Relative Affinities of Positive and Negative Modulators of Heteromeric GABA(A) Receptors for Pseudo-Symmetric Intersubunit Binding Sites

Abstract

Type-A γ-aminobutyric acid receptors (GABAARs) are pentameric ligand-gated ion channels that are ubiquitous to the central nervous system (CNS) and critical for regulating neuronal excitability. These inhibitory receptors, gated by γ-aminobutyric acid (GABA), can be potentiated and also directly activated by certain exogenous and endogenous lipophilic small molecules. Among the various endogenous modulators are derivatives of cholesterol called neurosteroids; neurosteroids that positively modulate GABAARs can function as natural sedatives, anesthetics, anxiolytics and anti-convulsants. The mechanisms underlying modulation of GABAARs by neurosteroids and exogenous modulators like general anesthetics are still poorly understood, in part due to the location of likely quasi-symmetric interaction sites in the transmembrane domain of the receptor, as well as inconclusive results from mutagenesis experiments. In particular, the extent to which binding modes of positively and negatively modulating neurosteroids overlap is unknown. Here we use molecular dynamics simulations and the thermodynamically rigorous alchemical free energy perturbation technique to rank pseudo-symmetric intersubunit binding modes by affinity for the positively modulating neurosteroid allopregnanolone, the negatively modulating neurosteroid pregnenolone sulfate, and a novel analog with complex dose-response: triiodothyronine (thyroid hormone). Differential rankings of intersubunit sites for positive and negative modulating neurosteroids, triiodothyronine, and the general anesthetics propofol and sevoflurane, are interpreted in the context of their distinctive functional effects.