Abstracts

Rationale: In thalamus, GABAergic feedback inhibition from the reticular nuclei (nRT) to the ventrobasal nuclei (VB) contributes to aberrant thalamic oscillations in absence seizures. Heterozygous loss-of-function mutations in the GABA_A receptor ( GABA_AR) 1 subunit, and heterozygous 1 subunit deletion (Het ₁KO), cause absence seizures in humans and mice, respectively. Recently, we reported that Het ₁KO increased 3 subunit-containing GABA_ARs in the cortex. Because cortical neurons express both 1 and 3 subunits, this finding may have resulted from an increased ability of 3 subunit to incorporate in functional GABA_ARs due to the lower 1 subunit expression. However, in VB and nRT, 1 and 3 subunits are not expressed in the same cells; VB expresses 1 but not 3 and nRT expresses 3 but not 1. Here, we determined the effects of Het ₁KO on GABA_AR expression and physiology in the thalamus.Methods: Immunofluorescence microscopy determined the effects of Het ₁KO on 1 and 3 subunit expression in VB and nRT, respectively. Brain slice biotinylation and Western blot assays allowed quantification of total and cell surface expression of the 1 and 3 subunits. Patch clamp electrophysiology determined the consequences of Het ₁KO on GABAergic miniature inhibitory postsynaptic currents (mIPSC).Results: Immunofluorescence revealed that 1 subunit was not present in the nRT of wild type or Het ₁KO mice and that Het ₁KO reduced 1 subunit expression homogenously throughout the VB. Biotinylation and Western blot assays demonstrated that Het ₁KO reduced total 1 subunit expression to 62% that of wild type and, in contrast to its effects in the cortex, did not compensate by increasing its relative surface expression. Electrophysiological recordings showed that Het ₁KO reduced peak mIPSC amplitudes in VB (57% of wild type) and did not alter the time course of mIPSC kinetics. However, Het ₁KO did reduce mIPSC frequency (38% of wild type). Immunofluorescence demonstrated that 3 was not present in the VB of wild type or Het ₁KO mice and, unexpectedly, that Het ₁KO increased its expression in nRT. Biotinylation and Western blot assays showed that Het ₁KO increased the total expression of the 3 subunit to 172% of wild type and had an even greater effect on its surface expression (205%).Conclusions: In contrast to its effects in cortex, Het ₁KO caused uncompensated reductions of 1 subunit expression and peak mIPSC amplitudes in VB. Surprisingly, Het ₁KO also increased 3 subunit expression in nRT. Unlike cortex, nRT neurons do not express 1 and thus the increased 3 expression in nRT cannot result from increased permissive assembly of 3 into functional GABA_ARs due to fewer competing 1 subunits. Rather, Het ₁KO likely modifies thalamic neuronal circuitry and the altered activity of nRT neurons indirectly increases 3 subunit expression. Increased intra-nRT GABAergic inhibition via increased 3 subunit-containing GABA_A Rs may be a protective mechanism that partially compensates for VB disinhibition.