Enantioselectivity of pregnanolone-induced gamma-aminobutyric acid(A) receptor modulation and anesthesia

This study reports the actions of enantiomer pairs of anesthetic steroids 3 alpha 5 alpha P/ent-3 alpha 5 alpha P and 3 alpha 5 beta P/ent-3 alpha 5 b eta P as modulators of gamma-aminobutyric acid (GABA)(A) receptors and as a nesthetics. The enantiomers of structurally related 17-carbonitrile analogs also are examined. These studies were aimed at 1) determining whether the steroid recognition site could distinguish between molecules differing in s hape, but not other physical properties (enantioselectivity); 2) providing further insight into the structure-activity relationships of anesthetic ste roids; and 3) determining whether modulation of GABA(A) receptor function c orrelates with anesthetic potency for anesthetic steroid enantiomers. Stere oselective actions of the compounds were evaluated in four different bioass ays: 1) noncompetitive displacement of [S-35]t-butylbicyclophosphorothionat e from the picrotoxin site of GABA(A) receptors present in rat brain membra ne preparations; 2) modulation of GABA currents in cultured rat hippocampal neurons; 3) loss of righting reflex in tadpoles; and 4) loss of righting r eflex in mice. The data indicate that 5 alpha-reduced steroids, but not 5 b eta-reduced steroids, show a high degree of enantioselectivity/enantiospeci ficity in their actions as modulators of GABA(A) receptors and as anestheti cs. For all compounds studied, the effects on GABA(A) receptor function clo sely tracked with anesthetic effects. These data show that the anesthetic s teroid recognition site is capable of distinguishing enantiomers, suggestin g a protein-binding site of specific dimensions and shape. The results are compatible either with a structural model of the binding site that can acco mmodate 3 alpha 5 alpha P, 3 alpha 5 beta P, and ent-3 alpha 5 beta P, but not ent-3 alpha 5 alpha P, or with two different binding sites for steroid anesthetics.