Oregon Health & Science University
This thesis is centered on acid-sensing ion channels (ASICs) and their properties of activation and desensitization. These channels are acid gated channels that are expressed throughout the nervous system. Although their physiological role is unclear, it is thought that they may be sensors of nociceptive signals brought on by changes in extracellular pH. What these channels are doing in the central nervous system is less clear. Recently, it has been proposed that they may play a pathological role during such ischemic conditions as stroke and seizure. Under ischemic conditions in the CNS, several ionic changes occur which may activate or enhance activation of ASICs: 1) anaerobic respiration causes the extracellular pH to decrease to 6.3, from a resting level of pH 7.4, 2) extracellular calcium can fall as low as 0.1 mM 3) extracellular zinc can increase to as high as 300 11M. In this thesis, I look at how these conditions affect activation of ASIC1a and ASIC1a/2a. I find that all of these conditions can activate or enhance ASICs in the CNS. In addition, these ASICs also exhibit a voltage dependent block by magnesium, which would be relieved by depolarizations induced during ischemia. Entry of sodium or calcium through ASICs during ischemia could then act to cause or enhance neuronal death through both necrotic and apoptotic pathways. ASICs exhibit desensitization under prolonged acidification. The time course of this can vary depending on the subtype from milliseconds to tens of seconds. I performed some structure function experiments to try and uncover both the mechanism of desensitization as well as identify what creates the distinct kinetics seen in different subtypes. I found a conserved region (T75-S82 in ASIC3) outside the first transmembrane domain that influences desensitization. Mutations in this region led to changes in desensitization kinetics and the pH dependence of desensitization. I also found that some variable residues, particularly residue 82, can create some of the differences seen in the kinetics of different subtypes. By using substituted cysteine accessibility experiments, I also showed that there is a conformational change that accompanies desensitization. This change buries a highly conserved glutamate at position 79 in ASIC3. This provides new evidence that there is a conformational change outside TMl during desensitization. This region appears to be critical for regulating desensitization of ASICs and is likely involved in a large conformational change that leads to the desensitized state.
Neuroscience Graduate Program
School of Medicine
Cushman, Kenneth A., "Activation and desensitization of acid-sensing ion channels" (2007). Scholar Archive. 798.