[3H]-methyllycaconitine: a high affinity radioligand that labels invertebrate nicotinic acetylcholine receptors

Robert J. Lind, David J. Hardick, Ian S. Blagbrough, Barry V. L. Potter, Adrian J. Wolstenholme, Andrew R. L. Davies, Martin S. Clough, Fergus G. P. Earley, Stuart E Reynolds, Susan Wonnacott

Research output: Contribution to journalArticle

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Abstract

Nicotinic acetylcholine receptors (nAChR) of insect and other invertebrates are heterogeneous and new tools are needed to dissect their multiplicity. [H-3]-Methyllycaconitine ([H-3]-MLA) is a novel radioligand which is a potent antagonist at vertebrate alpha7-type nAChR. Putative invertebrate nAChR of the aphid Myzus persicae, the moths Heliothis virescens and Manduca sexta, the fly Lucilia sericata, and the squid Loligo vulgaris were investigated in radioligand binding studies with [H-3]-MLA. Saturable binding was consistent with a single class of high affinity binding sites for each of these invertebrates, characterised by a dissociation constant, K-d, of approximately I nM and maximal binding capacities, B-max, between 749 and 1689 fmol/mg protein for the insects and 14,111 fmol/mg protein for squid. [H-3]-MLA binding to M. persicae membranes was characterised in more detail. Kinetic analysis demonstrated rapid association in a biphasic manner and slow, monophasic dissociation. Displacement studies demonstrate the nicotinic character of [H-3]-MLA binding sites. Data for all nicotinic ligands, except MLA itself, are consistent with displacement from a high and a low affinity site, indicating that displacement is occurring from two or more classes of nicotinic binding site that are not distinguished by MLA itself. Autoradiographic analysis of the distribution of [H-3]-MLA binding sites in Manduca sexta shows discrete labelling of neuropil areas of the optic and antennal lobes.
LanguageEnglish
Pages533-542
JournalInsect Biochemistry and Molecular Biology
Volume31
Issue number6-7
DOIs
StatusPublished - 2001

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Affinity Labels
cholinergic receptors
Nicotinic Receptors
Invertebrates
binding sites
invertebrates
Binding Sites
Manduca
Decapodiformes
Aphids
Myzus persicae
Manduca sexta
squid
alpha7 Nicotinic Acetylcholine Receptor
Loligo
Insect Proteins
Lucilia sericata
insect proteins
antennal lobe
optic lobe

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[3H]-methyllycaconitine : a high affinity radioligand that labels invertebrate nicotinic acetylcholine receptors. / Lind, Robert J.; Hardick, David J.; Blagbrough, Ian S.; Potter, Barry V. L.; Wolstenholme, Adrian J.; Davies, Andrew R. L.; Clough, Martin S.; Earley, Fergus G. P.; Reynolds, Stuart E; Wonnacott, Susan.

In: Insect Biochemistry and Molecular Biology, Vol. 31, No. 6-7, 2001, p. 533-542.

Research output: Contribution to journalArticle

Lind, Robert J. ; Hardick, David J. ; Blagbrough, Ian S. ; Potter, Barry V. L. ; Wolstenholme, Adrian J. ; Davies, Andrew R. L. ; Clough, Martin S. ; Earley, Fergus G. P. ; Reynolds, Stuart E ; Wonnacott, Susan. / [3H]-methyllycaconitine : a high affinity radioligand that labels invertebrate nicotinic acetylcholine receptors. In: Insect Biochemistry and Molecular Biology. 2001 ; Vol. 31, No. 6-7. pp. 533-542
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abstract = "Nicotinic acetylcholine receptors (nAChR) of insect and other invertebrates are heterogeneous and new tools are needed to dissect their multiplicity. [H-3]-Methyllycaconitine ([H-3]-MLA) is a novel radioligand which is a potent antagonist at vertebrate alpha7-type nAChR. Putative invertebrate nAChR of the aphid Myzus persicae, the moths Heliothis virescens and Manduca sexta, the fly Lucilia sericata, and the squid Loligo vulgaris were investigated in radioligand binding studies with [H-3]-MLA. Saturable binding was consistent with a single class of high affinity binding sites for each of these invertebrates, characterised by a dissociation constant, K-d, of approximately I nM and maximal binding capacities, B-max, between 749 and 1689 fmol/mg protein for the insects and 14,111 fmol/mg protein for squid. [H-3]-MLA binding to M. persicae membranes was characterised in more detail. Kinetic analysis demonstrated rapid association in a biphasic manner and slow, monophasic dissociation. Displacement studies demonstrate the nicotinic character of [H-3]-MLA binding sites. Data for all nicotinic ligands, except MLA itself, are consistent with displacement from a high and a low affinity site, indicating that displacement is occurring from two or more classes of nicotinic binding site that are not distinguished by MLA itself. Autoradiographic analysis of the distribution of [H-3]-MLA binding sites in Manduca sexta shows discrete labelling of neuropil areas of the optic and antennal lobes.",
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T2 - Insect Biochemistry and Molecular Biology

