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Proton and non-proton activation of ASIC channels

View Article: PubMed Central - PubMed

ABSTRACT

The Acid-Sensing Ion Channels (ASIC) exhibit a fast desensitizing current when activated by pH values below 7.0. By contrast, non-proton ligands are able to trigger sustained ASIC currents at physiological pHs. To analyze the functional basis of the ASIC desensitizing and sustained currents, we have used ASIC1a and ASIC2a mutants with a cysteine in the pore vestibule for covalent binding of different sulfhydryl reagents. We found that ASIC1a and ASIC2a exhibit two distinct currents, a proton-induced desensitizing current and a sustained current triggered by sulfhydryl reagents. These currents differ in their pH dependency, their sensitivity to the sulfhydryl reagents, their ionic selectivity and their relative magnitude. We propose a model for ASIC1 and ASIC2 activity where the channels can function in two distinct modes, a desensitizing mode and a sustained mode depending on the activating ligands. The pore vestibule of the channel represents a functional site for binding non-proton ligands to activate ASIC1 and ASIC2 at neutral pH and to prevent channel desensitization.

No MeSH data available.


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Representative tracings of ASIC1a-G430C currents elicited at different pH values after covalent modification with MTS-reagents.Oocytes expressing ASIC1-G430C were perfused at pH7.8 (blue line) and ASIC1a currents were elicited by a pH change (red line) to 7.4, 7.0, 6.5, or 6.0; amiloride (300 μM, dashed line) was added to bath upon pH return to 7.8. In (A) the oocyte expressing ASIC1a-G430C was not pre-incubated with MTS, while in all the other conditions oocytes were pre-incubated at pH 7.8 for 10 min with 100 μM of one of the following compounds: MTSET (B), MTSPTrEA (C), MTSEA-biotin (D), or MTSES (E).
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pone.0175293.g002: Representative tracings of ASIC1a-G430C currents elicited at different pH values after covalent modification with MTS-reagents.Oocytes expressing ASIC1-G430C were perfused at pH7.8 (blue line) and ASIC1a currents were elicited by a pH change (red line) to 7.4, 7.0, 6.5, or 6.0; amiloride (300 μM, dashed line) was added to bath upon pH return to 7.8. In (A) the oocyte expressing ASIC1a-G430C was not pre-incubated with MTS, while in all the other conditions oocytes were pre-incubated at pH 7.8 for 10 min with 100 μM of one of the following compounds: MTSET (B), MTSPTrEA (C), MTSEA-biotin (D), or MTSES (E).

Mentions: Fig 2 shows representative tracings of the ASIC1a-G430C-mediated inward currents elicited at different pH values, before and after covalent modification of the channel by exposure during 10 min to 100 μM of MTSET, MTSPTrEA, MTSEA-biotin, or MTSES. For the unmodified G430C, a fast transient current was observed at pHs below 7.0. After modification, an amiloride-sensitive, non-desensitizing inward current was already detected at pH 7.4 for MTSET, MTSPTrEA and MTSEA-biotin, but not for MTSES. At pH 7.0 and below, robust transient inward currents were observed followed by a non-desensitizing sustained current (Isust). After modification of G430C by negatively charged MTSES, no sustained current could be detected. These tracings suggest that the inward current elicited by protons on the G430C channel covalently modified by MTS-reagents results from two distinct currents, a non-desensitizing sustained current (Isust.) and a fast desensitizing current (Idesens).


Proton and non-proton activation of ASIC channels
Representative tracings of ASIC1a-G430C currents elicited at different pH values after covalent modification with MTS-reagents.Oocytes expressing ASIC1-G430C were perfused at pH7.8 (blue line) and ASIC1a currents were elicited by a pH change (red line) to 7.4, 7.0, 6.5, or 6.0; amiloride (300 μM, dashed line) was added to bath upon pH return to 7.8. In (A) the oocyte expressing ASIC1a-G430C was not pre-incubated with MTS, while in all the other conditions oocytes were pre-incubated at pH 7.8 for 10 min with 100 μM of one of the following compounds: MTSET (B), MTSPTrEA (C), MTSEA-biotin (D), or MTSES (E).
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC5383329&req=5

pone.0175293.g002: Representative tracings of ASIC1a-G430C currents elicited at different pH values after covalent modification with MTS-reagents.Oocytes expressing ASIC1-G430C were perfused at pH7.8 (blue line) and ASIC1a currents were elicited by a pH change (red line) to 7.4, 7.0, 6.5, or 6.0; amiloride (300 μM, dashed line) was added to bath upon pH return to 7.8. In (A) the oocyte expressing ASIC1a-G430C was not pre-incubated with MTS, while in all the other conditions oocytes were pre-incubated at pH 7.8 for 10 min with 100 μM of one of the following compounds: MTSET (B), MTSPTrEA (C), MTSEA-biotin (D), or MTSES (E).
Mentions: Fig 2 shows representative tracings of the ASIC1a-G430C-mediated inward currents elicited at different pH values, before and after covalent modification of the channel by exposure during 10 min to 100 μM of MTSET, MTSPTrEA, MTSEA-biotin, or MTSES. For the unmodified G430C, a fast transient current was observed at pHs below 7.0. After modification, an amiloride-sensitive, non-desensitizing inward current was already detected at pH 7.4 for MTSET, MTSPTrEA and MTSEA-biotin, but not for MTSES. At pH 7.0 and below, robust transient inward currents were observed followed by a non-desensitizing sustained current (Isust). After modification of G430C by negatively charged MTSES, no sustained current could be detected. These tracings suggest that the inward current elicited by protons on the G430C channel covalently modified by MTS-reagents results from two distinct currents, a non-desensitizing sustained current (Isust.) and a fast desensitizing current (Idesens).

View Article: PubMed Central - PubMed

ABSTRACT

The Acid-Sensing Ion Channels (ASIC) exhibit a fast desensitizing current when activated by pH values below 7.0. By contrast, non-proton ligands are able to trigger sustained ASIC currents at physiological pHs. To analyze the functional basis of the ASIC desensitizing and sustained currents, we have used ASIC1a and ASIC2a mutants with a cysteine in the pore vestibule for covalent binding of different sulfhydryl reagents. We found that ASIC1a and ASIC2a exhibit two distinct currents, a proton-induced desensitizing current and a sustained current triggered by sulfhydryl reagents. These currents differ in their pH dependency, their sensitivity to the sulfhydryl reagents, their ionic selectivity and their relative magnitude. We propose a model for ASIC1 and ASIC2 activity where the channels can function in two distinct modes, a desensitizing mode and a sustained mode depending on the activating ligands. The pore vestibule of the channel represents a functional site for binding non-proton ligands to activate ASIC1 and ASIC2 at neutral pH and to prevent channel desensitization.

No MeSH data available.


Related in: MedlinePlus