Neurosteroid Modulation of GABAA Receptor Function by Independent Action at Multiple Specific Binding Sites

Abstract

Neurosteroids are endogenous modulators of GABA_A receptors that mediate anxiety, pain, mood and arousal. The 3-hydroxyl epimers, allopregnanolone (3α-OH) and epiallopregnanolone (3β-OH) are both prevalent in the mammalian brain and produce opposite effects on GABA_A receptor function, acting as positive and negative allosteric modulators, respectively. This Perspective provides a model to explain the actions of 3α-OH and 3β-OH neurosteroids. The model is based on evidence that the neurosteroid epimers bind to an overlapping subset of specific sites on GABA_A receptors, with their net functional effect on channel gating being the sum of their independent effects at each site.

Keywords: GABA_A receptors; Neurosteroids; affinity labeling; desensitization; ion channels; structural biology.

Fig. (1)

Structure and actions of neurosteroids. Allopregnanolone (3α5α-P; top left) is an endogenous PAM neurosteroid characterized by a 3α-hydroxyl group. Its epimer, epi-allopregnanolone (3β5α-P; top right), is a NAM neurosteroid characterized by a 3β-hydroxyl group. The lower panels show simulated electrophysiological tracings of GABA_A-receptor currents constructed using a three-state (resting-active-desensitized) Monod-Wyman-Changeux model (simulation performed in ChanneLab™ ver. 2 software provided by Stephen F. Traynelis, Emory University School of Medicine). At low open probability (P_A), (left lower panel) 3α5α-P significantly potentiates the channel activation elicited by a low concentration of the orthosteric agonist GABA, whereas 3β5α-P (not shown) has no effect. In contrast, at high P_A (such as that produced by a saturating concentration of GABA) 3α5α-P weakly (lower middle panel) and 3β5α-P substantially desensitize the receptor, thus inhibiting the steady-state current.

Fig. (2)

Models of neurosteroid binding sites on α1β3 GABA_A receptors and the effects of their occupancy on channel gating. (Top left) Ribbon diagram of the interface between the β3(+) (salmon) and α1(-) (blue) subunits of a GABA_A receptor (based on pdb 6HUO) with the neurosteroid 3α5α-P in its preferred docking poses in: an intra-β-subunit site between β3(V290) on TM3 and β3(Y442) on TM4; a β3(+)/α1(-) intersubunit site between α1(Q242), α1(W246) on TM1 and β3(F301) on TM3 and; an intra-α-subunit site between α1(V227) on TM1 and α1(Y411 and N408) on TM4. (Top right) Hydrophobic surface representation of 3α5α-P docked to the α1β3 GABA_A receptor (brown most and turquoise least hydrophobic) illustrating that neurosteroids bind between the hydrophobic transmembrane α-helices on the receptor surface, interacting with both protein and annular lipid. (Lower panel) Cartoon illustrating that 3α5α-P (yellow steroid) occupancy of the β3(+)/α1(-) intersubunit site and/or the intra-α1-subunit site promotes channel activation. In contrast, occupancy of the intra-β3-subunit site by either 3α5α-P or 3β5α-P (blue steroid) and occupancy of the intra-α1-subunit site by 3β5α-P inhibits the receptor by promoting desensitization.