AU - Lind,Robert J.

AU - Hardick,David J.

AU - Blagbrough,Ian S.

AU - Potter,Barry V. L.

AU - Wolstenholme,Adrian J.

AU - Davies,Andrew R. L.

AU - Clough,Martin S.

AU - Earley,Fergus G. P.

AU - Reynolds,Stuart E

AU - Wonnacott,Susan

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N2 - Nicotinic acetylcholine receptors (nAChR) of insect and other invertebrates are heterogeneous and new tools are needed to dissect their multiplicity. [H-3]-Methyllycaconitine ([H-3]-MLA) is a novel radioligand which is a potent antagonist at vertebrate alpha7-type nAChR. Putative invertebrate nAChR of the aphid Myzus persicae, the moths Heliothis virescens and Manduca sexta, the fly Lucilia sericata, and the squid Loligo vulgaris were investigated in radioligand binding studies with [H-3]-MLA. Saturable binding was consistent with a single class of high affinity binding sites for each of these invertebrates, characterised by a dissociation constant, K-d, of approximately I nM and maximal binding capacities, B-max, between 749 and 1689 fmol/mg protein for the insects and 14,111 fmol/mg protein for squid. [H-3]-MLA binding to M. persicae membranes was characterised in more detail. Kinetic analysis demonstrated rapid association in a biphasic manner and slow, monophasic dissociation. Displacement studies demonstrate the nicotinic character of [H-3]-MLA binding sites. Data for all nicotinic ligands, except MLA itself, are consistent with displacement from a high and a low affinity site, indicating that displacement is occurring from two or more classes of nicotinic binding site that are not distinguished by MLA itself. Autoradiographic analysis of the distribution of [H-3]-MLA binding sites in Manduca sexta shows discrete labelling of neuropil areas of the optic and antennal lobes.

AB - Nicotinic acetylcholine receptors (nAChR) of insect and other invertebrates are heterogeneous and new tools are needed to dissect their multiplicity. [H-3]-Methyllycaconitine ([H-3]-MLA) is a novel radioligand which is a potent antagonist at vertebrate alpha7-type nAChR. Putative invertebrate nAChR of the aphid Myzus persicae, the moths Heliothis virescens and Manduca sexta, the fly Lucilia sericata, and the squid Loligo vulgaris were investigated in radioligand binding studies with [H-3]-MLA. Saturable binding was consistent with a single class of high affinity binding sites for each of these invertebrates, characterised by a dissociation constant, K-d, of approximately I nM and maximal binding capacities, B-max, between 749 and 1689 fmol/mg protein for the insects and 14,111 fmol/mg protein for squid. [H-3]-MLA binding to M. persicae membranes was characterised in more detail. Kinetic analysis demonstrated rapid association in a biphasic manner and slow, monophasic dissociation. Displacement studies demonstrate the nicotinic character of [H-3]-MLA binding sites. Data for all nicotinic ligands, except MLA itself, are consistent with displacement from a high and a low affinity site, indicating that displacement is occurring from two or more classes of nicotinic binding site that are not distinguished by MLA itself. Autoradiographic analysis of the distribution of [H-3]-MLA binding sites in Manduca sexta shows discrete labelling of neuropil areas of the optic and antennal lobes.

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DO - 10.1016/S0965-1748(00)00153-3

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JO - Insect Biochemistry and Molecular Biology

JF - Insect Biochemistry and Molecular Biology

SN - 0965-1748

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ER -