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A role for Piezo2 in EPAC1-dependent mechanical allodynia.

Eijkelkamp N, Linley JE, Torres JM, Bee L, Dickenson AH, Gringhuis M, Minett MS, Hong GS, Lee E, Oh U, Ishikawa Y, Zwartkuis FJ, Cox JJ, Wood JN - Nat Commun (2013)

Bottom Line: Human Piezo2 produces large mechanically gated currents that are enhanced by the activation of the cAMP-sensor Epac1 or cytosolic calcium but are unaffected by protein kinase C or protein kinase A and depend on the integrity of the cytoskeleton.Piezo2 knockdown also enhanced thresholds for light touch.Finally, 8-pCPT sensitizes responses to innocuous mechanical stimuli without changing the electrical excitability of sensory fibres.

View Article: PubMed Central - PubMed

Affiliation: Molecular Nociception Group, Wolfson Institute for Biomedical Research, University College London, London WC1E 6BT, UK. N.Eijkelkamp@umcutrecht.nl

ABSTRACT
Aberrant mechanosensation has an important role in different pain states. Here we show that Epac1 (cyclic AMP sensor) potentiation of Piezo2-mediated mechanotransduction contributes to mechanical allodynia. Dorsal root ganglia Epac1 mRNA levels increase during neuropathic pain, and nerve damage-induced allodynia is reduced in Epac1-/- mice. The Epac-selective cAMP analogue 8-pCPT sensitizes mechanically evoked currents in sensory neurons. Human Piezo2 produces large mechanically gated currents that are enhanced by the activation of the cAMP-sensor Epac1 or cytosolic calcium but are unaffected by protein kinase C or protein kinase A and depend on the integrity of the cytoskeleton. In vivo, 8-pCPT induces long-lasting allodynia that is prevented by the knockdown of Epac1 and attenuated by mouse Piezo2 knockdown. Piezo2 knockdown also enhanced thresholds for light touch. Finally, 8-pCPT sensitizes responses to innocuous mechanical stimuli without changing the electrical excitability of sensory fibres. These data indicate that the Epac1-Piezo2 axis has a role in the development of mechanical allodynia during neuropathic pain.

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DRG Epac1 expression is increased in and required for L5 spinal nerve transaction-induced allodynia.(a) The sensitivity to mechanical stimulation was determined in sham-operated mice (n=5), naive controls (n=10) or mice subjected to unilateral L5 SNT 4 weeks after surgery (n=10). Line with error bars represent mean±s.e.m. (b) Epac1 or (c) Epac2 mRNA levels in DRGs innervating the contralateral (contra) or ipsilateral (ipsi) side of sham-operated, naive controls and L5 SNL mice 4 weeks after surgery. Epac1/2 mRNA expression levels were corrected for GAPDH and β-actin mRNA expression levels. (d) Epac1 protein expression levels in ipsilateral (affected) and contralateral (unaffected) DRGs of mice subjected to unilateral L5 SNT 4 weeks after surgery. β-Actin and the neuron-specific β3-tubulin was used as loading control. (e) Epac1 protein expression in DRGs of WT (n=6), Epac1+/− (HE, n=4) and Epac1−/− (HO, n=4) mice. (f) The sensitivity to mechanical stimulation was determined in wild-type (n=10), Epac1+/− (n=10) and Epac1−/− (n=13) mice subjected to unilateral L5 SNT. Repeated measures one-way analysis of variance (ANOVA) showed a significant genotype effect F(2,30)=25,935, P<0.001. Bonferoni post hoc analysis showed a significant effect between Epac−/− and WT (P<0.001); Epac1−/− and Epac1+/− (P<0.001); and Epac1+/− and WT (P<0.05). All data are expressed as mean±s.e.m. (a–c) Data are analysed by ANOVA followed by the Bonferroni post hoc test. (d) Data are analysed using t-test. *P<0.05, **P<0.01, ***P<00.1.
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f1: DRG Epac1 expression is increased in and required for L5 spinal nerve transaction-induced allodynia.(a) The sensitivity to mechanical stimulation was determined in sham-operated mice (n=5), naive controls (n=10) or mice subjected to unilateral L5 SNT 4 weeks after surgery (n=10). Line with error bars represent mean±s.e.m. (b) Epac1 or (c) Epac2 mRNA levels in DRGs innervating the contralateral (contra) or ipsilateral (ipsi) side of sham-operated, naive controls and L5 SNL mice 4 weeks after surgery. Epac1/2 mRNA expression levels were corrected for GAPDH and β-actin mRNA expression levels. (d) Epac1 protein expression levels in ipsilateral (affected) and contralateral (unaffected) DRGs of mice subjected to unilateral L5 SNT 4 weeks after surgery. β-Actin and the neuron-specific β3-tubulin was used as loading control. (e) Epac1 protein expression in DRGs of WT (n=6), Epac1+/− (HE, n=4) and Epac1−/− (HO, n=4) mice. (f) The sensitivity to mechanical stimulation was determined in wild-type (n=10), Epac1+/− (n=10) and Epac1−/− (n=13) mice subjected to unilateral L5 SNT. Repeated measures one-way analysis of variance (ANOVA) showed a significant genotype effect F(2,30)=25,935, P<0.001. Bonferoni post hoc analysis showed a significant effect between Epac−/− and WT (P<0.001); Epac1−/− and Epac1+/− (P<0.001); and Epac1+/− and WT (P<0.05). All data are expressed as mean±s.e.m. (a–c) Data are analysed by ANOVA followed by the Bonferroni post hoc test. (d) Data are analysed using t-test. *P<0.05, **P<0.01, ***P<00.1.

