Secondary Logo

Journal Logo

Editorial: Editorials

Interesting Insights from a Very Simple (Unicellular) Source

Lynch, Carl III, MD, PhD

Author Information
doi: 10.1213/ANE.0b013e3181c5b72b
  • Free

The nicotinic acetylcholine receptors (nAChRs), ionotropic γ-aminobutyric acid (GABA) receptors (GABAA and GABAC), glycine receptors, and ionotropic serotonin receptors (5HT3) all have a similar structure in which 5 membrane subunits of similar size and shape combine to form a transmembrane channel, something similar to the staves of a barrel. A large portion of each subunit extends out from the membrane into the extracellular fluid. This extracellular region contains a distinctive disulfide linkage, providing the “Cys-loop” moniker for these receptors.1 Based on the species distribution, this superfamily of pentameric ligand-gated ion channels (pLGICs) was thought to be restricted to multicellular organisms. However, recently, pentameric proteins with large extracellular ligand-binding domains have been described in single-celled organisms.2–5 Although these channels lack the Cys-loop, their overall structure is remarkably similar to the pLGICs (see Fig. 1). Importantly, as demonstrated in the report by Weng et al.,6 the proton-activated currents of these channels are sensitive to both volatile anesthetics and propofol. But why should anyone care about the effects of anesthetics on a membrane protein from a Cyanobacteria (blue-green algae) Gloeobacter (aka GLIC)?

Figure 1.
Figure 1.:
Structures of the Gloeobacter ligand-gated ion channel (GLIC, left) and a nicotinic acetylcholine receptor (nAChR, right) channel derived from radiograph diffraction and reproduced from the National Center for Biotechnology Information (code 3EHZ or 3EI0 for GLIC; code 2BG9 for nAChR; available at: Each channel is composed of 5 subunits; 1 subunit in each channel is shown in cylinder (α-helix) and ribbon (β-sheet) format. The 5 M2 helices line the pore of the channel through which the ions pass. The M4 helix of each subunit actually extends most prominently into the lipid bilayer and is influenced by the order and structure of the lipid. The subunits of the AChR have an additional intracellular domain (MA) that forms a cage through which ions must pass. The linkage between the extracellular ligand-binding domain and the transmembrane bundle is mediated by salt bridges between these 2 regions (arrows). The approximate location of phospholipids and cholesterol, which constitute part of the bilayer, are shown in yellow.

Over the past 2 decades, by a combination of membrane electrophysiology, molecular biology, and structural proteomics (radiograph diffraction and nuclear magnetic resonance), we have come to a far more detailed understanding of the molecular basis of membrane excitation. Based on their primary structure/amino acid sequence, the multihelix bundles forming the structure of ion channels were proposed. Subsequent radiograph diffraction structural studies by MacKinnon7 and Unwin8,9 have defined the structure of ion channels at the atomic level and delineated the precise molecular rearrangements that occur as ion channels open and close. In Figure 1, the structure of GLIC deduced from radiograph diffraction data4,5 is shown next to the structure of an nAChR.8 Although the primary amino acid sequence shows a modest similarity of <20% with nAChR subunits, the similarity in secondary and tertiary structures is remarkable. Equally remarkable is the fact that when these channels are expressed in cells, they are inhibited by remarkably low concentrations of various anesthetic agents.

In considering the pLGICs it is noteworthy that although similar in structure, they have distinct behaviors. For example, the lining of transmembrane helices vary significantly such that nAChRs and 5HT3R transmit K+ and Na+ ions, whereas the GABA and glycine receptors transport Cl; the former depolarize the cell whereas the latter (usually) hyperpolarize or stabilize the membrane potential. More importantly, whereas anesthetics inhibit certain types of nAChR channels,10 anesthetics favor the opening of glycine and GABAA channels.11 For more than a decade, specific amino acid mutations in the nAChR12,13 and the GABAA14 receptor have been used to alter the sensitivity to volatile anesthetics and alcohol, and various investigators have inferred that specific binding sites exist on the channel protein. Are these differing anesthetic actions on various ion channels to be understood as actions of lipophilic molecules binding to distinct and randomly occurring amino acid sequences within each different membrane channel protein?

