Effectiveness of the Qube in Studying the Rapidly-Desensitizing Alpha7 Nicotinic Acetylcholine Receptor

Abstract

Listen

Translate article

Alpha7 nicotinic acetylcholine receptor (α7 nAChR) is an integral part of the excitatory cholinergic nervous system and must be maintained in humans. α7 nAChRs serve as a primary immune response and are highly expressed in the nervous system. Moreover, the ion channel has a direct link to schizophrenia and Alzheimer's disease including several other classes of neurological ailments such as cognitive and memory impairment, therefore, making α7 a critical target of ion channel drug discovery. Several different subunits and combinations can comprise the pentameric ligand-gated ion channel. In the case of α7, five identical α7 homomeric subunits form a functional channel comprised of an extracellular ligand-binding domain, a transmembrane domain, containing the ion gate and the pore, and an intracellular regulatory domain. Binding of a single ligand is enough to activate the channel. α7 contain three major conformational states: closed (C), open (O), and desensitized (D). In the open state, the channel conducts Na+, K+, and Ca2+ ions with Ca2+ displaying highest permeability. Historically, the very rapid agonist-induced channel activation and desensitization make α7 a challenging target to study where the data are limited by the temporal resolution of the protocol and the proper solution exchange rate used to measure current. This contrasts with chimeras comprising the ligand binding domain of α7 receptors and the ion pore domain of the homomeric α1 glycine receptors, which desensitize to a lesser degree than homomeric α7 counterparts. Our results demonstrated the reliability of the Qube, our high-throughput 384-well automated patch-clamp (APC), in recording the α7 nAChR and α7-nAChR/GlyR chimeras in recombinantly expressed HEK293 cells with our microfluidic QChips that allow complete solution exchange coupled to customized rapid-stacked solution-exchange.