Poster Presentation The 48th Lorne Conference on Protein Structure and Function 2023

Novel inhibitors of chloride conductance in human excitatory amino acid transporter 1 (#323)

Amy Cheng 1 , Renae Ryan 1
  1. School of Medical Sciences, Faculty of Medicine , University of Sydney, Sydney, NSW, Australia

Excitatory amino acid transporters (EAATs) clear glutamate from synapses into nearby neurons and astrocytes. By coupling substrate transport to that of Na+, H+ and K+, EAATs rapidly lower extracellular glutamate concentrations, terminating neurotransmission. In addition, EAATs mediate an uncoupled Cl- conductance during the substrate transport cycle [1]. Increasing evidence point to the physiological importance of these dual functions; in particular, dysfunction of the Cl- channel in human EAAT1 is associated with the neurological disease Episodic Ataxia type 6 (EA6). Site-directed mutagenesis including characterisation of EA6-linked mutations has provided much insight into the molecular determinants of the Cl- conductance [2, 3]. However, there are no pharmacological modulators that can selectively and reversibly inhibit Cl- conductance. The structure of GltPh, an archaeal homologue of EAATs, in the Cl--conducting state by recently been determined [4]. This enabled the discovery of active compounds targeting the Cl--conducting state using ultra-large scale virtual screening via OpenBench. Here we investigate the effects of hits from an in silico screen on the dual functions of human EAAT1. We have identified compounds that completely blocked Cl- conductance without significantly altering glutamate transport, a mode of action distinct from existing inhibitors. These compounds will be used to better understand the interplay between the dual function of EAATs and serve as scaffolds for novel series of modulators.

  1. Fairman WA, Vandenberg RJ, Arriza JL, Kavanaugh MP, Amara SG. An excitatory amino-acid transporter with properties of a ligand-gated chloride channel. Nature. 1995;375.
  2. Ryan RM, Mitrovic AD, Vandenberg RJ. The chloride permeation pathway of a glutamate transporter and its proximity to the glutamate translocation pathway. J Biol Chem. 2004;279.
  3. Wu Q, Akhter A, Pant S, Cho E, Zhu JX, Garner A, et al. Ataxia-linked SLC1A3 mutations alter EAAT1 chloride channel activity and glial regulation of CNS function. J Clin Invest. 2022;132(7).
  4. Chen I, Pant S, Wu Q, Cater RJ, Sobti M, Vandenberg RJ, et al. Glutamate transporters have a chloride channel with two hydrophobic gates. Nature. 2021;591(7849):327-31.