Molecular and Pharmacological Characterization of GABAAReceptor α1 Subunit Knockout Mice

Abstract

GABA_A receptors mediate fast inhibitory neurotransmission in the central nervous system (CNS), and approximately half of these receptors contain α1 subunits. GABA_A receptor α1 subunits are important for receptor assembly and specific pharmacological responses to benzodiazepines. Plasticity in GABA_A receptor α1 subunit expression is associated with changes in CNS excitability observed during normal brain development, in animal models of epilepsy, and upon withdrawal from alcohol and benzodiazepines. To examine the role of α1 subunit-containing GABA_A receptors in vivo, we characterized receptor subunit expression and pharmacological properties in cerebral cortex of knockout mice with a targeted deletion of the α1 subunit. The mice are viable but exhibit an intention tremor. Western blot analysis confirms the complete loss of α1 subunit peptide expression. Stable adaptations in the expression of several GABA_A receptor subunits are observed in the fifth to seventh generations, including decreased expression of β2/3 and γ2 subunits and increased expression of α2 and α3 subunits. There was no change in α4, α5, or δ subunit peptide levels in cerebral cortex. Knockout mice exhibit loss of over half of GABA_A receptors measured by [³H]muscimol, [³H]2-(3-carboxyl)-3-amino-6-(4-methoxyphenyl)-pyridazinium bromide ([³H]SR-95531), andt-butylbicyclophosphoro[³⁵S]thionate ([³⁵S]TBPS) binding. [³H]Ethyl-8-azido-5,6-dihydro-5-methyl-6-oxo-4H-imidazo[1,5-a][1,4]benzodiazepine-3-carboxylate ([³H]Ro15-4513) binding is reduced by variable amounts in different regions across brain. GABA_A receptor α1^−/− mice lose all high-affinity [³H]zolpidem binding and about half of [³H]flunitrazepam binding in the cerebral cortex. The potency and maximal efficacy of muscimol-stimulated³⁶Cl⁻ uptake in cerebral cortical synaptoneurosomes are reduced in α1^−/− mice. Furthermore, knockout mice exhibit increased bicuculline-induced seizure susceptibility compared with wild-type mice. These data emphasize the significance of α1 subunit expression and its involvement in the regulation of CNS excitability.