Allosteric modulation of a1b3g2 GABAA receptors by farnesol through the neurosteroid sites

In mammalian cells, all-trans farnesol, a 15-carbon isoprenol, is a product of the mevalonate pathway. It is the natural substrate of alcohol dehydrogenase and a substrate for CYP2E1, two enzymes implicated in ethanol metabolism. Studies have shown that farnesol is present in the human brain and inhibits voltage-gated Ca2 thorn channels at much lower con-centrations than ethanol. Here we show that farnesol modulates the activity of g-aminobutyric acid type A receptors (GABAARs), some of which also mediate the sedative activity of ethanol. Electrophysiology experiments performed in HEK cells expressing human a1b3g2 or a6b3g2 GABAARs revealed that farnesol increased chloride currents through positive allosteric modulation of these receptors and showed dependence on both the alcoholic functional group of farnesol and the length of the alkyl chain for activity. In silico studies using long-timescale unbiased all-atom molecular dynamics (MD) simulations of the human a1b3g2 GABAA receptors revealed that farnesol modulates the channel by directly binding to the transmembrane neurosteroid-binding site, after partitioning into the surrounding membrane and reaching the receptor by lateral diffusion. Channel activation by far-nesol was further characterized by several structural and dynamic variables, such as global twisting of the receptor's extracel-lular domain, tilting of the transmembrane M2 helices, radius, cross-sectional area, hydration status, and electrostatic potential of the channel pore. Our results expand the pharmacological activities of farnesol to yet another class of ion channels implicated in neurotransmission, thus providing a novel path for understanding and treatment of diseases involving GABAA receptor dysfunction. SIGNIFICANCE GABAARs are ligand-gated ion channels directly involved in inhibitory neurotransmission. GABAARs are the target of alcohol and numerous drugs used to treat CNS disorders. This study reveals, for the first time, the molecular recognition of GABAARs by farnesol, an endogenous alcohol present in the human brain and a product of the cholesterol synthesis pathway. We demonstrate that farnesol acts as an allosteric modulator of the receptors by directly binding to the transmembrane neurosteroid-binding site through a lipid path. The results provide novel insights into the allosteric communication affecting the affinity and binding modes of GABA and raise the possibility of functional and molecular crosstalk between GABA-mediated neurotransmission and the cholesterol synthesis pathway.