Two pools of Triton X‐100‐insoluble GABAA receptors are present in the brain, one associated to lipid rafts and another one to the post‐synaptic GABAergic complex

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Abstract
Rat forebrain synaptosomes were extracted with Triton X‐100 at 4°C and the insoluble material, which is enriched in post‐synaptic densities (PSDs), was subjected to sedimentation on a continuous sucrose gradient. Two pools of Triton X‐100‐insoluble γ‐aminobutyric acid type‐A receptors (GABAARs) were identified: (i) a higher‐density pool (ρ = 1.10–1.15 mg/mL) of GABAARs that contains the γ2 subunit (plus α and β subunits) and that is associated to gephyrin and the GABAergic post‐synaptic complex and (ii) a lower‐density pool (ρ = 1.06–1.09 mg/mL) of GABAARs associated to detergent‐resistant membranes (DRMs) that contain α and β subunits but not the γ2 subunit. Some of these GABAARs contain the δ subunit. Two pools of GABAARs insoluble in Triton X‐100 at 4°C were also identified in cultured hippocampal neurons: (i) a GABAAR pool that forms clusters that co‐localize with gephyrin and remains Triton X‐100‐insoluble after cholesterol depletion and (ii) a GABAAR pool that is diffusely distributed at the neuronal surface that can be induced to form GABAAR clusters by capping with an anti‐α1 GABAAR subunit antibody and that becomes solubilized in Triton X‐100 at 4°C after cholesterol depletion. Thus, there is a pool of GABAARs associated to lipid rafts that is non‐synaptic and that has a subunit composition different from that of the synaptic GABAARs. Some of the lipid raft‐associated GABAARs might be involved in tonic inhibition.