Mechanisms of reversible GABAA receptor plasticity after ethanol intoxication

The time-dependent effects of ethanol (EtOH) intoxication on GABA(A) receptor (GABA(A)R) composition and function were studied in rats. A cross-linking assay and Western blot analysis of microdissected CA1 area of hippocampal slices obtained 1 h after EtOH intoxication (5 g/kg, gavage), revealed decreases in the cell-surface fraction of alpha 4 and delta, but not alpha 1, alpha 5, or gamma 2 GABA(A)R subunits, without changes in their total content. This was accompanied (in CA1 neuron recordings) by decreased magnitude of the picrotoxin-sensitive tonic current (I-tonic), but not miniature IPSCs (mIPSCs), and by reduced enhancement of I-tonic by EtOH, but not by diazepam. By 48 h after EtOH dosing, cell-surface alpha 4 (80%) and gamma 2 (82%) subunit content increased, and cell-surface alpha 1 (-50%) and gamma (-79%) and overall content were decreased. This was paralleled by faster decay of mIPSCs, decreased diazepam enhancement of both mIPSCs and Itonic, and paradoxically increased mIPSC responsiveness to EtOH (10-100 mM). Sensitivity to isoflurane- or diazepam-induced loss of righting reflex was decreased at 12 and 24 h after EtOH intoxication, respectively, suggesting functional GABAAR tolerance. The plastic GABA(A)R changes were gradually and fully reversible by 2 weeks after single EtOH dosing, but unexplainably persisted long after withdrawal from chronic intermittent ethanol treatment, which leads to signs of alcohol dependence. Our data suggest that early tolerance to EtOH may result from excessive activation and subsequent internalization of alpha 4 beta delta extrasynaptic GABA(A)Rs. This leads to transcriptionally regulated increases in 4 and gamma 2 and decreases in alpha 1 subunits, with preferential insertion of the newly formed alpha 4 beta gamma 2 GABA(A)Rs at synapses.