title: Direct modulation of microtubule stability contributes to anthracene general anesthesia.
abstract: Recently, we identified 1-aminoanthracene as a fluorescent general anesthetic. To investigate the mechanism of action, a photoactive analogue, 1-azidoanthracene, was synthesized. Administration of 1-azidoanthracene to albino stage 40-47 tadpoles was found to immobilize animals upon near-UV irradiation of the forebrain region. The immobilization was often reversible, but it was characterized by a longer duration consistent with covalent attachment of the ligand to functionally important targets. IEF/SDS-PAGE examination of irradiated tadpole brain homogenate revealed labeled protein, identified by mass spectrometry as β-tubulin. In vitro assays with aminoanthracene-cross-linked tubulin indicated inhibition of microtubule polymerization, similar to colchicine. Tandem mass spectrometry confirmed anthracene binding near the colchicine site. Stage 40-47 tadpoles were also incubated 1 h with microtubule stabilizing agents, epothilone D or discodermolide, followed by dosing with 1-aminoanthracene. The effective concentration of 1-aminoanthracene required to immobilize the tadpoles was significantly increased in the presence of either microtubule stabilizing agent. Epothilone D similarly mitigated the effects of a clinical neurosteroid general anesthetic, allopregnanolone, believed to occupy the colchicine site in tubulin. We conclude that neuronal microtubules are "on-pathway" targets for anthracene general anesthetics and may also represent functional targets for some neurosteroid general anesthetics.
authors.affiliation: Department of Chemistry, University of Pennsylvania, 231 South 34th Street, Philadelphia, Pennsylvania 19104, USA.
authors.forename: Daniel J
authors.initials: DJ
authors.lastname: Emerson
authors.forename: Brian P
authors.initials: BP
authors.lastname: Weiser
authors.forename: John
authors.initials: J
authors.lastname: Psonis
authors.forename: Zhengzheng
authors.initials: Z
authors.lastname: Liao
authors.forename: Olena
authors.initials: O
authors.lastname: Taratula
authors.forename: Ashley
authors.initials: A
authors.lastname: Fiamengo
authors.forename: Xiaozhao
authors.initials: X
authors.lastname: Wang
authors.forename: Keizo
authors.initials: K
authors.lastname: Sugasawa
authors.forename: Amos B
authors.initials: AB
authors.lastname: Smith
authors.forename: Roderic G
authors.initials: RG
authors.lastname: Eckenhoff
authors.forename: Ivan J
authors.initials: IJ
authors.lastname: Dmochowski
mesh_terms: D000758:Anesthesia; D000818:Animals; D000873:Anthracenes / Q000008:administration & dosage / Q000737:chemistry / Q000494:pharmacology*; D001386:Azides / Q000008:administration & dosage / Q000737:chemistry / Q000494:pharmacology*; D007814:Larva / Q000187:drug effects; D008870:Microtubules / Q000187:drug effects* / Q000378:metabolism; D015394:Molecular Structure; D014466:Ultraviolet Rays; D014982:Xenopus laevis
chemical_list: C117941:1-azidoanthracene; D000873:Anthracenes; D001386:Azides; C049062:1-aminoanthracene