Stoichiometry of δ subunit containing GABA A receptors

Background and Purpose Although the stoichiometry of the major synaptic αβγ subunit‐containing GABA A receptors has consensus support for 2α:2β:1γ, a clear view of the stoichiometry of extrasynaptic receptors containing δ subunits has remained elusive. Here we examine the subunit stoichiometry of recombinant α4β3δ receptors using a reporter mutation and a functional electrophysiological approach. Experimental Approach Using site‐directed mutagenesis, we inserted a highly characterized 9′ serine to leucine mutation into the second transmembrane ( M 2) region of α4, β3 and δ subunits that increases receptor sensitivity to GABA . Whole‐cell, GABA ‐activated currents were recorded from HEK ‐293 cells co‐expressing different combinations of wild‐type ( WT ) and/or mutant α4( L 297 S ), β3( L 284 S ) and δ( L 288 S ) subunits. Key Results Recombinant receptors containing one or more mutant subunits showed increased GABA sensitivity relative to WT receptors by approximately fourfold, independent of the subunit class (α, β or δ) carrying the mutation. GABA dose–response curves of cells co‐expressing WT subunits with their respective L 9′ S mutants exhibited multiple components, with the number of discernible components enabling a subunit stoichiometry of 2α, 2β and 1δ to be deduced for α4β3δ receptors. Varying the cDNA transfection ratio by 10‐fold had no significant effect on the number of incorporated δ subunits. Conclusions and Implications Subunit stoichiometry is an important determinant of GABA A receptor function and pharmacology, and δ subunit‐containing receptors are important mediators of tonic inhibition in several brain regions. Here we demonstrate a preferred subunit stoichiometry for α4β3δ receptors of 2α, 2β and 1δ.