Dynamic conformational changes of acid-sensing ion channels in different desensitizing conditions

Acid-sensing ion channels (ASICs) are proton-gated cation channels that contribute to fast synaptic transmission and have roles in fear conditioning and nociception. Apart from activation at low pH, ASIC1a also undergoes several types of desensitization, including acute desensitization that terminates activation, steady-stated desensitization that occurs at sub-activating proton concentrations and limits subsequent activation, and tachyphylaxis that results in a progressive decrease in response during a series of activations. Structural insights from a desensitized state of ASIC1 have provided great spatial detail, but dynamic insights into conformational changes in different desensitizing conditions are largely missing. Here, we use electrophysiology and voltage-clamp fluorometry to follow the functional changes of the pore along with conformational changes at several positions in the extracellular and upper transmembrane domain via cysteine-labeled fluorophores. Acute desensitization terminates activation in wild-type but introducing an N414K mutation in the {beta}11-12 linker of mouse ASIC1a interfered with this process. The mutation also affected steady-state desensitization and led to pronounced tachyphylaxis. Common to all types of desensitization was that the extracellular domain remained sensitive to pH and underwent pH-dependent conformational changes. These conformational changes did, however, not necessarily lead to desensitization. N414K-containing channels remained sensitive to known peptide modulators that increased steady-state desensitization, indicating that the mutation only reduced, but not precluded, desensitization. Together, this study contributes to understanding the fundamental properties of ASIC1a desensitization, emphasizing the complex interplay between the conformational changes of the ECD and the pore during channel activation and desensitization. Statement of significanceAcid-sensing ion channels (ASICs) are proton-gated ion channels that contribute to synaptic activity and play roles in acidosis-related diseases. Prolonged acidosis can lead to desensitization in ASIC1a, and modulators that affect this desensitization have shown beneficial effects in pain and stroke. In this study, we investigated the functional and conformational changes during acute desensitization, steady-state desensitization, and tachyphylaxis through a mutation in the {beta}11-12 linker of ASIC1a. We found that the mutation retained pH-dependent conformational changes of the extracellular domain (ECD) but largely disconnected these movements from the channel pore. Collectively, our work emphasizes the critical role of the {beta}11-12 linker for the pH-dependent conformational interplay between the ECD and the channel pore.