GABAA receptor subtype selective loreclezole analogues

GABA-A receptors are channels present in the cell membrane of neurons. Upon binding of the messenger substance GABA they open and allow chloride-ions to flow into the cell. This electrical chloride current is an essential process because it leads to the reduction of excitability of the neuron and thus to a decrease of signalling between neurons. In mammals, GABA-A receptors are formed of five subunits (the most common are alpha, beta and gamma subunits) that can be combined differently. Importantly, GABA is not the only substance that binds to GABA-A receptors, but several clinically used drugs as sleeping aids, antiepileptics, antipanic medications or muscle relaxants do. These drugs bind to several known and unknown binding sites different from the GABA binding site. Recently, putative binding sites for loreclezole, an experimental anticonvulsant, were described to exist in a particular region of the GABA-A receptor. Four loreclezole analogues created by our group showed a clear preference for the subunit alpha6 over alpha1, meaning that these substances bound to the receptor and lead to an increased chloride current only when alpha6 subunits were present. During the past years the alpha6 subunit raised interest thanks to its highly specific expression pattern in the central and peripheral nervous system and its possible involvement in hyper-locomotion, pain perception, inflammation and depression. One of our aims is therefore the generation of alpha6 specific modulatory substances. Our modulators could so be used not only as tool compounds for the detection of alpha6 containing receptors and their distribution in the brain, but on a long term especially as drugs that might be suited to treat some neuropsychiatric conditions. Our second aim is to localize the exact binding site of our substances in the GABA-A receptor. Given that the alpha isoforms differ in only one amino acid of the candidate site, we hypothesize that this amino acid may be responsible for the observed subtype preference. The methods to be used comprise chemical construction of the computationally designed substances, mutations of the one amino acid thought to be responsible for the binding, and an electrophysiological technique to study the chloride currents and their modulation by our substances on the GABA receptors. The binding site with which loreclezole interacts has never been utilized to develop ligands with selectivity for any alpha isoform. Thus, our approach is completely novel and addresses a target of high scientific and medical interest.