While we know that these pLGICs have a major role in mediating anesthetic actions, what can these bacterial channel members tell us? Despite divergent behavior and very modest homology in amino acid sequence, the similarity in overall structure of these pentameric proteins suggests that some common mechanisms of action may exist. Part of the problem in attempting to find a common theme is that in all these channels, the binding of a simple molecule to a cleft between the subunit extracellular regions results in significant movement of transmembrane helices so that there is large enough space for ions to pass through. The connection between the ligand-binding site and the transmembrane helices occurs over significant molecular distance (more than the thickness of the bilayer). Although yet to be completely elucidated, the change in structure seems to be mediated not by a fixed link but via electrostatic interactions (salt bridges) between the extracellular domain and transmembrane α-helices.15 This interaction site is at the plane of the membrane interfacial region—at the phospholipids head groups and polar portion of cholesterol. It is also in this region where tryptophan residues play an important role in anchoring the helical portion of the protein at the membrane surface.16 It seems more than coincidental that it is at the interfacial region of the membrane where volatile anesthetics reside.17,18

Another emerging clue is that many membrane proteins do not merely float in a sea of nonspecific lipid but instead have specific lipid requirements. For example, the function of the AChR is markedly enhanced and in some sense seems to require the presence of cholesterol in the bilayer,19,20 whereas various lipids can alter channel function21,22 or drug actions.23 Certain lipids are immobilized and remain tightly associated with pLGICs, while conversely membrane proteins also can cause specific organization of membrane lipids.24 Interestingly, the steroid hormone progesterone has anesthetic potency and similar to anesthetics inhibits AChR channels but activates GABAA channels. Anesthetically active steroids are large and bulky molecules, and it is difficult to imagine that it would occupy the same site as a small halogenated ether.

The view that anesthetics are acting at either a lipid or protein site may end up being far too simplistic. One suggestion is that the mechanical properties of the membrane lipids can mediate alterations in channel function.25–27 Although such global properties of the membrane may contribute, we are beginning to understand that there is a complex and sophisticated interaction between the specific lipids with various segments of the membrane proteins. For example, the “soft” interaction of the ligand-binding region with the transmembrane helices may require certain interfacial lipids. The linkage between the extracellular ligand-binding domain with the ion-gating region might be altered by a change in a critical amino acid in the region, a change in the lipid, or by anesthetic disruption of lipid-protein interaction. It is not impossible that lipids and anesthetics might weaken the linkage in one pLGIC (AChR) but strengthen it in another (GABA).

But what of these bacterial channels? The ability to mutate these molecules and produce them in sufficient quantity to study their function and their structure make them an exciting model for our more complex glycine, GABAA, nicotinic ACh, and 5HT3 channels. Photoactivated anesthetics may be used to determine with which regions of the receptor the drugs interact. Their protein structure can be mutated to determine the location of amino acids critical for permitting anesthetic action, and whether lipids will indeed have a role. Study of GLIC and ELIC3 (Erwinia chrysanthemi LIC, yet another bacterial LIC) will begin to answer how our structurally simple agents produce their panoply of actions.


