Extracellular acidification is a hallmark of various physiological and pathological processes, regulating a plethora of plasma membrane proteins. In the nervous system, acidification is a large determinant of synaptic regulation and neuronal death translated by Acid-Sensing Ion Channels (ASICs). As proton-gated ion channels, ASICs are activated by reductions in extracellular pH89, where protonation of the extracellular domain elicits a conformational cascade ultimately leading to pore opening and sodium permeation. Despite persistent acidification, these channels undergo an intrinsic failsafe mechanism where over time the pore closes permanently for the rest of the duration of acidification. This process is known as desensitization, a common phenomenon that occurs in many neurotransmitter-gated ion channels. In the first act of this dissertation, we delve into the molecular mechanisms and regulation behind the desensitization process in ASICs. Utilizing outside-out and whole cell patch clamp electrophysiology, we functionally assess a region within the extracellular domain, the ?11-12 linker, a region that undergoes a large conformational change when going from the activated or open state to the desensitized state. Located between the distal extracellular regions and the pore, we hypothesized as being the clutch that is flipped to uncouple these regions to allow for pore closure ultimately governing desensitization. Through site-directed mutagenesis, we found the side chain profile of two residues within the linker of chicken ASIC1, L414 and N415, had a substantial effect on channel desensitization and recovery from desensitization kinetics. Further, we interrogated the influence of residues that surround the ?11-12 linker on channel kinetics through mutagenesis. I found several key interactions that play a role in the stabilization of L414 and N415 in their respective desensitized conformation. Finally, utilizing non-canonical amino acid (ncAA) incorporation of Bpa, a UV-sensitive photocrosslinking amino acid, we determined that the conformational flip of the ?11-12 linker was necessary for channel desensitization. In the second act of this dissertation, we ventured into investigating the role of the ?4-5 interface of the thumb domain during channel activation. During the process of activation, the ?5 helix rotates into a region known as the acidic pocket, creating the largest conformational change in the extracellular domain, known as the collapse of the acidic pocket. To address the effects of this collapse on ASIC1a gating, we incorporated two photocrosslinkable amino acids, Bpa and AzF, into positions within the ?4-5 interface of the thumb. Here, we found that restriction of movement via crosslinking induced acceleration of desensitization and deactivation kinetics, similar to chloride ablation, while significantly reducing the pH sensitivity of the channel. According to solved structures of the channel, this region contains a chloride binding site, where chloride is only bound in the collapsed conformation of the thumb domain. We hypothesized that chloride binds at this interface, stabilizing the open state of human ASIC1a. Combining outside-out patch clamp electrophysiology, anion substitution and site-directed mutagenesis, we revealed that ablation of chloride binding elicits two profound effects on channel gating: acceleration of channel desensitization and deactivation, two processes that involve withdrawing of the channel from the open state. Further, we revealed through state-dependent experiments using anion substitution, that chloride likely binds exclusively to the protonated, collapsed conformation of the thumb domain to elicit these effects. In summary, these findings show that collapse of the thumb domain allows for a stabilization of the open state of ASIC1a, partially through chloride binding.