Shisa7-Dependent Regulation of GABA_A Receptor Single-Channel Gating Kinetics

Abstract

GABA_A receptors (GABA_ARs) mediate the majority of fast inhibitory transmission throughout the brain. Although it is widely known that pore-forming subunits critically determine receptor function, it is unclear whether their single-channel properties are modulated by GABA_AR-associated transmembrane proteins. We previously identified Shisa7 as a GABA_AR auxiliary subunit that modulates the trafficking, pharmacology, and deactivation properties of these receptors. However, whether Shisa7 also regulates GABA_AR single-channel properties has yet to be determined. Here, we performed single-channel recordings of α2β3γ2L GABA_ARs cotransfected with Shisa7 in HEK293T cells and found that while Shisa7 does not change channel slope conductance, it reduced the frequency of receptor openings. Importantly, Shisa7 modulates GABA_AR gating by decreasing the duration and open probability within bursts. Through kinetic analysis of individual dwell time components, activation modeling, and macroscopic simulations, we demonstrate that Shisa7 accelerates GABA_AR deactivation by governing the time spent between close and open states during gating. Together, our data provide a mechanistic basis for how Shisa7 controls GABA_AR gating and reveal for the first time that GABA_AR single-channel properties can be modulated by an auxiliary subunit. These findings shed light on processes that shape the temporal dynamics of GABAergic transmission.

SIGNIFICANCE STATEMENT Although GABA_A receptor (GABA_AR) single-channel properties are largely determined by pore-forming subunits, it remains unknown whether they are also controlled by GABA_AR-associated transmembrane proteins. Here, we show that Shisa7, a recently identified GABA_AR auxiliary subunit, modulates GABA_AR activation by altering single-channel burst kinetics. These results reveal that Shisa7 primarily decreases the duration and open probability of receptor burst activity during gating, leading to accelerated GABA_AR deactivation. These experiments are the first to assess the gating properties of GABA_ARs in the presence of an auxiliary subunit and provides a kinetic basis for how Shisa7 modifies temporal attributes of GABAergic transmission at the single-channel level.