Two pools of Triton X-100-insoluble GABA(A) receptors are present in the brain, one associated to lipid rafts and another one to the post-synaptic GABAergic complex

J Neurochem. 2007 Aug;102(4):1329-45. doi: 10.1111/j.1471-4159.2007.04635.x.

Abstract

Rat forebrain synaptosomes were extracted with Triton X-100 at 4 degrees 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 gamma-aminobutyric acid type-A receptors (GABA(A)Rs) were identified: (i) a higher-density pool (rho = 1.10-1.15 mg/mL) of GABA(A)Rs that contains the gamma2 subunit (plus alpha and beta subunits) and that is associated to gephyrin and the GABAergic post-synaptic complex and (ii) a lower-density pool (rho = 1.06-1.09 mg/mL) of GABA(A)Rs associated to detergent-resistant membranes (DRMs) that contain alpha and beta subunits but not the gamma2 subunit. Some of these GABA(A)Rs contain the delta subunit. Two pools of GABA(A)Rs insoluble in Triton X-100 at 4 degrees C were also identified in cultured hippocampal neurons: (i) a GABA(A)R pool that forms clusters that co-localize with gephyrin and remains Triton X-100-insoluble after cholesterol depletion and (ii) a GABA(A)R pool that is diffusely distributed at the neuronal surface that can be induced to form GABA(A)R clusters by capping with an anti-alpha1 GABA(A)R subunit antibody and that becomes solubilized in Triton X-100 at 4 degrees C after cholesterol depletion. Thus, there is a pool of GABA(A)Rs associated to lipid rafts that is non-synaptic and that has a subunit composition different from that of the synaptic GABA(A)Rs. Some of the lipid raft-associated GABA(A)Rs might be involved in tonic inhibition.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Animals
  • Brain / cytology
  • Brain / metabolism*
  • Brain / ultrastructure
  • Cells, Cultured
  • Detergents / pharmacology
  • Embryo, Mammalian
  • Flunitrazepam / pharmacokinetics
  • GABA Modulators / pharmacokinetics
  • Membrane Microdomains / drug effects
  • Membrane Microdomains / metabolism*
  • Membrane Microdomains / ultrastructure
  • Membrane Proteins / metabolism
  • Microscopy, Electron, Transmission / methods
  • Neurons / cytology
  • Neurons / drug effects
  • Neurons / metabolism
  • Octoxynol / pharmacology
  • Radioligand Assay / methods
  • Rats
  • Rats, Sprague-Dawley
  • Receptors, GABA-A / classification*
  • Receptors, GABA-A / metabolism*
  • Saponins / pharmacology
  • Subcellular Fractions / metabolism
  • Subcellular Fractions / ultrastructure
  • Synaptosomes / drug effects
  • Synaptosomes / metabolism*
  • Synaptosomes / ultrastructure
  • gamma-Aminobutyric Acid / metabolism*

Substances

  • Detergents
  • GABA Modulators
  • Membrane Proteins
  • Receptors, GABA-A
  • Saponins
  • gamma-Aminobutyric Acid
  • Flunitrazepam
  • Octoxynol