Premium
A novel fluorescent α‐conotoxin for the study of α7 nicotinic acetylcholine receptors
Author(s) -
Hone Arik J.,
Whiteaker Paul,
Christensen Sean,
Xiao Yingxian,
Meyer Erin L.,
McIntosh J. Michael
Publication year - 2009
Publication title -
journal of neurochemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.75
H-Index - 229
eISSN - 1471-4159
pISSN - 0022-3042
DOI - 10.1111/j.1471-4159.2009.06299.x
Subject(s) - homomeric , receptor , nicotinic agonist , acetylcholine receptor , biology , acetylcholine , xenopus , fluorescence , biophysics , conotoxin , microbiology and biotechnology , biochemistry , chemistry , protein subunit , pharmacology , peptide , physics , quantum mechanics , gene
Homomeric α7 nicotinic acetylcholine receptors are a well‐established, pharmacologically distinct subtype. The more recently identified α9 subunit can also form functional homopentamers as well as α9α10 heteropentamers. Current fluorescent probes for α7 nicotinic ACh receptors are derived from α‐bungarotoxin (α‐BgTx). However, α‐BgTx also binds to α9* and α1* receptors which are coexpressed with α7 in multiple tissues. We used an analog of α‐conotoxin ArIB to develop a highly selective fluorescent probe for α7 receptors. This fluorescent α‐conotoxin, Cy3‐ArIB[V11L;V16A], blocked ACh‐evoked α7 currents in Xenopus laevis oocytes with an IC 50 value of 2.0 nM. Observed rates of blockade were minute‐scale with recovery from blockade even slower. Unlike FITC‐conjugated α‐BgTx, Cy3‐ArIB[V11L;V16A] did not block α9α10 or α1β1δε receptors. In competition binding assays, Cy3‐ArIB[V11L;V16A] potently displaced [ 125 I]‐α‐BgTx binding to mouse hippocampal membranes with a K i value of 21 nM. Application of Cy3‐ArIB[V11L;V16A] resulted in specific punctate labeling of KXα7R1 cells but not KXα3β2R4, KXα3β4R2, or KXα4β2R2 cells. This labeling could be abolished by pre‐treatment with α‐cobratoxin. Thus, Cy3‐ArIB[V11L;V16A] is a novel and selective fluorescent probe for α7 receptors.