TY - JOUR
T1 - The influence of neuroactive steroid lipophilicity on GABAA receptor modulation
T2 - Evidence for a low-affinity interaction
AU - Chisari, Mariangela
AU - Eisenman, Lawrence N.
AU - Krishnan, Kathiresan
AU - Bandyopadhyaya, Achintya K.
AU - Wang, Cunde
AU - Taylor, Amanda
AU - Benz, Ann
AU - Covey, Douglas F.
AU - Zorumski, Charles F.
AU - Mennerick, Steven
PY - 2009/8
Y1 - 2009/8
N2 - Anesthetic steroids with actions at γ-aminobutyric acid type A receptors (GABAARs) may access transmembrane domain binding site(s) directly from the plasma cell membrane. Accordingly, the effective concentration in lipid phase and the ability of the steroid to meet pharmacophore requirements for activity will both contribute to observed steady-state potency. Furthermore, onset and offset of receptor effects may be rate limited by lipid partitioning. Here we show that several GABA-active steroids, including naturally occurring neurosteroids, of different lipophilicity differ in kinetics and potency at GABAARs. The hydrophobicity ranking predicted relative potency of GABAAR potentiation and predicted current offset kinetics. Kinetic offset differences among steroids were largely eliminated by γ-cyclodextrin, a scavenger of unbound steroid, suggesting that affinity differences among the analogues are dwarfed by the contributions of nonspecific accumulation. A 7-nitrobenz-2-oxa-1,3-diazole (NBD)-tagged fluorescent analogue of the low-lipophilicity alphaxalone (C17-NBD-alphaxalone) exhibited faster nonspecific accumulation and departitioning than those of a fluorescent analogue of the high-lipophilicity (3α,5α)-3-hydroxypregnan-20-one (C17-NBD-3α5αA). These differences were paralleled by differences in potentiation of GABAAR function. The enantiomer of C17-NBD-3α5αA, which does not satisfy pharmacophore requirements for steroid potentiation, exhibited identical fluorescence kinetics and distribution to C17-NBD-3α5αA, but was inactive at GABA ARs. Simple simulations supported our major findings, which suggest that neurosteroid binding affinity is low. Therefore both specific (e.g., fulfilling pharmacophore requirements) and nonspecific (e.g., lipid solubility) properties contribute to the potency and longevity of anesthetic steroid action.
AB - Anesthetic steroids with actions at γ-aminobutyric acid type A receptors (GABAARs) may access transmembrane domain binding site(s) directly from the plasma cell membrane. Accordingly, the effective concentration in lipid phase and the ability of the steroid to meet pharmacophore requirements for activity will both contribute to observed steady-state potency. Furthermore, onset and offset of receptor effects may be rate limited by lipid partitioning. Here we show that several GABA-active steroids, including naturally occurring neurosteroids, of different lipophilicity differ in kinetics and potency at GABAARs. The hydrophobicity ranking predicted relative potency of GABAAR potentiation and predicted current offset kinetics. Kinetic offset differences among steroids were largely eliminated by γ-cyclodextrin, a scavenger of unbound steroid, suggesting that affinity differences among the analogues are dwarfed by the contributions of nonspecific accumulation. A 7-nitrobenz-2-oxa-1,3-diazole (NBD)-tagged fluorescent analogue of the low-lipophilicity alphaxalone (C17-NBD-alphaxalone) exhibited faster nonspecific accumulation and departitioning than those of a fluorescent analogue of the high-lipophilicity (3α,5α)-3-hydroxypregnan-20-one (C17-NBD-3α5αA). These differences were paralleled by differences in potentiation of GABAAR function. The enantiomer of C17-NBD-3α5αA, which does not satisfy pharmacophore requirements for steroid potentiation, exhibited identical fluorescence kinetics and distribution to C17-NBD-3α5αA, but was inactive at GABA ARs. Simple simulations supported our major findings, which suggest that neurosteroid binding affinity is low. Therefore both specific (e.g., fulfilling pharmacophore requirements) and nonspecific (e.g., lipid solubility) properties contribute to the potency and longevity of anesthetic steroid action.
UR - http://www.scopus.com/inward/record.url?scp=68149160907&partnerID=8YFLogxK
U2 - 10.1152/jn.00346.2009
DO - 10.1152/jn.00346.2009
M3 - Article
C2 - 19553485
AN - SCOPUS:68149160907
SN - 0022-3077
VL - 102
SP - 1254
EP - 1264
JO - Journal of neurophysiology
JF - Journal of neurophysiology
IS - 2
ER -