Cerebral GABAB receptor: proposed mechanisms of action and purification procedures.
The GABAB receptor in brain is one of the GABA receptor subtypes, and has been found to be negatively coupled to adenylate cyclase and phosphatidylinositide turnover. This receptor easily solubilizes from cerebral synaptic membrane preparations by 3-[(3-cholamidopropyl) dimethylammonio]-1-propanesulfonate (CHAPS) in the presence of asolectin. GABAB receptor solubilized from bovine cerebral cortex was purified using baclofen-coupled affinity beads (baclofen-coupled Toyopearl beads). Using these procedures, almost pure GABAB receptor (80 KDa protein) was obtained in the affinity eluate. A monoclonal antibody has been also raised against the purified GABAB receptor. The antibody recognized a protein of about 80 KDa in bovine brain synaptic membrane. Immunoabsorbent agarose beads conjugated with the antibody were able to remove more than 90% of the baclofen suppressive GABA binding activity in the solubilized synaptic membrane, and this system was found to be useful for the immunoaffinity column chromatographic separation of GABAB receptor. Preliminary studies of immunohistochemical visualization of GABAB receptor in the rat cerebellum suggested that this receptor may be exclusively localized at the presynaptic site of GABAergic neurons.
Department of Pharmacology, Kyoto Prefectural University of Medicine, Japan.
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K
Kuriyama
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H
Nakayasu
H
H
Mizutani
R
R
Narihara
T
T
Ichida
D000262:Adenylyl Cyclases / Q000378:metabolism; D000818:Animals; D001921:Brain / Q000378:metabolism*; D001923:Brain Chemistry; D019204:GTP-Binding Proteins / Q000502:physiology; D007118:Immunoassay; D011963:Receptors, GABA-A / Q000302:isolation & purification / Q000502:physiology*; D013570:Synaptic Membranes / Q000737:chemistry / Q000378:metabolism
D011963:Receptors, GABA-A; D019204:GTP-Binding Proteins; D000262:Adenylyl Cyclases