Mentions: We measured Epac1 and Epac2 mRNA expression in a neuropathic pain model. Four weeks after a unilateral L5 nerve transection (L5 SNT), mice displayed mechanical allodynia in the ipsilateral paw. Thresholds to mechanical stimulation were unaffected in contralateral paws, sham-operated mice or in naive untreated mice (Fig. 1a). At this time point, Epac1 mRNA levels in dorsal root ganglia (DRG) innervating the ipsilateral paw increased in comparison with DRG innervating the contralateral paw, from sham-operated animals, or from untreated mice by ~1.8-fold (Fig. 1b). Epac2 mRNA expression levels in DRG innervating the ipsilateral or contralateral paw were similar to Epac2 expression levels in DRG from sham-operated animals or naive untreated animals (Fig. 1c). The increase in Epac1 mRNA levels in the DRG innervating the ipsilateral paw was also observed at the protein level (Fig. 1d).


A role for Piezo2 in EPAC1-dependent mechanical allodynia.

Eijkelkamp N, Linley JE, Torres JM, Bee L, Dickenson AH, Gringhuis M, Minett MS, Hong GS, Lee E, Oh U, Ishikawa Y, Zwartkuis FJ, Cox JJ, Wood JN - Nat Commun (2013)

DRG Epac1 expression is increased in and required for L5 spinal nerve transaction-induced allodynia.(a) The sensitivity to mechanical stimulation was determined in sham-operated mice (n=5), naive controls (n=10) or mice subjected to unilateral L5 SNT 4 weeks after surgery (n=10). Line with error bars represent mean±s.e.m. (b) Epac1 or (c) Epac2 mRNA levels in DRGs innervating the contralateral (contra) or ipsilateral (ipsi) side of sham-operated, naive controls and L5 SNL mice 4 weeks after surgery. Epac1/2 mRNA expression levels were corrected for GAPDH and β-actin mRNA expression levels. (d) Epac1 protein expression levels in ipsilateral (affected) and contralateral (unaffected) DRGs of mice subjected to unilateral L5 SNT 4 weeks after surgery. β-Actin and the neuron-specific β3-tubulin was used as loading control. (e) Epac1 protein expression in DRGs of WT (n=6), Epac1+/− (HE, n=4) and Epac1−/− (HO, n=4) mice. (f) The sensitivity to mechanical stimulation was determined in wild-type (n=10), Epac1+/− (n=10) and Epac1−/− (n=13) mice subjected to unilateral L5 SNT. Repeated measures one-way analysis of variance (ANOVA) showed a significant genotype effect F(2,30)=25,935, P<0.001. Bonferoni post hoc analysis showed a significant effect between Epac−/− and WT (P<0.001); Epac1−/− and Epac1+/− (P<0.001); and Epac1+/− and WT (P<0.05). All data are expressed as mean±s.e.m. (a–c) Data are analysed by ANOVA followed by the Bonferroni post hoc test. (d) Data are analysed using t-test. *P<0.05, **P<0.01, ***P<00.1.
© Copyright Policy - open-access
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC3644070&req=5

