Synthesis and receptor binding of 5-amino[3H]2methyl-3-isothiazolol ([3H]thiomuscimol), a specific GABAA agonist photoaffinity label

The synthesis of tritium labelled thiomuscimol (5-amino[3H]2methyl-3-isothiazolol) (7c), a specific and high-affinity agonist photoaffinity label for GABAA, receptors, is described. The synthesis of 7c is based on a procedure fo the preparation of 5-amino[2H]2methyl-3-isothiazolol (7b). Methyl 3-ethoxyisothiazole-5-carboxylate (3), synthesized from 3-hydroxyisothiazole-5-carboxamide (1) via the corresponding methyl ester (2), was reduced with sodium borotritide to give 3-ethoxy-5-hydroxy[3H]2methyl-3-isothiazole (4c). 3-Ethoxy-5-phtalimido[3H]2-methylisothiazole (6c), synthesized from the 5-chloro[3H]2methyl analogue (5c) of (4c), was deprotected by treatment with concentrated hydrochloric acid to give 7c with a specific radioactivity of 29 Ci/mmol. In pilot binding assays, 7c was shown to bind to membranes from rat forebrain in a saturable manner and with KD and Bmax values of 28 ± 6 nM and 50 ± 4 fmol/mg original tissue, respectively. (Author)