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Functional Reconstitution of α‐Amino‐3‐Hydroxy‐5‐Methylisoxazole‐4‐Propionate (AMPA) Receptors from Rat Brain
Author(s) -
Bahr Ben A.,
Vodyanoy Vitaly,
Hall Randy A.,
Suppiramaniam Vishnu,
Kessler Markus,
Sumikawa Katumi,
Lynch Gary
Publication year - 1992
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.1471-4159.1992.tb11038.x
Subject(s) - ampa receptor , forebrain , glutamate receptor , biochemistry , receptor , protein subunit , amino acid , biology , photoaffinity labeling , propionate , chemistry , neuroscience , central nervous system , gene
Glutamate receptors belonging to the subclass specifically activated by α‐amino‐3‐hydroxy‐5‐methylisoxazole‐4‐propionic acid (AMPA) were solubilized from rat forebrain membranes with Triton X‐100 and partially purified through a series of three chromatographic steps. Specific [ 3 H]AMPA binding increased 30–60‐fold during the isolation procedure. A protein band recognized by antibodies against specific amino acid sequences of the glutamate receptor‐A subunit was enriched with each purification step; the molecular mass of this band (105 kDa) corresponded to that of cloned AMPA receptor subunits. Photoaffinity labeling of forebrain membranes with 6‐cyano‐7‐[ 3 H]hitroquinoxaline‐2,3‐dione, a specific antagonist of the AMPA receptor, labeled a single band that comigrated with the immunolabeled protein. On reconstitution of the partially purified material into bilayer patches, singlechannel current fluctuations were elicited by 300 n M AMPA and blocked by 1 μ M 6,7‐dinitroquinoxaline‐2,3‐dione.