Bonferonni post-hoc comparisons.
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Figure 7: TRPA1-dependent 15d-PGJ2-mediated inhibition of AITC in vitro and mechanical hypersensivity in vivo. (A) In the presence of a specific TRPA1 antagonist, HC-030031 (100 μM), 100 μM 15d-PGJ2 did not block 50 μM AITC responses. (B) We also examined the reversibility of AITC-response inhibition in DRG neurons by 15d-PGJ2. 100 μM 15d-PGJ2 inhibited subsequent responses to 50 μM AITC for up to 16 min with continuous washout. Traces depict representative cells from 1 or more experiments in which >100 neurons were analyzed per experimental condition. (C) 15d-PGJ2 (10 μL ipl.) attenuates CFA-induced mechanical hypersensitivity in WT but not TRPA1 −/− mice. Vehicle or 15 mM 15d-PGJ2 was injected into the hindpaw of TRPA1 or WT littermates (n = 8 per group) 1 h prior to von Frey measurements. **p < 0.01 in comparing WT mice injected with 15d-PGJ2 vs. WT mice injected with vehicle. #p < 0.05, ##p < 0.01 in comparing WT mice injected with 15d-PGJ2 vs. TRPA1 KO mice injected with 15d-PGJ2. Values expressed as mean ± SEM. Data were analyzed using RMANOVA with Bonferonni post-hoc comparisons.
Mentions: To address the possibility that 15d-PGJ2 inhibits DRG neuron responses to AITC via a mechanism separate from TRPA1 activation, we performed co-application studies in DRG using HC-030031, a selective TRPA1 antagonist. After a 4 min exposure to 100 μM 15d-PGJ2 in the presence of antagonist, 50 μM AITC responses of DRG neurons were equivalent to those after vehicle exposure (Figure 7A and data not shown). These data support that the inhibitory effect of 15d-PGJ2 pre-application on DRG neuron responses to AITC is mediated via TRPA1. We also examined the reversibility of desensitization of TRPA1 in DRG neurons by 15d-PGJ2. As shown in Figure 7B, 100 μM 15d-PGJ2 inhibited subsequent responses to 50 μM AITC for up to 16 min (4–16 min intervals tested) with continuous washout, while 50 mM KCl responses were unaltered, indicating that general neuronal viability remained intact.

Bottom Line: Ca2+ imaging studies of DRG neurons demonstrated that 15d-PGJ2 pre-exposure reduced the magnitude and number of neuronal responses to AITC, but not CAP.AITC responses were not reduced when neurons were pre-exposed to 15d-PGJ2 combined with HC-030031 (TRPA1 antagonist), demonstrating that inhibitory effects of 15d-PGJ2 depend on TRPA1 activation.Collectively, our studies support that TRPA1 agonists may be useful as pain therapeutics.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Anesthesiology, Washington University Pain Center, St. Louis, MO 63110, USA.

ABSTRACT

Background: The Transient Receptor Potential (TRP) ion channel TRPA1 is a key player in pain pathways. Irritant chemicals activate ion channel TRPA1 via covalent modification of N-terminal cysteines. We and others have shown that 15-Deoxy-Δ12, 14-prostaglandin J2 (15d-PGJ2) similarly activates TRPA1 and causes channel-dependent nociception. Paradoxically, 15d-PGJ2 can also be anti-nociceptive in several pain models. Here we hypothesized that activation and subsequent desensitization of TRPA1 in dorsal root ganglion (DRG) neurons underlies the anti-nociceptive property of 15d-PGJ2. To investigate this, we utilized a battery of behavioral assays and intracellular Ca2+ imaging in DRG neurons to test if pre-treatment with 15d-PGJ2 inhibited TRPA1 to subsequent stimulation.

Results: Intraplantar pre-injection of 15d-PGJ2, in contrast to mustard oil (AITC), attenuated acute nocifensive responses to subsequent injections of 15d-PGJ2 and AITC, but not capsaicin (CAP). Intraplantar 15d-PGJ2-administered after the induction of inflammation-reduced mechanical hypersensitivity in the Complete Freund's Adjuvant (CFA) model for up to 2 h post-injection. The 15d-PGJ2-mediated reduction in mechanical hypersensitivity is dependent on TRPA1, as this effect was absent in TRPA1 knockout mice. Ca2+ imaging studies of DRG neurons demonstrated that 15d-PGJ2 pre-exposure reduced the magnitude and number of neuronal responses to AITC, but not CAP. AITC responses were not reduced when neurons were pre-exposed to 15d-PGJ2 combined with HC-030031 (TRPA1 antagonist), demonstrating that inhibitory effects of 15d-PGJ2 depend on TRPA1 activation. Single daily doses of 15d-PGJ2, administered during the course of 4 days in the CFA model, effectively reversed mechanical hypersensitivity without apparent tolerance or toxicity.

Conclusions: Taken together, our data support the hypothesis that 15d-PGJ2 induces activation followed by persistent inhibition of TRPA1 channels in DRG sensory neurons in vitro and in vivo. Moreover, we demonstrate novel evidence that 15d-PGJ2 is analgesic in mouse models of pain via a TRPA1-dependent mechanism. Collectively, our studies support that TRPA1 agonists may be useful as pain therapeutics.

Show MeSH
Related in: MedlinePlus