Two sets of chimeras between alphaxalone-sensitive GABA(A) receptor alpha2 or beta1 subunits and the alphaxalone-insensitive glycine receptor alpha1 subunit were constructed to determine the structural domains important for the modulatory actions of neuroactive steroids. These data suggest that the site of action for neurosteroids on GABA(A) receptors is not the same as that for volatile anesthetics and ethanol, but is on the N-terminal side of the middle of TM2.