Diazepam-insensitive GABA A receptors in rat cerebellum and thalamus

Abstract

There major populations of GABA A receptor binding sites are present in cerebellar membranes: diazepam-sensitive [ 3H]Ro15-4513 binding sites, diazepam-insensitive [ 3H]Ro15-4513 binding sites and high-affinity [ 3H]muscimol binding sites. All three populations contain a β subunit as shown by immunoprecipitation with antibodies that recognize all β subunits. The β3 subtype of β subunit is contained in all three populations, but only a similar low fraction (< 20%) in each. Thus, the majority contain β subunits other than β3 (β2 and β1) and β3 subunits are not selectively associated with nor lacking in any of the three binding populations. Antibodies to the γ2 subunit precipitated similar fractions of [ 3H]Ro15-4513, [ 3H]flunitrazepam and [ 3H]muscimol binding sites, showing that γ2 subunits are present in high-affinity muscimol binding isoforms, as well as a significant fraction of the diazepam-insensitive [ 3H]Ro15-4513 binding sites. Under conditions that identify the 56 kDa α6 subunit on SDS-PAGE as the diazepam-insensitive site of [ 3H]Ro15-4513 binding in cerebellum, no polypeptide showing diazepam-insensitive binding of [ 3H]Ro15-4513 could be photoaffinity-labeled in rat thalamus. These results suggest that α4 subunits in the thalamus participate primarily in subunit combinations which bind muscimol but not any benzodiazepine site ligands.