Lipids influence the ability of Cys-loop receptors to gate open in response to neurotransmitter binding, but the underlying mechanisms are poorly understood. With the nicotinic acetylcholine receptor (nAChR) from Torpedo, current models suggest that lipids modulate the natural equilibrium between resting and desensitized conformations. We show that the lipidinactivated nAChR is not desensitized, instead it adopts a novel conformation where the allosteric coupling between its neurotransmitter-binding sites and transmembrane pore is lost. The uncoupling is accompanied by an unmasking of previously buried residues, suggesting weakened association between structurally intact agonist-binding and transmembrane domains. These data combined with the extensive literature on Cys-loop receptor-lipid interactions suggest that the M4 transmembrane helix plays a key role as a lipid-sensor, translating bilayer properties into altered nAChR function.

The structural and functional properties of reconstituted nicotinic acetylcholine receptor membranes composed of phosphatidyl choline either with or without cholesterol and/or phosphatidic acid have been examined to test the hypothesis that receptor conformational equilibria are modulated by the physical properties of the surrounding lipid environment. Spectroscopic and chemical labeling data indicate that the receptor in phosphatidylcholine alone is stabilized in a desensitizedlike state, whereas the presence of either cholesterol or phosphatidic acid favors a resting-like conformation. Membranes that effectively stabilize a resting-like state exhibit a relatively large proportion of non-hydrogen-bonded lipid ester carbonyls, suggesting a relatively tight packing of the lipid head groups and thus a well ordered membrane. Functional reconstituted membranes also exhibit gel-to-liquid crystal phase transition temperatures that are higher than those of nonfunctional reconstituted membranes composed of phosphatidylcholine alone. Significantly, incorporation of the receptor into phosphatidic acid-containing membranes leads to a dramatic increase in both the lateral packing densities and the gel-to-liquid crystal phase transition temperatures of the reconstituted lipid bilayers. These results suggest a functional link between the nicotinic acetylcholine receptor and the physical properties of phosphatidic acid-containing membranes that could underlie the mechanism by which this lipid preferentially enhances receptor function.

Potentiation of neuronal nicotinic acetylcholine receptors by exogenous ligands is a promising strategy for treatment of neurological disorders including Alzheimer's disease and Schizophrenia. To gain insight into molecular mechanisms underlying potentiation, we examined ACh-induced single-channel currents through the human neuronal α7 acetylcholine receptor in the presence of the α7-specific potentiator PNU-120596 (PNU). Compared to the unusually brief single-channel opening episodes elicited by agonist alone, channel opening episodes in the presence of agonist and PNU are dramatically prolonged. Dwell time analysis reveals that PNU introduces two novel components into open-time histograms, indicating at least two degrees of PNU-induced potentiation. Openings of the longest potentiated class coalesce into clusters whose frequency and duration change over a narrow range of PNU concentration. At PNU concentrations approaching saturation, these clusters last up to several minutes, prolonging the sub-millisecond α7 opening episodes by several orders of magnitude. Mutations known to reduce PNU potentiation at the whole-cell level still give rise to multi-second long single-channel clusters. However mutation of five residues lining a cavity within each subunit's transmembrane domain abolishes PNU potentiation, defining minimal structural determinants of PNU potentiation.

Pentameric ligand-gated ion channels (pLGICs) mediate fast synaptic communication by converting chemical signals into an electrical response. Recently solved agonist-bound and agonist-free structures greatly extend our understanding of these complex molecular machines. A key challenge to a full description of function, however, is the ability to unequivocally relate determined structures to the canonical resting, open, and desensitized states. Here, we review current understanding of pLGIC structure, with a focus on the conformational changes underlying channel gating. We compare available structural information and review the evidence supporting the assignment of each structure to a particular conformational state. We discuss multiple factors that may complicate the interpretation of crystal structures, highlighting the potential influence of lipids and detergents. We contend that further advances in the structural biology of pLGICs will require deeper insight into the nature of pLGIC-lipid interactions.

Mukhtasimova et al. describe experimental modifications of the patch clamp technique that improve temporal resolution of currents through single acetylcholine receptor channels. The study not only distinguishes between the priming and gating steps, but it also reveals how rate and equilibrium constants change as a function of agonist occupancy.