Rescue of two trafficking-defective variants of the neuronal glycine transporter GlyT2 associated to hyperekplexia

Neuropharmacology. 2021 May 15:189:108543. doi: 10.1016/j.neuropharm.2021.108543. Epub 2021 Mar 29.


Hyperekplexia is a rare sensorimotor syndrome characterized by pathological startle reflex in response to unexpected trivial stimuli for which there is no specific treatment. Neonates suffer from hypertonia and are at high risk of sudden death due to apnea episodes. Mutations in the human SLC6A5 gene encoding the neuronal glycine transporter GlyT2 may disrupt the inhibitory glycinergic neurotransmission and cause a presynaptic form of the disease. The phenotype of missense mutations giving rise to protein misfolding but maintaining residual activity could be rescued by facilitating folding or intracellular trafficking. In this report, we characterized the trafficking properties of two mutants associated with hyperekplexia (A277T and Y707C, rat numbering). Transporter molecules were partially retained in the endoplasmic reticulum showing increased interaction with the endoplasmic reticulum chaperone calnexin. One transporter variant had export difficulties and increased ubiquitination levels, suggestive of enhanced endoplasmic reticulum-associated degradation. However, the two mutant transporters were amenable to correction by calnexin overexpression. Within the search for compounds capable of rescuing mutant phenotypes, we found that the arachidonic acid derivative N-arachidonoyl glycine can rescue the trafficking defects of the two variants in heterologous cells and rat brain cortical neurons. N-arachidonoyl glycine improves the endoplasmic reticulum output by reducing the interaction transporter/calnexin, increasing membrane expression and improving transport activity in a comparable way as the well-established chemical chaperone 4-phenyl-butyrate. This work identifies N-arachidonoyl glycine as a promising compound with potential for hyperekplexia therapy.

Keywords: 4-Phenylbutyric acid (PubChem CID: 4775); ALX1393 (o-[(2-benzyloxyphenyl-3-flurophenyl)methyl]-l-serine) (PubChem CID: 16078939); Bupropion hydrochloride (PubChem CID: 62884); Calnexin; Chemical chaperone; Glycine; Hyperekplexia; Ibogaine hydrochloride (PubChem CID: 197059); Mutation; N-arachidonoyl glycine (PubChem CID: 5283389); Transport.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Arachidonic Acids / pharmacology
  • Arachidonic Acids / therapeutic use*
  • COS Cells
  • Cells, Cultured
  • Chlorocebus aethiops
  • Female
  • Genetic Variation / drug effects
  • Genetic Variation / physiology*
  • Glycine / analogs & derivatives*
  • Glycine / pharmacology
  • Glycine / therapeutic use
  • Glycine Plasma Membrane Transport Proteins / genetics*
  • Glycine Plasma Membrane Transport Proteins / metabolism
  • Hyperekplexia / drug therapy
  • Hyperekplexia / genetics*
  • Hyperekplexia / metabolism
  • Mutation, Missense / drug effects
  • Mutation, Missense / physiology*
  • Neurons / drug effects
  • Neurons / physiology*
  • Protein Transport / drug effects
  • Protein Transport / physiology
  • Rats
  • Rats, Wistar


  • Arachidonic Acids
  • Glycine Plasma Membrane Transport Proteins
  • N-arachidonylglycine
  • Slc6a5 protein, rat
  • Glycine