title: First demonstration of a functional role for central nervous system betaine/{gamma}-aminobutyric acid transporter (mGAT2) based on synergistic anticonvulsant action among inhibitors of mGAT1 and mGAT2.
abstract: In a recent study, EF1502 [N-[4,4-bis(3-methyl-2-thienyl)-3-butenyl]-3-hydroxy-4-(methylamino)-4,5,6,7-tetrahydrobenzo [d]isoxazol-3-ol], which is an N-substituted analog of the GAT1-selective GABA uptake inhibitor exo-THPO (4-amino-4,5,6,7-tetrahydrobenzo[d]isoxazol-3-ol), was found to inhibit GABA transport mediated by both GAT1 and GAT2 in human embryonic kidney (HEK) cells expressing the mouse GABA transporters GAT1 to 4 (mGAT1-4). In the present study, EF1502 was found to possess a broad-spectrum anticonvulsant profile in animal models of generalized and partial epilepsy. When EF1502 was tested in combination with the clinically effective GAT1-selective inhibitor tiagabine [(R)-N-[4,4-bis(3-methyl-2-thienyl)-3-butenyl]nipecotic acid] or LU-32-176B [N-[4,4-bis(4-fluorophenyl)-butyl]-3-hydroxy-4-amino-4,5,6,7-tetrahydrobenzo[d]isoxazol-3-ol], another GAT1-selective N-substituted analog of exo-THPO, a synergistic rather than additive anticonvulsant interaction was observed in the Frings audiogenic seizure-susceptible mouse and the pentylenetetrazol seizure threshold test. In contrast, combination of the two mGAT1-selective inhibitors, tiagabine and LU-32-176B, resulted in only an additive anticonvulsant effect. Importantly, the combination of EF1502 and tiagabine did not result in a greater than additive effect in the rotarod behavioral impairment test. In subsequent in vitro studies conducted in HEK-293 cells expressing the cloned mouse GAT transporters mGAT1 and mGAT2, EF1502 was found to noncompetitively inhibit both mGAT1 and the betaine/GABA transporter mGAT2 (K(i) of 4 and 5 muM, respectively). Furthermore, in a GABA release study conducted in neocortical neurons, EF1502 did not act as a substrate for the GABA carrier. Collectively, these findings support a functional role for mGAT2 in the control of neuronal excitability and suggest a possible utility for mGAT2-selective inhibitors in the treatment of epilepsy.
authors.affiliation: University of Utah, Anticonvulsant Drug Development Program, Department of Pharmacology and Toxicology, 20 S. 2030 E., Room 408, Salt Lake City, UT 84112, USA. swhite@hsc.utah.edu
authors.forename: H Steve
authors.initials: HS
authors.lastname: White
authors.forename: William P
authors.initials: WP
authors.lastname: Watson
authors.forename: Suzanne L
authors.initials: SL
authors.lastname: Hansen
authors.forename: Scott
authors.initials: S
authors.lastname: Slough
authors.forename: Jens
authors.initials: J
authors.lastname: Perregaard
authors.forename: Alan
authors.initials: A
authors.lastname: Sarup
authors.forename: Tina
authors.initials: T
authors.lastname: Bolvig
authors.forename: Gitte
authors.initials: G
authors.lastname: Petersen
authors.forename: Orla M
authors.initials: OM
authors.lastname: Larsson
authors.forename: Rasmus P
authors.initials: RP
authors.lastname: Clausen
authors.forename: Bente
authors.initials: B
authors.lastname: Frølund
authors.forename: Erik
authors.initials: E
authors.lastname: Falch
authors.forename: Povl
authors.initials: P
authors.lastname: Krogsgaard-Larsen
authors.forename: Arne
authors.initials: A
authors.lastname: Schousboe
mesh_terms: D000161:Acoustic Stimulation; D000217:Acyltransferases / Q000037:antagonists & inhibitors* / Q000235:genetics / Q000502:physiology*; D000679:Amygdala / Q000502:physiology; D000818:Animals; D000927:Anticonvulsants / Q000494:pharmacology*; D001522:Behavior, Animal / Q000187:drug effects; D002478:Cells, Cultured; D003001:Cloning, Molecular; D003292:Convulsants; D004357:Drug Synergism; D004827:Epilepsy / Q000139:chemically induced / Q000517:prevention & control; D020195:Epilepsy, Reflex / Q000139:chemically induced / Q000517:prevention & control; D007696:Kindling, Neurologic; D008297:Male; D051379:Mice; D017351:N-Acetylglucosaminyltransferases; D009474:Neurons / Q000187:drug effects; D009557:Nipecotic Acids / Q000494:pharmacology; D010433:Pentylenetetrazole; D010862:Pilocarpine; D011596:Psychomotor Disorders / Q000139:chemically induced / Q000517:prevention & control; D011994:Recombinant Proteins / Q000037:antagonists & inhibitors / Q000235:genetics; D012640:Seizures / Q000139:chemically induced / Q000517:prevention & control; D000078308:Tiagabine; D005680:gamma-Aminobutyric Acid / Q000378:metabolism
chemical_list: D000927:Anticonvulsants; D003292:Convulsants; D009557:Nipecotic Acids; D011994:Recombinant Proteins; D010862:Pilocarpine; D005680:gamma-Aminobutyric Acid; D000217:Acyltransferases; C021802:2-acylglycerol O-acyltransferase; C459903:MGAT1 protein, mouse; D017351:N-Acetylglucosaminyltransferases; D010433:Pentylenetetrazole; D000078308:Tiagabine