Dissecting the role of diazepam-sensitive γ-aminobutyric acid type A receptors in defensive behavioral reactivity to mild threat

Moderate reductions in synaptic γ-aminobutyric acid_A receptors (GABA_ARs) have been associated with an enhanced defensive behavioral reactivity to mild threat, sensitive to diazepam. We here tested whether a deficit in α2 subunit-containing GABAergic synapses is sufficient to cause this anxiety-related phenotype and to prevent its attenuation by the benzodiazepine. Wild type (α2 +/+), heterozygous (α2 +/-) and homozygous (α2 -/-) knock-out littermates were tested in the free-choice exploratory (FCE) and the light/dark choice (LDC) paradigms. α2 -/- mice, double mutant α1H101Rα2 -/- and α3H126Rα2 -/- mice, which combine a lack of α2-GABA _ARs with point-mutated diazepam-insensitive either α1H101R or α3H126R-GABA_ARs, and double point-mutated α1H101Rα 2H101R and α1H101Rα3H126R mice were used to uncover the GABA _AR subtype(s) mediating the drug effects. Data show that in the FCE, α2 -/- mice exhibited more retractions (i.e. risk assessment) and longer latencies to first occurrence into the novel compartment and less transitions and time spent inside it in comparison to α2 +/- and α2 +/+ mice. In the LDC, α2 -/- mice visited and spent less time in the lit box and stayed longer in the tunnel than the other two groups. Minor differences were found between α2 +/- and α2 +/+ mice in the two paradigms. Diazepam (1.5 mg/kg per os) normalized retractions and latencies in the FCE in α2 -/- and α3H126Rα2 -/- mice, but not in α1H101Rα2 -/- mice. The same drug treatment failed to attenuate behavioral aversion in both paradigms in all mutants with impaired α2-GABA_AR function. These results reveal α2-containing GABA_ARs as key molecular determinants in the regulation of anxiety-related responses elicited by exposure to relative novelty and mild threat. In the absence of these receptors, diazepam through activation of α1-GABA_ARs remains effective in reducing risk assessment, but not behavioral aversion.