Abstract
GABA(A) receptors mediate most inhibitory synaptic transmission in the brain of vertebrates. Following GABA binding and fast activation, these receptors undergo a slower desensitization, the conformational pathway of which remains largely elusive. To explore the mechanism of desensitization, we used concatemeric α1β2γ2 GABA(A) receptors to selectively introduce gain-of-desensitization mutations one subunit at a time. A library of twenty-six mutant combinations was generated and their bi-exponential macroscopic desensitization rates measured. Introducing mutations at the different subunits shows a strongly asymmetric pattern with a key contribution of the γ2 subunit, and combining mutations results in marked synergistic effects indicating a non-concerted mechanism. Kinetic modelling indeed suggests a pathway where subunits move independently, the desensitization of two subunits being required to occlude the pore. Our work thus hints towards a very diverse and labile conformational landscape during desensitization, with potential implications in physiology and pharmacology.