Structure-Function Evaluation of Imidazopyridine Derivatives Selective for δ-Subunit-Containing γ-Aminobutyric Acid Type A (GABAA) Receptors.

δ-Selective compounds 1 and 2 (DS1, compound 22; DS2, compound 16) were introduced as functionally selective modulators of δ-containing GABA type A receptors (GABAAR). In our hands, [3H]EBOB-binding experiments with recombinant GABAAR and compound 22 showed no proof of δ-selectivity, although there was a minimally higher preference for the α4ß3δ and α6ß2/3δ receptors with respect to potency. In order to delineate the structural determinants of δ preferences, we synthesized 25 derivatives of DS1 and DS2, and investigated their structure-activity relationships (SAR). Four of our derivatives showed selectivity for α6ß3δ receptors (29, 38, 39, and 41). For all of them, the major factors that distinguished them from compound 22 were variations at the para-positions of their benzamide groups. However, two compounds (29 and 39), when tested in the presence of GABA, revealed effects at several additional GABAAR. The newly synthesized compounds will still serve as useful tools to investigate α6ß3δ receptors.

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

The betaine/γ-aminobutyric acid (GABA) transporter 1 (BGT1) is one of the four GABA transporters (GATs) involved in the termination of GABAergic neurotransmission. 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-carboxylic acid (ATPCA), a selective substrate for BGT1 over GAT1/GAT3, but also an agonist for GABAA receptors. With the aim of providing new functional insight into BGT1, we here present the synthesis and pharmacological characterization of the tritiated analogue, [3H]ATPCA. Using traditional uptake assays at recombinant transporters expressed in cell lines, [3H]ATPCA displayed a striking selectivity for BGT1 among the four GATs ( Km and Vmax values of 21 µM and 3.6 nmol ATPCA/(min × 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 [3H]ATPCA uptake in neurons, but not in astrocytes. The neuronal uptake could be inhibited by GABA, ATPCA, and a noncompetitive BGT1-selective inhibitor, indicating functional BGT1 in neurons. In conclusion, we report [3H]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.

Glial GABA Transporters as Modulators of Inhibitory Signalling in Epilepsy and Stroke.

Imbalances in GABA-mediated tonic inhibition are involved in several pathophysiological conditions. A classical way of controlling tonic inhibition is through pharmacological intervention with extrasynaptic GABAA receptors that sense ambient GABA and mediate a persistent GABAergic conductance. An increase in tonic inhibition may, however, also be obtained indirectly by inhibiting glial GABA transporters (GATs). These are sodium-coupled membrane transport proteins that normally act to terminate GABA neurotransmitter action by taking up GABA into surrounding astrocytes. The aim of the review is to provide an overview of glial GATs in regulating tonic inhibition, especially in epilepsy and stroke. This entails a comprehensive summary of changes known to occur in GAT expression levels and signalling following epileptic and ischemic insults. Further, we discuss the accumulating pharmacological evidence for targeting GATs in these diseases.