1. Sine SM, Englel AG. Recent advances in Cys-loop receptor structure and function. Nature 2006;440:448–55
2. Bocquet N, de Carvalho LP, Cartaud J, Neyton J, Le Poupon C, Taly A, Grutter T, Changeux J-P, Corringer P-J. A prokaryotic proton-gated ion channel from the nicotinic acetylcholine receptor family. Nature 2007;445:116–9
3. Hilf RJC, Dutzler R. X-ray structure of a prokaryotic pentameric ligand-gated ion channel. Nature 2008;452:375–9
4. Bocquet N, Nury H, Baaden M, Le Poupon C, Changeux J-P, Delarue M, Corringer P-J. X-ray structure of a pentameric ligand-gated ion channel in an apparently open configuration. Nature 2009;457:111–4
5. Hilf RJC, Dutzler R. Structure of a potentially open state of a proton-activated pentameric ligand-gated ion channel. Nature 2009;457:115–8
6. Weng Y, Yang L, Corringer P-J, Sonner JM. Anesthetic sensitivity of the Gloeobacter violaceus proton-gated ion channel. Anesth Analg 2010;110:59–63
7. MacKinnon R. Potassium channels and the atomic basis of selective ion conduction. In: Frängsmyr T, ed. The Nobel Prizes. Stockholm: Nobel Foundation, 2003
8. Unwin N. Refined structure of the nicotinic acetylcholine receptor at 4 Å resolution. J Mol Biol 2005;346:967–89
9. Unwin N. Acetylcholine receptor channel imaged in the open state. Nature 1995;373:37–43
10. Flood P, Ramirez-Latorre J, Role L. α4β2 neuronal nicotinic acetylcholine receptors in the central nervous system are inhibited by isoflurane and propofol, but α7 nicotinic acetylcholine receptors are unaffected. Anesthesiology 1997;86:859–65
11. Jones MV, Brooks PA, Harrison NL. Enhancement of γ-aminobutyric acid-activated Cl currents in cultured rat hippocampal neurones by three volatile anesthetics. J Physiol 1992;449:279–93
12. Yamakura T, Borghese CM, Harris RA. A transmembrane site determines sensitivity of neuronal nicotinic acetylcholine receptors to general anesthetics. J Biol Chem 2000;275:40879–86
13. Wenningmann I, Barann M, Vidal AM, Dilger JP. The effects of isoflurane on acetylcholine receptor channels: 3. Effects of conservative polar-to-nonpolar mutations within the channel pore. Mol Pharmacol 2001;60:584–94
14. Mihic SJ, Ye Q, Wick MJ, Koltchine VV, Krasowski MC, Finn SE, Mascia MP, Valenzuela CF, Hanson KK, Greenblatt EP, Harris RA, Harrison NL. Sites of alcohol and volatile anaesthetic action on GABAA and glycine receptors. Nature 1997;389:385–9
15. Law RJ, Lightstone FC. Modeling neuronal nicotinic and GABA receptors: important interface salt-links and protein dynamics. Biophys J 2009;97:1586–94
16. Yau WM, Wimley WC, Gawrisch K, White SH. The preference of tryptophan for membrane interfaces. Biochemistry 1998;37: 14713–8
17. Baber J, Ellena JF, Cafiso DS. Distribution of general anesthetics in phospholipid bilayers determined using 2H NMR and 1H-1H NOE spectroscopy. Biochemistry 1995;34:6533–9
18. Tang P, Yan B, Xu Y. Different distribution of fluorinated anesthetics and nonanesthetics in model membrane: a 19F NMR study. Biophys J 1997;72:1676–82
19. Lechleiter J, Wells M, Gruener R. Halothane-induced changes in acetylcholine receptor channel kinetics are attenuated by cholesterol. Biochim Biophys Acta 1986;856:640–5
20. Barrantes FJ. Structural basis for lipid modulation of nicotinic acetylcholine receptor function. Brain Res Brain Res Rev 2004;47:71–95
21. Nievas GAF, Barrantes FJ, Antollini SS. Modulation of nicotinic acetylcholine receptor conformational state by free fatty acids and steroids. J Biol Chem 2008;283:21478–86
22. Nievas GAF, Barrantes FJ, Antollini SS. Conformation-sensitive steroid and fatty acid sites in the transmembrane domain of the nicotinic acetylcholine receptor. Biochemistry 2007;46:3503–12
23. Baenziger JE, Ryan SE, Goodreid MM, Vuong NQ, Sturgeon RM, daCosta CJB. Lipid composition alters drug action at the nicotinic acetylcholine receptor. Mol Pharmacol 2008;73:880–90
24. Wenz JJ, Barrantes FJ. Nicotinic acetylcholine receptor induces lateral segregation of phosphatidic acid and phosphatidylcholine in reconstituted membranes. Biochemistry 2005;44:398–410
25. Lundbæk JA, Andersen OS. Lysophospholipids modulate channel function by altering the mechanical properties of lipid bilayers. J Gen Physiol 1994;44:645–73
26. Cantor RS. The lateral pressure profile in membranes: a physical mechanism of general anesthesia. Biochemistry 1997;36:2339–44
27. Søgaard R, Werge TM, Bertelsen C, Lundbye C, Madsen KL, Nielsen CH, Lundbæk JA. GABAA receptor function is regulated by lipid bilayer fluidity. Biochemistry 2006;45:13118–29
© 2010 International Anesthesia Research Society