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Extracellular chloride modulates the desensitization kinetics of acid-sensing ion channel 1a (ASIC1a).

ABSTRACT: Acid-sensing ion channels (ASICs) are sodium channels gated by extracellular protons. The recent crystallization of ASIC1a identified potential binding sites for Cl(-) in the extracellular domain that are highly conserved between ASIC isoforms. However, the significance of Cl(-) binding is unknown. We investigated the effect of Cl(-) substitution on heterologously expressed ASIC1a current and H(+)-gated currents from hippocampal neurons recorded by whole-cell patch clamp. Replacement of extracellular Cl(-) with the impermeable and inert anion methanesulfonate (MeSO(3)(-)) caused ASIC1a currents to desensitize at a faster rate and attenuated tachyphylaxis. However, peak current amplitude, pH sensitivity, and selectivity were unchanged. Other anions, including Br(-), I(-), and thiocyanate, also altered the kinetics of desensitization and tachyphylaxis. Mutation of the residues that form the Cl(-)-binding site in ASIC1a abolished the modulatory effects of anions. The results of anion substitution on native ASIC channels in hippocampal neurons mirrored those in heterologously expressed ASIC1a and altered acid-induced neuronal death. Anion modulation of ASICs provides new insight into channel gating and may prove important in pathological brain conditions associated with changes in pH and Cl(-).

SUBMITTER: Kusama N

PROVIDER: S-EPMC2878506 | biostudies-literature | 2010 Jun

REPOSITORIES: biostudies-literature

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Extracellular chloride modulates the desensitization kinetics of acid-sensing ion channel 1a (ASIC1a).

Kusama Nobuyoshi N Harding Anne Marie S AM Benson Christopher J CJ

The Journal of biological chemistry 20100412 23

Acid-sensing ion channels (ASICs) are sodium channels gated by extracellular protons. The recent crystallization of ASIC1a identified potential binding sites for Cl(-) in the extracellular domain that are highly conserved between ASIC isoforms. However, the significance of Cl(-) binding is unknown. We investigated the effect of Cl(-) substitution on heterologously expressed ASIC1a current and H(+)-gated currents from hippocampal neurons recorded by whole-cell patch clamp. Replacement of extracel ...[more]

PMID: 20385551

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Project description:Background and Purpose- Sex differences in the incidence and outcome of stroke have been well documented. The severity of stroke in women is, in general, significantly lower than that in men, which is mediated, at least in part, by the protective effects of β-estradiol. However, the detailed mechanisms underlying the neuroprotection by β-estradiol are still elusive. Recent studies have demonstrated that activation of ASIC1a (acid-sensing ion channel 1a) by tissue acidosis, a common feature of brain ischemia, plays an important role in ischemic brain injury. In the present study, we assessed the effects of β-estradiol on acidosis-mediated and ischemic neuronal injury both in vitro and in vivo and explored the involvement of ASIC1a and underlying mechanism. Methods- Cultured neurons and NS20Y cells were subjected to acidosis-mediated injury in vitro. Cell viability and cytotoxicity were measured by methylthiazolyldiphenyl-tetrazolium bromide and lactate dehydrogenase assays, respectively. Transient (60 minutes) focal ischemia in mice was induced by suture occlusion of the middle cerebral artery in vivo. ASIC currents were recorded using whole-cell patch-clamp technique while intracellular Ca2+ concentration was measured with fluorescence imaging using Fura-2. ASIC1a expression was detected by Western blotting and quantitative real-time polymerase chain reaction. Results- Treatment of neuronal cells with β-estradiol decreased acidosis-induced cytotoxicity. ASIC currents and acid-induced elevation of intracellular Ca2+ were all attenuated by β-estradiol treatment. In addition, we showed that β-estradiol treatment reduced ASIC1a protein expression, which was mediated by increased protein degradation, and that estrogen receptor α was involved. Finally, we showed that the level of ASIC1a protein expression in brain tissues and the degree of neuroprotection by ASIC1a blockade were lower in female mice, which could be attenuated by ovariectomy. Conclusions- β-estradiol can protect neurons against acidosis-mediated neurotoxicity and ischemic brain injury by suppressing ASIC1a protein expression and channel function. Visual Overview- An online visual overview is available for this article.

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Extracellular chloride modulates the desensitization kinetics of acid-sensing ion channel 1a (ASIC1a).

Publications

Extracellular chloride modulates the desensitization kinetics of acid-sensing ion channel 1a (ASIC1a).

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