Functional Characterization of Neurotransmitter Activation and Modulation in a Nematode Model Ligand-gated Ion Channel
2016 (English)In: Journal of Neurochemistry, ISSN 0022-3042, E-ISSN 1471-4159, Vol. 138, no 2, 243-253 p.Article in journal (Refereed) Published
The superfamily of pentameric ligand-gated ion channels includes neurotransmitter receptors that mediate fast synaptic transmission in vertebrates, and are targets for drugs including alcohols, anesthetics, benzodiazepines and anticonvulsants. However, the mechanisms of ion channel opening, gating and modulation in these receptors leave many open questions, despite their pharmacological importance. Subtle conformational changes in both the extracellular and transmembrane domains are likely to influence channel opening, but have been difficult to characterize given the limited structural data available for human membrane proteins. Recent crystal structures of a modifiedCaenorhabditis elegans glutamate-gated chloride channel (GluCl) in multiple states offer an appealing model system for structure-function studies. However, the pharmacology of the crystallographic GluCl construct is not well established. To establish the functional relevance of this system, we used two-electrode voltage-clamp electrophysiology in Xenopus oocytes to characterize activation of crystallographic and native-like GluCl constructs by L-glutamate and ivermectin. We also tested modulation by ethanol and other anesthetic agents, and used site-directed mutagenesis to explore the role of a region of Loop F which was implicated in ligand gating by molecular dynamics simulations. Our findings indicate that the crystallographic construct functionally models concentration-dependent agonism and allosteric modulation of pharmacologically relevant receptors. Specific substitutions at residue Leu174 in loop F altered direct L-glutamate activation, consistent with computational evidence for this region's role in ligand binding. These insights demonstrate conservation of activation and modulation properties in this receptor family, and establish a framework for GluCl as a model system, including new possibilities for drug discovery.
Place, publisher, year, edition, pages
John Wiley & Sons, 2016. Vol. 138, no 2, 243-253 p.
anesthetic, cys-loop receptor, GluCl, Ion channel, pLGIC
Biochemistry and Molecular Biology
IdentifiersURN: urn:nbn:se:kth:diva-187250DOI: 10.1111/jnc.13644ISI: 000380263700004PubMedID: 27102368ScopusID: 2-s2.0-84979036020OAI: oai:DiVA.org:kth-187250DiVA: diva2:929437
FunderSwedish Research Council, 2013-5901Swedish e‐Science Research Center
QC 201605182016-05-182016-05-182016-09-07Bibliographically approved