Pharmacological Characterization of [H-3]ATPCA as a Substrate for Studying the Functional Role of the Betaine/GABA Transporter 1 and the Creatine Transporter

0489983 - ÚOCHB 2019 RIV US eng J - Journal Article
Al-Khawaja, A. - Haugaard, A. S. - Marek, Aleš - Löffler, R. - Thiesen, L. - Santiveri, M. - Damgaard, M. - Bundgaard, C. - Frolund, B. - Wellendorph, P.
Pharmacological Characterization of [H-3]ATPCA as a Substrate for Studying the Functional Role of the Betaine/GABA Transporter 1 and the Creatine Transporter.
ACS Chemical Neuroscience. Roč. 9, č. 3 (2018), s. 545-554. ISSN 1948-7193. E-ISSN 1948-7193
Institutional support: RVO:61388963
Keywords : betaine/gamma-aminobutyric acid transporter 1 * BGT1 * GABA uptake assay * creatine transporter * ATPCA
OECD category: Biochemistry and molecular biology
Impact factor: 3.861, year: 2018

The betaine/gamma-aminobutyric acid (GABA) transporter 1 (BGT1) is one of the four GABA transporters (GATs) involved in the termination of GABAergic neuro-transmission. Although suggested to be implicated in seizure management, the exact functional importance of BGT1 in the brain is still elusive. This is partly owing to the lack of potent and selective pharmacological tool compounds that can be used to probe its function. We previously reported the identification of 2-amino-1,4,5,6-tetrahydropyrimidine-5-car-boxylic acid (ATPCA), a selective substrate for BGT1 over GAT1/GAT3, but also an agonist for GABA(A) receptors. With the aim of providing new functional insight into BGT1, we here present the synthesis and pharmacological characterization of the tritiated analogue, [H-3]ATPCA. Using traditional uptake assays at recombinant transporters expressed in cell lines, [H-3]ATPCA displayed a striking selectivity for BGT1 among the four GATs (K-m and V-max values of 21 mu M and 3.6 nmol ATPCA/(min x mg protein), respectively), but was also found to be a substrate for the creatine transporter (CreaT). In experiments with mouse cortical cell cultures, we observed a Na+-dependent [H-3]ATPCA uptake in neurons, but not in astrocytes. The neuronal uptake could be inhibited by GABA, ATPCA, and a noncompetitive BGTI-selective inhibitor, indicating functional BGT1 in neurons. In conclusion, we report [H-3]ATPCA as a novel radioactive substrate for both BGT1 and CreaT. The dual activity of the radioligand makes it most suitable for use in recombinant studies.
Permanent Link: http://hdl.handle.net/11104/0284271