Intracellular alkalization causes pain sensation through activation of TRPA1 in mice
Fumitaka Fujita, … , Takaaki Sokabe, Makoto Tominaga
Fumitaka Fujita, … , Takaaki Sokabe, Makoto Tominaga
Published November 13, 2008
Citation Information: J Clin Invest. 2008;118(12):4049-4057. https://doi.org/10.1172/JCI35957.
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Research Article Neuroscience

Intracellular alkalization causes pain sensation through activation of TRPA1 in mice

Abstract

Vertebrate cells require a very narrow pH range for survival. Cells accordingly possess sensory and defense mechanisms for situations where the pH deviates from the viable range. Although the monitoring of acidic pH by sensory neurons has been attributed to several ion channels, including transient receptor potential vanilloid 1 channel (TRPV1) and acid-sensing ion channels (ASICs), the mechanisms by which these cells detect alkaline pH are not well understood. Here, using Ca2+ imaging and patch-clamp recording, we showed that alkaline pH activated transient receptor potential cation channel, subfamily A, member 1 (TRPA1) and that activation of this ion channel was involved in nociception. In addition, intracellular alkalization activated TRPA1 at the whole-cell level, and single-channel openings were observed in the inside-out configuration, indicating that alkaline pH activated TRPA1 from the inside. Analyses of mutants suggested that the two N-terminal cysteine residues in TRPA1 were involved in activation by intracellular alkalization. Furthermore, intraplantar injection of ammonium chloride into the mouse hind paw caused pain-related behaviors that were not observed in TRPA1-deficient mice. These results suggest that alkaline pH causes pain sensation through activation of TRPA1 and may provide a molecular explanation for some of the human alkaline pH–related sensory disorders whose mechanisms are largely unknown.

Authors

Fumitaka Fujita, Kunitoshi Uchida, Tomoko Moriyama, Asako Shima, Koji Shibasaki, Hitoshi Inada, Takaaki Sokabe, Makoto Tominaga

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Figure 2

Alkaline pH solution and NH4Cl activate TRPA1 in HEK293 cells.

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Alkaline pH solution and NH4Cl activate TRPA1 in HEK293 cells. (A) A...
(A) A representative whole-cell current trace activated by pH 8 pipette solution or AITC (20 μM) in the presence of extracellular Ca2+ in HEK293 cells expressing TRPA1. The insets indicate representative I-V curves of pH 8 pipette solution– or AITC-activated currents showing an outward rectification. Holding potential (Vh), –60 mV. Horizontal bars indicate the duration of compound application. (B) A representative current trace activated by pH 9 Tris buffer solution or AITC (20 μM) using the unbuffered pipette solution in the presence of extracellular Ca2+. The inset indicates a representative I-V curve of pH 9 Tris buffer solution–activated current showing an outward rectification. (C) Comparison of current densities activated by pH 9 Tris buffer solution using buffered (n = 8) or unbuffered (n = 8) pipette solution. *P < 0.05. (D) A representative current trace activated by 30 mM NH4Cl or AITC (20 μM) in the presence of extracellular Ca2+. The inset indicates a representative I-V curve of NH4Cl-activated current showing an outward rectification. (E) Comparison of times to maximum current responses evoked by pH 8 pipette solution (n = 4), pH 9 Tris buffer solution (n = 8), or 30 mM NH4Cl (n = 31) using unbuffered pipette solution. **P < 0.01.