Journal of Neuroscience, Vol 8, 615-622, Copyright © 1988 by Society for Neuroscience
Monoclonal antibodies and conventional antisera to the GABAA receptor/benzodiazepine receptor/Cl- channel complex
J Vitorica, D Park, G Chin and AL de Blas
Department of Neurobiology and Behavior, State University of New York, Stony Brook 11794.
Monoclonal antibodies (mAbs) and conventional antisera were raised to the
affinity-purified GABAA receptor/benzodiazepine receptor/Cl- channel
complex. The antibodies immunoprecipitated the affinity- purified complex
in Triton X-100 and also reacted with the complex in a solid-phase
radioimmunoassay. Immunoblots indicated that the mAb 62-3G1 reacted with
the 57,000 Mr peptide subunit of the affinity-purified complex, while the
antisera mainly reacted with the 51,000 Mr peptide subunit. The mAbs and
the antisera also immunoprecipitated the GABAA receptor/benzodiazepine
receptor/Cl- channel complex after being solubilized from cerebral cortex
membranes by the zwitterionic detergent CHAPS. The immunoprecipitated
complex bound 3H-muscimol, 3H- flunitrazepam (FNZ) and
35S-t-butylbicyclophosphorothionate (TBPS). The 3H-FNZ binding was
stimulated by GABA, indicating that the functional interactions among the
immunoprecipitated components of the complex were preserved. The mAb 62-3G1
also recognized the 57,000 Mr peptide in immunoblots with crude brain
membranes. Immunocytochemistry experiments showed that the binding of both
the mAb 62-3G1 and 3H-muscimol colocalized throughout the brain. The
results suggest that (1) the 57,000 Mr peptide is the muscimol (GABAA
receptor agonist) binding peptide of the complex, and (2) in the cerebral
cortex, most of the GABAA receptors (GABARs), benzodiazepine receptors
(BDZRs), and Cl- channels are physically coupled to one another.
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