Immunochemical Characterization of the β2 Subunit of the GABAAReceptor

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Abstract
To date three beta subunits of the GABAA receptor have been identified in rat brain as a result of cDNA library screening. The beta 2 subunit has been reported to have a wide distribution in rat brain based on in situ hybridization studies quantifying beta 2 mRNA. To study the beta 2 subunit more directly, we have raised a polyclonal antibody to a synthetic peptide representing residues 315-334 of the intracellular loop of the beta 2 subunit. The antibody, which had been affinity-purified, recognized the beta 2 peptide but did not immunolabel homologous beta 1 and beta 3 subunit peptides, indicating that this antibody is specific for the beta 2 subunit of the receptor. In western blots of the purified receptor, the antibody recognized a major diffuse band of 54-58 kDa and exhibited minor labeling of lower-molecular-mass polypeptides. In western blots of cortex homogenate, the antibody exhibited nervous system-specific labeling of a 55-kDa band that comigrated with the 55-kDa band of the purified receptor. Quantitative immunolabeling of this 55-kDa polypeptide permitted direct determination of the relative amounts of the beta 2 subunit in different brain regions. The brainstem contained the highest relative specific activity of the beta 2 subunit, followed by the inferior colliculus, olfactory lobe, and cerebellum. Lower levels of immunolabeling were seen in hypothalamus, hippocampus, thalamus, and cortex.

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