Background: Acid-sensing ion channels (ASICs) are activated by extracellular protons and are inhibited by amiloride. Results: Amiloride activates and sensitizes ASIC3 channels depending on the nonproton ligand sensing domain. Conclusion: ASICs can sense nonproton ligands in addition to protons. Significance: The results indicate caution in the use of amiloride for studying ASIC physiology and in the development of amiloride-derived ASIC inhibitors for treating pain syndromes.
Acid-sensing ion channels (ASICs) have emerged as important, albeit challenging therapeutic targets for pain, stroke, etc. One approach to developing therapeutic agents could involve the generation of functional antibodies against these channels. To select such antibodies, we used channels assembled in nanodiscs, such that the target ASIC1a has a configuration as close as possible to its natural state in the plasma membrane. This methodology allowed selection of functional antibodies that inhibit acid-induced opening of the channel in a dose-dependent way. In addition to regulation of pH, these antibodies block the transport of cations, including calcium, thereby preventing acid-induced cell death in vitro and in vivo. As proof of concept for the use of these antibodies to modulate ion channels in vivo, we showed that they potently protect brain cells from death after an ischemic stroke. Thus, the methodology described here should be general, thereby allowing selection of antibodies to other important ASICs, such as those involved in pain, neurodegeneration, and other conditions.
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