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A TRPA1-dependent mechanism for the pungent sensation of weak acids.

Wang YY, Chang RB, Allgood SD, Silver WL, Liman ER - J. Gen. Physiol. (2011)

Bottom Line: Our results show that heterologously expressed TRPA1 currents can be induced by a series of weak organic acids, including acetic, propionic, formic, and lactic acid, but not by strong acids.Importantly, responses of trigeminal neurons to weak acids were highly overrepresented in the subpopulation of TRPA1-expressing neurons and were severely reduced in neurons from TRPA1 knockout mice.We conclude that TRPA1 is a general sensor for weak acids that produce intracellular acidification and suggest that it functions within the pain pathway to mediate sensitivity to cellular acidosis.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Biological Sciences, Section of Neurobiology, University of Southern California, Los Angeles, CA 90089, USA.

ABSTRACT
Acetic acid produces an irritating sensation that can be attributed to activation of nociceptors within the trigeminal ganglion that innervate the nasal or oral cavities. These sensory neurons sense a diverse array of noxious agents in the environment, allowing animals to actively avoid tissue damage. Although receptor mechanisms have been identified for many noxious chemicals, the mechanisms by which animals detect weak acids, such as acetic acid, are less well understood. Weak acids are only partially dissociated at neutral pH and, as such, some can cross the cell membrane, acidifying the cell cytosol. The nociceptor ion channel TRPA1 is activated by CO(2), through gating of the channel by intracellular protons, making it a candidate to more generally mediate sensory responses to weak acids. To test this possibility, we measured responses to weak acids from heterologously expressed TRPA1 channels and trigeminal neurons with patch clamp recording and Ca(2+) microfluorometry. Our results show that heterologously expressed TRPA1 currents can be induced by a series of weak organic acids, including acetic, propionic, formic, and lactic acid, but not by strong acids. Notably, the degree of channel activation was predicted by the degree of intracellular acidification produced by each acid, suggesting that intracellular protons are the proximate stimulus that gates the channel. Responses to weak acids produced a Ca(2+)-independent inactivation that precluded further activation by weak acids or reactive chemicals, whereas preactivation by reactive electrophiles sensitized TRPA1 channels to weak acids. Importantly, responses of trigeminal neurons to weak acids were highly overrepresented in the subpopulation of TRPA1-expressing neurons and were severely reduced in neurons from TRPA1 knockout mice. We conclude that TRPA1 is a general sensor for weak acids that produce intracellular acidification and suggest that it functions within the pain pathway to mediate sensitivity to cellular acidosis.

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PA activates TRPA1 in Ca2+ imaging. Agonist-induced elevation of intracellular Ca2+ in HEK-293 cells transfected with TRPV1 or TRPA1. (A) TRPA1-expressing HEK cells responded to both 100 mM PA, pH 6.5, and 100 µM Cin. (B) TRPV1-expressing HEK cells responded only to 1 µM Cap and showed nonspecific responses to PA. Scatter plot shows the amplitude of the responses to PA as a function of the responses to Cin (A) or Cap (B). Average responses to PA and Cin (A) or PA and Cap (B) are shown in the bar graph. Data are represented by the mean ± SEM. Significance was determined with the two-tailed Student’s t test. ***, P < 0.001.
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fig8: PA activates TRPA1 in Ca2+ imaging. Agonist-induced elevation of intracellular Ca2+ in HEK-293 cells transfected with TRPV1 or TRPA1. (A) TRPA1-expressing HEK cells responded to both 100 mM PA, pH 6.5, and 100 µM Cin. (B) TRPV1-expressing HEK cells responded only to 1 µM Cap and showed nonspecific responses to PA. Scatter plot shows the amplitude of the responses to PA as a function of the responses to Cin (A) or Cap (B). Average responses to PA and Cin (A) or PA and Cap (B) are shown in the bar graph. Data are represented by the mean ± SEM. Significance was determined with the two-tailed Student’s t test. ***, P < 0.001.

Mentions: The preceding experiments show that TRPA1 is activated by acetic acid and PA, but not by HCl, and, therefore, has the pharmacological properties expected for a sensor of weak acids in the trigeminal system. But is it required for the sensory response? To test this possibility, we measured responses of sensory neurons to acids with Ca2+ imaging. To avoid confounding our data with well-described effects of external pH acting through TRPV1 or ASIC channels that are coexpressed with TRPA1 in the nociceptors, the stimulus that we used was 100 mM PA at pH 6.5, which strongly activates sensory nerves in an intact preparation (Fig. 1 and Silver and Moulton, 1982; Bryant and Moore, 1995). This stimulus produced a large Ca2+ elevation in HEK-293 cells expressing TRPA1 (12/13 cells responded; Fig. 8 A), and the response had a characteristic biphasic time course. In contrast, only a small monophasic response was detected in cells transfected with TRPV1 (Fig. 8 B; 0/7 GFP+ cells). Because this small response was detected in all cells tested, we attributed it to previously described effects of intracellular pH on the emission of the Ca2+ indicator, rather than to a true elevation of intracellular Ca2+ (Lattanzio and Bartschat, 1991). Thus, 100 mM PA, pH 6.5, can be used to specifically test the effects of intracellular acidification under conditions that minimize extracellular acidification, in the context of native sensory neurons.


A TRPA1-dependent mechanism for the pungent sensation of weak acids.

