1: J Chromatogr B Analyt Technol Biomed Life Sci. 2003 Nov 25;797(1-2):373-9.


Enantioselective interactions of dextromethorphan and levomethorphan with the alpha 3 beta 4-nicotinic acetylcholine receptor: comparison of chromatographic and functional data.

Jozwiak K, Hernandez SC, Kellar KJ, Wainer IW.

Bioanalytical and Drug Discovery Unit, National Institute on Aging, National Institutes of Health, Gerontology Research Center, 5600 Nathan Shock Drive, Baltimore, MD 21224-6825, USA.

The enantioselectivity of the interaction of dextromethorphan (DM) and levomethorphan (LM) with an immobilized alpha 3 beta 4 subtype of the nicotinic acetylcholine receptor (nAChR) liquid chromatographic stationary phase has been compared to DM- and LM-induced non-competitive blockade of nicotine-stimulated 86Rb(+) efflux from cells expressing the alpha 3 beta 4-nAChR. The association rate constants (k(on)) and dissociation rate constants (k(off)) for the formation of the DM and LM complexes with the nAChR were determined using non-linear chromatographic techniques and the k(off) value for DM (1.01+/-0.01 s(-1)) was significantly lower than the k(off) for LM (1.55+/-0.002s(-1)) while the k(on) values did not significantly differ (23.66+/-0.61 and 18.61+/-0.36 microM(-1)s(-1), respectively). In thermodynamic studies using the van't Hoff approach, the enthalpy change (Delta H degrees) of the DM-nAChR complex was 330 calmol(-1) more stable than the LM-nAChR complex, while there was no significant difference in the entropy change (DeltaS degrees ). In the functional in vitro cell-based studies, there was no significant difference in the observed IC(50) values for DM (10.1+/-1.01 microM) and LM (10.9+/-1.08 microM), but the recovery from the DM-induced blockade was slower than the recovery from LM-induced blockade; after 7 min: 38.25+/-15.46% recovery from DM blockade, 63.30+/-16.08% from LM blockade; after 4h: 76.20+/-4.51% recovery from DM blockade and 93.12+/-8.76% from LM blockade. The enantioselective differences in the functional effects are consistent with the chromatographic and thermodynamic data and indicate that this difference is due to increased stability of the DM-nAChR complex. The results suggest that the chromatographic approach can be used to probe the interaction of non-competitive inhibitors (NCIs) with nAChRs and to predict relative duration of functional blockades.

Publication Types:
Comparative Study
Review

PMID: 14630163 [PubMed - indexed for MEDLINE]

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2: Mol Membr Biol. 2002 Oct-Dec;19(4):277-84.


Lipid matters: nicotinic acetylcholine receptor-lipid interactions (Review).

Barrantes FJ.

Instituto de Investigaciones Bioquímicas de Bahía Blanca, BZW8000 Bahía Blanca, Argentina. rtfjb1@criba.edu.ar

Ligand-gated ion channels mediate fast intercellular communication in response to endogenous neurotransmitters. The nicotinic acetylcholine receptor (AChR) is the archetype molecule in the superfamily of these membrane proteins. Early electron spin resonance studies led to the discovery of a lipid fraction in direct contact with the AChR, with rotational dynamics 50-fold slower than those of the bulk lipids. This AChR-vicinal lipid region has since been postulated to be a possible site of lipid modulation of receptor function. The polarity and molecular dynamics of solvent dipoles-mainly water-of AChR-vicinal lipids in the membrane have been studied with Laurdan extrinsic fluorescence, and Forster-type resonance energy transfer (FRET) was introduced to characterize the receptor-associated lipid microenvironment. FRET enabled one to discriminate between the bulk lipid and the AChR-vicinal lipid. The latter is in a liquid-ordered phase and exhibits a higher degree of order than the bulk bilayer lipid. Changes in FRET efficiency induced by fatty acids, phospholipids and cholesterol also led to the identification of discrete sites for these lipids on the AChR protein. After delineating the topography of the AChR membrane-embedded domains with fluorescence methods, sites for steroids are being explored with site-directed mutagenesis and patch-clamp recording. Pyrene-labelled Cys residues in alphaM1, alphaM4, gammaM1 and gammaM4 transmembrane regions were found to lie in a shallow position. For M4 segments, this is in agreement with a canonical linear alpha-helix; for M1, it is necessary to postulate a substantial amount of non-helical structure, and/or of kinks, to rationalize the shallow location of Cys residues. Mutations of Thr422, a residue close to the extracellular-facing membrane hemilayer in alphaM4, affect the steroid modulation of AChR function, suggesting its involvement in steroid-AChR interactions.

Publication Types:
Research Support, Non-U.S. Gov't
Review

PMID: 12512774 [PubMed - indexed for MEDLINE]

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3: Toxicol Lett. 1998 Nov 23;100-101:169-78.


Direct interactions of anesthetics and nonanesthetics with the nicotinic acetylcholine receptor pore.

Forman SA.

Department of Anesthesia and Critical Care, CLN-3, Massachusetts General Hospital, Boston 02114, USA. forman@helix.mgh.harvard.edu

(1) We review evidence that anesthetics inhibit peripheral nAChR cation translocation by binding directly to a protein site in the transmembrane pore. (2) This site is near the middle of the pore-forming M2 domains on alpha and beta subunits, but further from the homologous portions of gamma and delta subunits. (3) Interactions between both anesthetics and nonanesthetics with the nAChR pore site are determined primarily by hydrophobic forces rather than steric factors. (4) Anesthetics and nonanesthetics display different state-dependent accessibility to this site, suggesting a mechanism for the different in vivo actions of these two classes of drugs.

Publication Types:
Review

PMID: 10049138 [PubMed - indexed for MEDLINE]

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4: Md Med J. 1992 Jul;41(7):623-31.


Progress in understanding the nicotinic acetylcholine receptor function at central and peripheral nervous system synapses through toxin interactions.

Swanson KL, Albuquerque EX.

Department of Pharmacology and Experimental Therapeutics, University of Maryland School of Medicine.

The need to treat diseases affecting the nicotinic AChR is great, but therapeutic options are few. Through careful correlation of structure-activity relationships of AnTX analogs, we may ultimately be led to the development of diagnostic and therapeutic drugs with specific nicotinic agonist or antagonist activities in the central nervous system that would be of major importance in the treatment of Alzheimer's disease.

Publication Types:
Research Support, U.S. Gov't, Non-P.H.S.
Research Support, U.S. Gov't, P.H.S.
Review

PMID: 1640819 [PubMed - indexed for MEDLINE]