An outline of desensitization in pentameric ligand-gated ion channel receptors

Abstract

Pentameric ligand-gated ion channel (pLGIC) receptors exhibit desensitization, the progressive reduction in ionic flux in the prolonged presence of agonist. Despite its pathophysiological importance and the fact that it was first described over half a century ago, surprisingly little is known about the structural basis of desensitization in this receptor family. Here, we explain how desensitization is defined using functional criteria. We then review recent progress into reconciling the structural and functional basis of this phenomenon. The extracellular-transmembrane domain interface is a key locus. Activation is well known to involve conformational changes at this interface, and several lines of evidence suggest that desensitization involves a distinct conformational change here that is incompatible with activation. However, major questions remain unresolved, including the structural basis of the desensitization-induced agonist affinity increase and the mechanism of pore closure during desensitization.

Fig. 1

a A top view schematic of a pLGIC showing its five-fold symmetry and the central ion permeation pathway. The five M2 α-helical segments, located within transmembrane domain, line a pore. Each subunit is given a different colour. b A side view schematic of a pLGIC showing two of the three functional domains, the extracellular and transmembrane (demarcated by black horizontal bars). The intracellular domain is not shown. The images were made using the crystal structure of the C. elegans α GluClR 3RIF [5]

Fig. 2

a Schematic of a single subunit indicating the extracellular-transmembrane domain and transmembrane-intracellular domain interfaces (arrows). The intracellular domain has been omitted. b A higher magnification view of a single subunit showing the elements of the contributing to the interface between extracellular and transmembrane domains. The elements are; loop 9 (green), loop 2 (light blue), loop 7 (dark blue), pre-M1 (purple) and N terminus of M1 (olive). Also shown are the other transmembrane α helices, including the pore-lining M2 (orange). The images were made using the crystal structure of the C. elegans α GluClR 3RIF [5]

Fig. 3

a Whole-cell current recorded from an HEK293 cell expressing homomeric α1 GlyRs showing the phases of an ensemble current in response to a saturating concentration of glycine. b Single channel currents recorded from outside-out patches expressing α1 GlyRs. The records show short bursts of activity at low ligand concentrations (top), which group into clusters at saturating ligand concentrations. The clusters are separated by non-conducting periods where the channel adopts desensitized states (bottom)

Fig. 4

A general reaction mechanism describing any ligand-gated ion channel that accommodates the binding of two ligand molecules. Un-, mono- and di-liganded shut (S), open (O), and desensitized (D) configurations are shown. The forward and backward arrows between connected states are associated with corresponding rate constants, which quantify the transition frequencies to and from the states. Only the diliganded opening (β), shutting (α), entry into desensitized (δ), and ligand dissociation (k₋₁) rate constants are shown

Fig. 5

A cartoon schematic illustrating the salient features of channel activation and desensitization and the two-gate mechanism. The unliganded shut (S) channel binds two molecules of ligand (orange), which initially induce a structural reorientation of the extracellular domain (dark grey) producing a liganded shut state (A ₂ S). This is followed by a structural change in the transmembrane domain (red), which disrupts the activation gate to produce an open channel (A ₂ O). The continued presence of ligand elicits a further structural change in the extracellular domain (light brown) and transmembrane domain (dark brown) that occludes the pore with a second desensitization gate

Fig. 6

Combined current and fluorescence recording of rhodamine-labeled A52C and A52C–A248L mutant GlyRs reveals a conformational change specifically associated with receptor desensitization. The A248L mutation in the M1–M2 loop induces fast desensitization. a Examples of current (black) and fluorescence (red) responses induced by a saturating glycine concentration in both GlyRs. Note the biphasic fluorescence response and the glycine-induced plateau in the double mutant GlyR. b Averaged time constants for current and fluorescence decay recorded from double mutant GlyRs plotted as a function of glycine concentration. There was no significant difference at any concentration, confirming that the fluorescence response reported a conformational change associated with desensitization. Figure modified from [30]