Synthesis of GABAA receptor agonists and evaluation of their α-subunit selectivity and orientation in the GABA binding site

Drugs used to treat various disorders target GABAA receptors. To develop a subunit selective compounds, we synthesized 5-(4-piperidyl)-3-isoxazolol (4-PIOL) derivatives. The 3-isoxazolol moiety was substituted by 1,3,5-oxadiazol-2-one, 1,3,5-oxadiazol-2-thione, and substituted 1,2,4-triazol-3-ol heterocycles with modifications to the basic piperidine substituent as well as substituents without basic nitrogen. Compounds were screened by [3 H]muscimol binding and in patch-clamp experiments with heterologously expressed GABAA UikY2 receptors (i = 1-6). The effects of 5-aminomethyl-3H-[1,3,4]oxadiazol-2-one 5d were comparable to GABA for all a subunit isoforms. 5-piperidin-4-yl-3H-[1,3,4]oxadiazol-2-one 5a and 5 -piperidi n-4-yl -3H- [ 1, 3,4] oxadiazol - 2- thi one 6a were weak agonists at a,-, (X3-, and a5-containing receptors. When coapplied with GABA, they were antagonistic in a,-, a4-, and 0-6-containing receptors and potentiated U3-containing receptors. 6a protected GABA binding site cysteine-substitution mutants ajF64C and ajS68C from reacting with methanethiosulfonate-ethylsulfonate. 6a specifically covalently modified the ajR66C thiol, in the GABA binding site, through its oxadiazolethione sulfur. These results demonstrate the feasibility of synthesizing a subtype selective GABA mimetic drugs.