A substituted anilino enaminone acts as a novel positive allosteric modulator of GABA(A) receptors in the mouse brain

• Published in: The Journal of pharmacology and experimental therapeutics 2011-03, Vol.336 (3), p.916-924
• Main Authors: Wang, Ze-Jun, Sun, Liqin, Jackson, Patrice L, Scott, Kenneth R, Heinbockel, Thomas
• Format: Article
• Language: English
• Subjects: Action Potentials - drug effects; Action Potentials - physiology; Allosteric Regulation - drug effects; Allosteric Regulation - physiology; Animals; Anticonvulsants - chemistry; Anticonvulsants - metabolism; Anticonvulsants - pharmacology; Brain - drug effects; Brain - metabolism; GABA Modulators - chemistry; GABA Modulators - metabolism; GABA Modulators - pharmacology; Mice; Mice, Inbred C57BL; Protein Binding - physiology; Receptors, GABA-A - physiology
• ISSN: 1521-0103
• DOI: 10.1124/jpet.110.173740

A small library of anilino enaminones was analyzed for potential anticonvulsant agents. We examined the effects of three anilino enaminones on neuronal activity of output neurons, mitral cells (MC), in an olfactory bulb brain slice preparation using whole-cell patch-clamp recording. These compounds are known to be effective in attenuating pentylenetetrazol-induced convulsions. Among the three compounds tested, 5-methyl-3-(4-trifluoromethoxy-phenylamino)-cyclohex-2-enone (KRS-5Me-4-OCF₃) showed potent inhibition of MC activity with an EC₅₀ of 24.5 μM. It hyperpolarized the membrane potential of MCs accompanied by suppression of spontaneous firing. Neither ionotropic glutamate receptor blockers nor a GABA(B) receptor blocker prevented the KRS-5Me-4-OCF(3)-evoked inhibitory effects. In the presence of GABA(A) receptor antagonists, KRS-5Me-4-OCF(3) completely failed to evoke inhibition of MC spiking activity, suggesting that KRS-5Me-4-OCF₃-induced inhibition may be mediated by direct action on GABA(A) receptors or indirect action through the elevation of tissue GABA levels. Neither vigabatrin (a selective GABA-T inhibitor) nor 1,2,5,6-tetrahydro-1-[2-[[(diphenylmethylene)amino]oxy]ethyl]-3-pyridinecarboxylic acid hydrochloride (NNC-711) (a selective inhibitor of GABA uptake by GABA transporter 1) eliminated the effect of KRS-5ME-4-OCF₃ on neuronal excitability, indicating that the inhibitory effect of the enaminone resulted from direct activation of GABA(A) receptors. The concentration-response curves for GABA are left-shifted by KRS-5Me-4-OCF₃, demonstrating that KRS-5Me-4-OCF₃ enhanced GABA affinity and acted as a positive allosteric modulator of GABA(A) receptors. The effect of KRS-5Me-4-OCF₃ was blocked by applying a benzodiazepine site antagonist, suggesting that KRS-5Me-4-OCF₃ binds at the classic benzodiazepine site to exert its pharmacological action. The results suggest clinical use of enaminones as anticonvulsants in seizures and as a potential anxiolytic in mental disorders.