Complex interactions between the steroid derivative RU 5135 and the GABAA-receptor complex

The modulation of [S-35]t-butylbicyclophosporothionate ([S-35]TBPS) binding was used to evaluate the actions of the steroid derivative RU 5135 at the gamma-aminobutyric acid, (GABA(A)) receptor complex. The inhibition of [S-35]TBPS binding by GABA in the presence of various concentrations of RU 5135 was consistent with the hypothesis that RU 5135 is a competitive antagonist at the GABA(A) receptor. Despite common structural features (i.e., 3-alpha-hydroxylated, 5-beta-reduced A ring) with GABA(A) receptor-active neurosteroids, RU 5135 did not appear to be competitive at the putative steroid site on the GABA(A) receptor-active, as demonstrated by Schild analysis of 5-alpha-pregnane-3-alpha-ol-20-one (3-alpha,5-alpha-P) modulation of [S-35]TBPS binding in the presence of different concentrations of RU 5135. On the other hand, the reduced potency of 3-alpha,5-alpha-P as an inhibitor of [S-35]TBPS binding in the presence of RU 5135, as well as blockade of 5-alpha-pregnane-3-alpha-20-alpha-diol (5-alpha-pregnanediol) inhibition of [S-35]TBPS binding by RU 5135 provide further-support for the GABA(A) receptor antagonist properties of RU 5135. Moreover, this amidine steroid was able to partially inhibit [S-35]TBPS binding independent of GABA with nanomolar potency; yet the mechanism by which this occurs remains to be determined.