f1: DRG Epac1 expression is increased in and required for L5 spinal nerve transaction-induced allodynia.(a) The sensitivity to mechanical stimulation was determined in sham-operated mice (n=5), naive controls (n=10) or mice subjected to unilateral L5 SNT 4 weeks after surgery (n=10). Line with error bars represent mean±s.e.m. (b) Epac1 or (c) Epac2 mRNA levels in DRGs innervating the contralateral (contra) or ipsilateral (ipsi) side of sham-operated, naive controls and L5 SNL mice 4 weeks after surgery. Epac1/2 mRNA expression levels were corrected for GAPDH and β-actin mRNA expression levels. (d) Epac1 protein expression levels in ipsilateral (affected) and contralateral (unaffected) DRGs of mice subjected to unilateral L5 SNT 4 weeks after surgery. β-Actin and the neuron-specific β3-tubulin was used as loading control. (e) Epac1 protein expression in DRGs of WT (n=6), Epac1+/− (HE, n=4) and Epac1−/− (HO, n=4) mice. (f) The sensitivity to mechanical stimulation was determined in wild-type (n=10), Epac1+/− (n=10) and Epac1−/− (n=13) mice subjected to unilateral L5 SNT. Repeated measures one-way analysis of variance (ANOVA) showed a significant genotype effect F(2,30)=25,935, P<0.001. Bonferoni post hoc analysis showed a significant effect between Epac−/− and WT (P<0.001); Epac1−/− and Epac1+/− (P<0.001); and Epac1+/− and WT (P<0.05). All data are expressed as mean±s.e.m. (a–c) Data are analysed by ANOVA followed by the Bonferroni post hoc test. (d) Data are analysed using t-test. *P<0.05, **P<0.01, ***P<00.1.
Mentions: We measured Epac1 and Epac2 mRNA expression in a neuropathic pain model. Four weeks after a unilateral L5 nerve transection (L5 SNT), mice displayed mechanical allodynia in the ipsilateral paw. Thresholds to mechanical stimulation were unaffected in contralateral paws, sham-operated mice or in naive untreated mice (Fig. 1a). At this time point, Epac1 mRNA levels in dorsal root ganglia (DRG) innervating the ipsilateral paw increased in comparison with DRG innervating the contralateral paw, from sham-operated animals, or from untreated mice by ~1.8-fold (Fig. 1b). Epac2 mRNA expression levels in DRG innervating the ipsilateral or contralateral paw were similar to Epac2 expression levels in DRG from sham-operated animals or naive untreated animals (Fig. 1c). The increase in Epac1 mRNA levels in the DRG innervating the ipsilateral paw was also observed at the protein level (Fig. 1d).

Bottom Line: Human Piezo2 produces large mechanically gated currents that are enhanced by the activation of the cAMP-sensor Epac1 or cytosolic calcium but are unaffected by protein kinase C or protein kinase A and depend on the integrity of the cytoskeleton.Piezo2 knockdown also enhanced thresholds for light touch.Finally, 8-pCPT sensitizes responses to innocuous mechanical stimuli without changing the electrical excitability of sensory fibres.

View Article: PubMed Central - PubMed

Affiliation: Molecular Nociception Group, Wolfson Institute for Biomedical Research, University College London, London WC1E 6BT, UK. N.Eijkelkamp@umcutrecht.nl

ABSTRACT
Aberrant mechanosensation has an important role in different pain states. Here we show that Epac1 (cyclic AMP sensor) potentiation of Piezo2-mediated mechanotransduction contributes to mechanical allodynia. Dorsal root ganglia Epac1 mRNA levels increase during neuropathic pain, and nerve damage-induced allodynia is reduced in Epac1-/- mice. The Epac-selective cAMP analogue 8-pCPT sensitizes mechanically evoked currents in sensory neurons. Human Piezo2 produces large mechanically gated currents that are enhanced by the activation of the cAMP-sensor Epac1 or cytosolic calcium but are unaffected by protein kinase C or protein kinase A and depend on the integrity of the cytoskeleton. In vivo, 8-pCPT induces long-lasting allodynia that is prevented by the knockdown of Epac1 and attenuated by mouse Piezo2 knockdown. Piezo2 knockdown also enhanced thresholds for light touch. Finally, 8-pCPT sensitizes responses to innocuous mechanical stimuli without changing the electrical excitability of sensory fibres. These data indicate that the Epac1-Piezo2 axis has a role in the development of mechanical allodynia during neuropathic pain.

Show MeSH
Related in: MedlinePlus