title: Silent, fluorescent labeling of native neuronal receptors.
abstract: We have developed a minimally-perturbing strategy that enables labeling and subcellular visualization of endogenous dendritic receptors on live, wild-type neurons. Specifically, calcium-permeable non-NMDA glutamate receptors expressed in hippocampal neurons can be targeted with this novel synthetic tri-functional molecule. This ligand-directed probe was targeted towards AMPA receptors and bears an electrophilic group for covalent bond formation with an amino acid side chain on the extracellular side of the ion channel. This molecule was designed in such a way that the use-dependent, polyamine-based ligand accumulates the chemically-reactive group at the extracellular side of these polyamine-sensitive receptors, thereby allowing covalent bond formation between an electrophilic moiety on the nanoprobe and a nucleophilic amino acid sidechain on the receptor. Bioconjugation of this molecule results in a stable covalent bond between the nanoprobe and the target receptor. Subsequent photolysis of a portion of the nanoprobe may then be employed to effect ligand release allowing the receptor to re-enter the non-liganded state, all the while retaining the fluorescent beacon for visualization. This technology allows for rapid fluorescent labeling of native polyamine-sensitive receptors and further advances the field of fluorescent labeling of native biological molecules.
authors: Department of Chemistry, University of Massachusetts Amherst, Amherst, MA 01003, USA.
authors: Devaiah
authors: D
authors: Vytla
authors: Rosamund E
authors: RE
authors: Combs-Bachmann
authors: Amanda M
authors: AM
authors: Hussey
authors: Ismail
authors: I
authors: Hafez
authors: James J
authors: JJ
authors: Chambers
mesh_terms: D000818:Animals; D002478:Cells, Cultured; D005456:Fluorescent Dyes / Q000737:chemistry*; D006624:Hippocampus / Q000737:chemistry*; D006801:Humans; D015394:Molecular Structure; D051381:Rats; D017207:Rats, Sprague-Dawley; D018091:Receptors, AMPA / Q000032:analysis* / Q000737:chemistry
chemical_list: D005456:Fluorescent Dyes; D018091:Receptors, AMPA