Wang YY, Chang RB, Allgood SD, Silver WL, Liman ER - J. Gen. Physiol. (2011)

PA activates TRPA1 in Ca2+ imaging. Agonist-induced elevation of intracellular Ca2+ in HEK-293 cells transfected with TRPV1 or TRPA1. (A) TRPA1-expressing HEK cells responded to both 100 mM PA, pH 6.5, and 100 µM Cin. (B) TRPV1-expressing HEK cells responded only to 1 µM Cap and showed nonspecific responses to PA. Scatter plot shows the amplitude of the responses to PA as a function of the responses to Cin (A) or Cap (B). Average responses to PA and Cin (A) or PA and Cap (B) are shown in the bar graph. Data are represented by the mean ± SEM. Significance was determined with the two-tailed Student’s t test. ***, P < 0.001.
© Copyright Policy - openaccess
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC3105510&req=5

fig8: PA activates TRPA1 in Ca2+ imaging. Agonist-induced elevation of intracellular Ca2+ in HEK-293 cells transfected with TRPV1 or TRPA1. (A) TRPA1-expressing HEK cells responded to both 100 mM PA, pH 6.5, and 100 µM Cin. (B) TRPV1-expressing HEK cells responded only to 1 µM Cap and showed nonspecific responses to PA. Scatter plot shows the amplitude of the responses to PA as a function of the responses to Cin (A) or Cap (B). Average responses to PA and Cin (A) or PA and Cap (B) are shown in the bar graph. Data are represented by the mean ± SEM. Significance was determined with the two-tailed Student’s t test. ***, P < 0.001.
Mentions: The preceding experiments show that TRPA1 is activated by acetic acid and PA, but not by HCl, and, therefore, has the pharmacological properties expected for a sensor of weak acids in the trigeminal system. But is it required for the sensory response? To test this possibility, we measured responses of sensory neurons to acids with Ca2+ imaging. To avoid confounding our data with well-described effects of external pH acting through TRPV1 or ASIC channels that are coexpressed with TRPA1 in the nociceptors, the stimulus that we used was 100 mM PA at pH 6.5, which strongly activates sensory nerves in an intact preparation (Fig. 1 and Silver and Moulton, 1982; Bryant and Moore, 1995). This stimulus produced a large Ca2+ elevation in HEK-293 cells expressing TRPA1 (12/13 cells responded; Fig. 8 A), and the response had a characteristic biphasic time course. In contrast, only a small monophasic response was detected in cells transfected with TRPV1 (Fig. 8 B; 0/7 GFP+ cells). Because this small response was detected in all cells tested, we attributed it to previously described effects of intracellular pH on the emission of the Ca2+ indicator, rather than to a true elevation of intracellular Ca2+ (Lattanzio and Bartschat, 1991). Thus, 100 mM PA, pH 6.5, can be used to specifically test the effects of intracellular acidification under conditions that minimize extracellular acidification, in the context of native sensory neurons.

Bottom Line: Our results show that heterologously expressed TRPA1 currents can be induced by a series of weak organic acids, including acetic, propionic, formic, and lactic acid, but not by strong acids.Importantly, responses of trigeminal neurons to weak acids were highly overrepresented in the subpopulation of TRPA1-expressing neurons and were severely reduced in neurons from TRPA1 knockout mice.We conclude that TRPA1 is a general sensor for weak acids that produce intracellular acidification and suggest that it functions within the pain pathway to mediate sensitivity to cellular acidosis.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Biological Sciences, Section of Neurobiology, University of Southern California, Los Angeles, CA 90089, USA.

ABSTRACT
Acetic acid produces an irritating sensation that can be attributed to activation of nociceptors within the trigeminal ganglion that innervate the nasal or oral cavities. These sensory neurons sense a diverse array of noxious agents in the environment, allowing animals to actively avoid tissue damage. Although receptor mechanisms have been identified for many noxious chemicals, the mechanisms by which animals detect weak acids, such as acetic acid, are less well understood. Weak acids are only partially dissociated at neutral pH and, as such, some can cross the cell membrane, acidifying the cell cytosol. The nociceptor ion channel TRPA1 is activated by CO(2), through gating of the channel by intracellular protons, making it a candidate to more generally mediate sensory responses to weak acids. To test this possibility, we measured responses to weak acids from heterologously expressed TRPA1 channels and trigeminal neurons with patch clamp recording and Ca(2+) microfluorometry. Our results show that heterologously expressed TRPA1 currents can be induced by a series of weak organic acids, including acetic, propionic, formic, and lactic acid, but not by strong acids. Notably, the degree of channel activation was predicted by the degree of intracellular acidification produced by each acid, suggesting that intracellular protons are the proximate stimulus that gates the channel. Responses to weak acids produced a Ca(2+)-independent inactivation that precluded further activation by weak acids or reactive chemicals, whereas preactivation by reactive electrophiles sensitized TRPA1 channels to weak acids. Importantly, responses of trigeminal neurons to weak acids were highly overrepresented in the subpopulation of TRPA1-expressing neurons and were severely reduced in neurons from TRPA1 knockout mice. We conclude that TRPA1 is a general sensor for weak acids that produce intracellular acidification and suggest that it functions within the pain pathway to mediate sensitivity to cellular acidosis.

Show MeSH
Related in: MedlinePlus