TY - JOUR

T1 - The four human ¿-aminobutyric acid (GABA) transporters

T2 - pharmacological characterization and validation of a highly efficient screening assay

AU - Kvist, Trine

AU - Christiansen, Bolette

AU - Jensen, Anders Asbjørn

AU - Bräuner-Osborne, Hans

N1 - Keywords: Drug Evaluation, Preclinical; GABA Plasma Membrane Transport Proteins; High throughput screening assay; Humans; Tritium; gamma-Aminobutyric Acid; Subtype selectivity;

PY - 2009

Y1 - 2009

N2 - The neurotransmission mediated by gamma-aminobutyric acid (GABA) in the mammalian brain is terminated by a family of four GABA transporters (GATs). Inhibition of GATs is currently used in the treatment of epilepsy and these proteins are generally considered as important drug targets. In this study, we perform the first elaborate pharmacological characterization of all four human GAT subtypes. We conduct the experiments in parallel in a [3H]GABA uptake assay using 14 standard GAT substrates and inhibitors. This setup enables direct comparison of the absolute values of inhibitory activities of the compounds between the different GAT subtypes. The results are overall in agreement with data reported by other groups for the orthologous murine GATs. However, there do seem to be some minor variations among species. In contrast to the several subtype selective ligands identified for the GAT-1 subtype, no subtype selective ligands have been reported for the three remaining GATs. Given the potential therapeutic relevance of the individual GAT subtypes, a search for novel structures displaying selectivities for specific GAT subtypes is important. In this study, we validate our [3H]GABA uptake assay for use in high throughput screening. We find that the assay is categorized by high Z'-factors (Z' > 0.5) for all four GAT subtypes, demonstrating that the assay is excellent for a high throughput screen. This [3H]GABA uptake assay therefore enables future high through put screening of compound libraries at the four human GATs.

AB - The neurotransmission mediated by gamma-aminobutyric acid (GABA) in the mammalian brain is terminated by a family of four GABA transporters (GATs). Inhibition of GATs is currently used in the treatment of epilepsy and these proteins are generally considered as important drug targets. In this study, we perform the first elaborate pharmacological characterization of all four human GAT subtypes. We conduct the experiments in parallel in a [3H]GABA uptake assay using 14 standard GAT substrates and inhibitors. This setup enables direct comparison of the absolute values of inhibitory activities of the compounds between the different GAT subtypes. The results are overall in agreement with data reported by other groups for the orthologous murine GATs. However, there do seem to be some minor variations among species. In contrast to the several subtype selective ligands identified for the GAT-1 subtype, no subtype selective ligands have been reported for the three remaining GATs. Given the potential therapeutic relevance of the individual GAT subtypes, a search for novel structures displaying selectivities for specific GAT subtypes is important. In this study, we validate our [3H]GABA uptake assay for use in high throughput screening. We find that the assay is categorized by high Z'-factors (Z' > 0.5) for all four GAT subtypes, demonstrating that the assay is excellent for a high throughput screen. This [3H]GABA uptake assay therefore enables future high through put screening of compound libraries at the four human GATs.

KW - Former Faculty of Pharmaceutical Sciences

M3 - Journal article

C2 - 19275529

VL - 12

SP - 241

EP - 249

JO - Combinatorial Chemistry & High Throughput Screening

JF - Combinatorial Chemistry & High Throughput Screening

SN - 1386-2073

IS - 3

ER -