Frances A. Bromidge

Identification of amino acid residues responsible for the α5 subunit binding selectivity of L‐655,708, a benzodiazepine binding site ligand at the GABAA receptor

L‐655,708 is a ligand for the benzodiazepine site of the γ‐aminobutyric acid type A (GABAA) receptor that exhibits a 100‐fold higher affinity for α5‐containing receptors compared with α1‐containing receptors. Molecular biology approaches have been used to determine which residues in the α5 subunit are responsible for this selectivity. Two amino acids have been identified, α5Thr208 and α5Ile215, each of which individually confer approximately 10‐fold binding selectivity for the ligand and which together account for the 100‐fold higher affinity of this ligand at α5‐containing receptors. L‐655,708 is a partial inverse agonist at the GABAA receptor which exhibited no functional selectivity between α1‐ and α5‐containing receptors and showed no change in efficacy at receptors containing α1 subunits where amino acids at both of the sites had been altered to their α5 counterparts (α1ΔSer205‐Thr,Val212‐Ile). In addition to determining the binding selectivity of L‐655,708, these amino acid residues also influence the binding affinities of a number of other benzodiazepine (BZ) site ligands. They are thus important elements of the BZ site of the GABAA receptor, and further delineate a region just N‐terminal to the first transmembrane domain of the receptor α subunit that contributes to this binding site.

GABAA α1 subunit knock-out mice do not show a hyperlocomotor response following amphetamine or cocaine treatment

The GABA(A) receptor system provides the major inhibitory control in the CNS, with the alpha 1 beta 2 gamma 2 subunit combination being the most abundant and widely distributed form of the receptor. The alpha1 subunit knock-out (alpha1 KO) mice had a surprisingly mild overt phenotype, despite having lost approximately 60% of all GABA(A) receptors. The alpha1 KO mice had normal spontaneous locomotor activity, but were more sensitive to the sedating/ataxic effects of diazepam than wildtype (WT) mice. Pharmacological modulation of dopamine and N-methyl-D-aspartate (NMDA) receptors also produced altered responses in alpha1 KO mice compared with WT mice. As expected, the NMDA receptor antagonist MK801, amphetamine and cocaine increased locomotor activity in WT mice. Although MK801 increased locomotor activity in alpha1 KO mice, amphetamine and cocaine induced stereotypy not hyperlocomotion. Binding studies showed no gross changes in the total number of D1, D2 or NMDA receptors. Furthermore, pre-pulse inhibition of acoustic startle and the effects of cocaine in conditioned place preference were similar in both alpha1 KO and WT mice, indicating selective rather that global changes in response to dopaminergic agents. These data demonstrate subtle changes in behaviours mediated by neurotransmitters other than GABA in alpha1 KO mice and suggest that compensation may have occurred beyond the GABAergic system.

N-(Indol-3-ylglyoxylyl)piperidines: high affinity agonists of human GABA-A receptors containing the α1 subunit

A new class of N-(indol-3-ylglyoxylyl)piperidines are high affinity agonists at the benzodiazepine binding site of human GABA-A receptor ion-channels, with modest selectivity for receptors containing the alpha1 subunit over alpha2 and alpha3. All three receptor subtypes discriminate substantially between the two enantiomers of the chiral ligand 10.