Modulation of Acid-sensing Ion Channel 1a by Intracellular pH and Its Role in Ischemic Stroke

Ming-Hua Li,Tian-Dong Leng,Xue-Chao Feng,Tao Yang,Roger P Simon,Zhi-Gang Xiong

doi:10.1074/jbc.m115.713636

Abstract

An important contributor to brain ischemia is known to be extracellular acidosis, which activates acid-sensing ion channels (ASICs), a family of proton-gated sodium channels. Lines of evidence suggest that targeting ASICs may lead to novel therapeutic strategies for stroke. Investigations of the role of ASICs in ischemic brain injury have naturally focused on the role of extracellular pH in ASIC activation. By contrast, intracellular pH (pHi) has received little attention. This is a significant gap in our understanding because the ASIC response to extracellular pH is modulated by pHi, and activation of ASICs by extracellular protons is paradoxically enhanced by intracellular alkalosis. Our previous studies show that acidosis-induced cell injury in in vitro models is attenuated by intracellular acidification. However, whether pHi affects ischemic brain injury in vivo is completely unknown. Furthermore, whereas ASICs in native neurons are composed of different subunits characterized by distinct electrophysiological/pharmacological properties, the subunit-dependent modulation of ASIC activity by pHi has not been investigated. Using a combination of in vitro and in vivo ischemic brain injury models, electrophysiological, biochemical, and molecular biological approaches, we show that the intracellular alkalizing agent quinine potentiates, whereas the intracellular acidifying agent propionate inhibits, oxygen-glucose deprivation-induced cell injury in vitro and brain ischemia-induced infarct volume in vivo Moreover, we find that the potentiation of ASICs by quinine depends on the presence of the ASIC1a, ASIC2a subunits, but not ASIC1b, ASIC3 subunits. Furthermore, we have determined the amino acids in ASIC1a that are involved in the modulation of ASICs by pHi.

Highlights

A common feature of brain ischemia is acidosis [1,2,3,4,5], which plays a critical role in the ensuing brain injury
Similar to lactate dehydrogenase (LDH) measurements, fluorescein diacetate and propidium iodide staining of alive and dead neurons taken at 6 h following the 1 h of OGD treatment in the absence or presence of quinine or propionate revealed that increasing intracellular pH potentiates and intracellular acidification inhibits OGD-induced neuronal injury (Fig. 1A)
Effect of Intracellular pH on acid-sensing ion channels (ASICs) Currents Carried by ASIC1a Point Mutations—we aimed to identify specific amino acid(s) that may be involved in the modulation of Quinine on kinetics of desensitization of Chinese hamster ovary (CHO) cells expressing ASIC1a and ASIC2a subunits

Summary

Introduction

A common feature of brain ischemia is acidosis [1,2,3,4,5], which plays a critical role in the ensuing brain injury. Quinine is a widely used agent for intracellular alkalization, whereas addition of propionate or NH4Cl withdrawal induces intracellular acidification [36] These agents do not directly activate ASIC currents in either cultured cortical neurons or ASIC1a-expressing Chinese hamster ovary (CHO) cells by themselves, but they modulate ASIC activity induced by extracellular acidosis [38]. This suggests that the function of ASICs is modulated by extracellular pH and intracellular pH. Using in vitro (oxygen-glucose deprivation, OGD) and in vivo (a transient middle cerebral artery occlusion, MCAO) ischemia models, combined with electrophysiological, biochemical, and molecular biological approaches, here we determined whether changing intracellular pH affects ischemic brain injury. The potential domains and/or site(s) involved in the modulation of ASICs by intracellular pH were examined

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Conclusion