Negative allosteric modulation of wild-type and mutant AMPA receptors by GYKI 53655.
Benzothiadiazides such as cyclothiazide potentiate alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptor responses, whereas 2,3-benzodiazepines such as 1-(4-aminophenyl)-3-methylcarbamyl-4-methyl-7,8-methylenedioxy-3,4 -dihydro- 5H-2,3-benzodiazepine (GYKI 53655) act as noncompetitive antagonists; both drugs act through allosteric modulation. Controversy exists as to whether cyclothiazide and GYKI 53655 act at a common site. Recent mutational analysis has led to the identification of a serine residue in flip splice variants that is critical for directing the interaction of cyclothiazide with AMPA receptors. We tested whether the mutation of this residue to glutamine, which abolishes potentiation by cyclothiazide, can in addition block antagonism by 2,3-benzodiazepines, as would be predicted for action at a common site. We found that the S to Q mutation does not alter antagonism by 2,3-benzodiazepines, suggesting that the molecular determinants directing the interaction between GYKI 53655 and AMPA receptors are not identical to those controlling sensitivity to cyclothiazide. Additional support for this was obtained from analysis of the responses of AMPA receptor flip/flop splice variants, which, despite differences in equilibrium desensitization and sensitivity to cyclothiazide, show only small differences in sensitivity to 2,3-benzodiazepines. Furthermore, introduction of the flip exon from GluRA into GluR6, conferred sensitivity to cyclothiazide but did not increase sensitivity to 2,3-benzodiazepines. Of interest, experiments with native AMPA receptors generated from hippocampal and forebrain poly(A)+ mRNA revealed greater sensitivity to 2,3-benzodiazepines than receptors generated by expression of recombinant AMPA receptors, possibly indicating the existence of an unidentified accessory protein or novel receptor subunit.
Partin
K M
KM
Laboratory of Cellular and Molecular Neurophysiology, National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, Maryland 20892-4495, USA.
Mayer
M L
ML
D000494:Allosteric Regulation; D000595:Amino Acid Sequence; D000818:Animals; D001569:Benzodiazepines / Q000494:pharmacology*; D001581:Benzothiadiazines / Q000494:pharmacology; D018691:Excitatory Amino Acid Antagonists / Q000494:pharmacology*; D008969:Molecular Sequence Data; D016548:Prosencephalon / Q000378:metabolism; D012326:RNA Splicing; D012333:RNA, Messenger / Q000235:genetics; D051381:Rats; D018091:Receptors, AMPA / Q000037:antagonists & inhibitors* / Q000235:genetics / Q000378:metabolism; D011994:Recombinant Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D014982:Xenopus laevis
D001581:Benzothiadiazines; D018691:Excitatory Amino Acid Antagonists; D012333:RNA, Messenger; D018091:Receptors, AMPA; D011994:Recombinant Proteins; D001569:Benzodiazepines; C079692:GYKI 53655; C004639:cyclothiazide