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Involvement of the alpha3 subunit in central nicotinic binding populations.

ABSTRACT: The alpha3 subunit gene was one of the first neuronal nicotinic acetylcholine receptor (nAChR) subunits to be cloned (Boulter et al., 1986), but direct evidence of alpha3 subunit contributions to mammalian central nAChR populations has not been presented. The studies reported here used mice engineered to contain a null mutation in the alpha3 nAChR subunit gene (Xu et al., 1999) to examine the involvement of the alpha3 subunit in central nAChR populations. Heterologously expressed alpha3beta2 and alpha3beta4 nAChRs are pharmacologically similar to native [125I]alpha-conotoxin MII (alpha-CtxMII)-binding and 3-(2(S)-azetidinylmethoxy)pyridine dihydrochloride (A85380)-resistant [125I]epibatidine-binding nAChR subtypes, respectively. The hypothesis that both native sites are alpha3-subtype nAChRs was tested using quantitative autoradiography in alpha3-null mutant mice. Somewhat surprisingly, deletion of the alpha3 nAChR subunit gene did not affect expression of the great majority of [125I]alpha-CtxMII-binding sites, indicating that they do not correspond to heterologously expressed alpha3beta2 nAChRs. The only exception to this was observed in the habenulointerpeduncular tract, where alpha3-dependent [125I]alpha-CtxMII binding was observed. This finding may suggest the presence of an additional, minor nicotinic population in this pathway. In contrast, most -resistant [125I]epibatidine-binding nAChRs were dependent on alpha3 gene expression, suggesting that they do indeed correspond to an alpha3 nAChR subtype. However, widespread but lower levels of alpha3-independent -resistant [125I]epibatidine binding were also seen. Again, this may indicate the existence of an additional, minor population of non-alpha3 -resistant sites.

SUBMITTER: Whiteaker P

PROVIDER: S-EPMC6758302 | biostudies-literature | 2002 Apr

REPOSITORIES: biostudies-literature

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Involvement of the alpha3 subunit in central nicotinic binding populations.

Whiteaker Paul P Peterson Cyrus G CG Xu Wei W McIntosh J Michael JM Paylor Richard R Beaudet Arthur L AL Collins Allan C AC Marks Michael J MJ

The Journal of neuroscience : the official journal of the Society for Neuroscience 20020401 7

The alpha3 subunit gene was one of the first neuronal nicotinic acetylcholine receptor (nAChR) subunits to be cloned (Boulter et al., 1986), but direct evidence of alpha3 subunit contributions to mammalian central nAChR populations has not been presented. The studies reported here used mice engineered to contain a null mutation in the alpha3 nAChR subunit gene (Xu et al., 1999) to examine the involvement of the alpha3 subunit in central nAChR populations. Heterologously expressed alpha3beta2 and ...[more]

PMID: 11923417

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Project description:Rapid neurotransmission is mediated through a superfamily of Cys-loop receptors that includes the nicotinic acetylcholine (nAChR), gamma-aminobutyric acid (GABA(A)), serotonin (5-HT(3)) and glycine receptors. A class of ligands, including galanthamine, local anesthetics and certain toxins, interact with nAChRs non-competitively. Suggested modes of action include blockade of the ion channel, modulation from undefined extracellular sites, stabilization of desensitized states, and association with annular or boundary lipid. Alignment of mammalian Cys-loop receptors shows aromatic residues, found in the acetylcholine or ligand-binding pocket of nAChRs, are conserved in all subunit interfaces of neuronal nAChRs, including those that are not formed by alpha subunits on the principal side of the transmitter binding site. The amino-terminal domain containing the ligand recognition site is homologous to the soluble acetylcholine-binding protein (AChBP) from mollusks, an established structural and functional surrogate. We assess ligand specificity and employ X-ray crystallography with AChBP to demonstrate ligand interactions at subunit interfaces lacking vicinal cysteines (i.e. the non-alpha subunit interfaces in nAChRs). Non-competitive nicotinic ligands bind AChBP with high affinity (K(d) 0.015-6 microM). We mutated the vicinal cysteine residues in loop C of AChBP to mimic the non-alpha subunit interfaces of neuronal nAChRs and other Cys loop receptors. Classical nicotinic agonists show a 10-40-fold reduction in binding affinity, whereas binding of ligands known to be non-competitive are not affected. X-ray structures of cocaine and galanthamine bound to AChBP (1.8 A and 2.9 A resolution, respectively) reveal interactions deep within the subunit interface and the absence of a contact surface with the tip of loop C. Hence, in addition to channel blocking, non-competitive interactions with heteromeric neuronal nAChR appear to occur at the non-alpha subunit interface, a site presumed to be similar to that of modulating benzodiazepines on GABA(A) receptors.

Dataset Information

Involvement of the alpha3 subunit in central nicotinic binding populations.

Publications

Involvement of the alpha3 subunit in central nicotinic binding populations.

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