A Novel Fluorescent Î±-Conotoxin For The Study Of Î±7 Nicotinic Acetylcholine Receptors
Homomeric Î±7 nicotinic acetylcholine receptors are a well-established, pharmacologically distinct subtype. The more recently identified Î±9 subunit can also form functional homopentamers as well as Î±9Î±10 heteropentamers. Current fluorescent probes for Î±7 nicotinic ACh receptors are derived from Î±-bungarotoxin (Î±-BgTx). However, Î±-BgTx also binds to Î±9* and Î±1* receptors which are coexpressed with Î±7 in multiple tissues. We used an analog of Î±-conotoxin ArIB to develop a highly selective fluorescent probe for Î±7 receptors. This fluorescent Î±-conotoxin, Cy3-ArIB[V11L;V16A], blocked ACh-evoked Î±7 currents in Xenopus laevis oocytes with an IC50 value of 2.0 nM. Observed rates of blockade were minute-scale with recovery from blockade even slower. Unlike FITC-conjugated Î±-BgTx, Cy3-ArIB[V11L;V16A] did not block Î±9Î±10 or Î±1Î²1Î”Îµ receptors. In competition binding assays, Cy3-ArIB[V11L;V16A] potently displaced [125I]-Î±-BgTx binding to mouse hippocampal membranes with a Ki value of 21 nM. Application of Cy3-ArIB[V11L;V16A] resulted in specific punctate labeling of KXÎ±7R1 cells but not KXÎ±3Î²2R4, KXÎ±3Î²4R2, or KXÎ±4Î²2R2 cells. This labeling could be abolished by pre-treatment with Î±-cobratoxin. Thus, Cy3-ArIB[V11L;V16A] is a novel and selective fluorescent probe for Î±7 receptors. Â© 2009 International Society for Neurochemistry.
Journal of Neurochemistry
Digital Object Identifier (DOI)
Hone, Arik J.; Whiteaker, Paul; Christensen, Sean; Xiao, Yingxian; Meyer, Erin L.; and McIntosh, J. Michael, "A Novel Fluorescent Î±-Conotoxin For The Study Of Î±7 Nicotinic Acetylcholine Receptors" (2009). Neurobiology. 377.