The Sulfated Steroids Pregnenolone Sulfate and Dehydroepiandrosterone Sulfate Inhibit the α1β3γ2L GABAA Receptor by Stabilizing a Novel Nonconducting State

Pierce, Spencer R.; Germann, Allison L.; Steinbach, Joseph H.

doi:10.1124/molpharm.121.000385

Cited by 16 publications

(15 citation statements)

References 42 publications

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“…These findings may reflect conditions related to cryo-EM sample preparation, favoring a desensitized-like structure not perfectly representative of the predominant physiological state 34 . Alternatively, these results may support a recent hypothesis based on detailed kinetic modeling of the α1β3γ2 GABA A receptor, where PS binding stabilizes a nonconducting state distinct from both open and desensitized 29 . Such a model would recapitulate several functional features we observe here, including increased apparent GABA affinity in the presence of PS and a transient increase in current amplitude upon PS washout.…”

Section: Discussionsupporting

confidence: 78%

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Divergent mechanisms of steroid inhibition in the human ρ1 GABA_Areceptor

Fan,

Cowgill,

Howard

et al. 2024

Preprint

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ρ-type γ-aminobutyric acid-A (GABAA) receptors are widely distributed in the retina and brain, and are potential drug targets for the treatment of visual, sleep and cognitive disorders. Endogenous neuroactive steroids including β-estradiol and pregnenolone sulfate negatively modulate the function of ρ1 GABAAreceptors, but their inhibitory mechanisms are not clear. By combining four new cryo-EM structures with electrophysiology and molecular dynamics simulations, we characterize binding sites and negative modulation mechanisms of β-estradiol and pregnenolone sulfate at the human ρ1 GABAAreceptor. β-estradiol binds in a pocket at the interface between extracellular and transmembrane domains, apparently specific to the ρ subfamily, and disturbs allosteric conformational transitions linking GABA binding to pore opening. In contrast, pregnenolone sulfate binds inside the pore to block ion permeation, with a preference for activated structures. These results illuminate contrasting mechanisms of ρ1 inhibition by two different neuroactive steroids, with potential implications for subtype-specific gating and pharmacological design.

show abstract

Section: Discussionsupporting

confidence: 78%

“…To explore alternative mechanisms of ρ1 inhibition, we then characterized the 3β-sulfated neurosteroid PS (Figure 3A), a negative modulator of this and several other GABA A -receptor subtypes 15,29 . Similar to previous reports 13 , 100 μM PS reduced ρ1-EM currents by roughly 35% (Figure 3B).…”

Section: Resultsmentioning

confidence: 99%

Divergent mechanisms of steroid inhibition in the human ρ1 GABA_Areceptor

Fan,

Cowgill,

Howard

et al. 2024

Preprint

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“…More details are available in our recent publications. 17,18 The current responses were amplified with an OC-725C (Warner Instruments) or Axoclamp 900A amplifier (Molecular Devices, San Jose, CA), digitized with a Digidata 1200 or 1320 series digitizer (Molecular Devices), and stored using Clampex (pClamp software package, Molecular Devices). The current traces were low-pass filtered at 10 Hz, digitized at 50 Hz, and analyzed using Clampfit (pClamp) to determine the amplitude of the response.…”

Section: ■ Methodsmentioning

confidence: 99%

“…All experiments were conducted at room temperature. More details are available in our recent publications. , …”

Section: Methodsmentioning

confidence: 99%

Comparison of Behavioral Effects of GABAergic Low- and High-Efficacy Neuroactive Steroids in the Zebrafish Larvae Assay

Germann,

Xu,

Covey

et al. 2024

ACS Chem. Neurosci.

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Activation of the GABA A receptor is associated with numerous behavioral end points ranging from anxiolysis to deep anesthesia. The specific behavioral effect of a GABAergic compound is considered to correlate with the degree of its functional effect on the receptor. Here, we tested the hypothesis that a low-efficacy allosteric potentiator of the GABA A receptor may act, due to a ceiling effect, as a sedative with reduced and limited action. We synthesized a derivative, named (3α,5β)-20methyl-pregnane-3,20-diol (KK-235), of the GABAergic neurosteroid 5β-pregnane-3α,20α-diol. Using electrophysiology, we showed that KK-235 is a low-efficacy potentiator of the synaptictype α1β2γ2L GABA A receptor. In the zebrafish larvae behavioral assay, KK-235 was found to only partially block the inverted photomotor response (PMR) and to weakly reduce swimming behavior, whereas the high-efficacy GABAergic steroid (3α,5α,17β)-3-hydroxyandrostane-17-carbonitrile (ACN) fully blocked PMR and spontaneous swimming. Coapplication of KK-235 reduced the potentiating effect of ACN in an electrophysiological assay and dampened its sedative effect in behavioral experiments. We propose that low-efficacy GABAergic potentiators may be useful as sedatives with limited action.

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“…To explore alternative mechanisms of ρ1 inhibition, we then characterized the 3β-sulfated neurosteroid PS (Figure 3A), a negative modulator of this and several other GABAA-receptor subtypes 15,29 . Similar to previous reports 13 , 100 μM PS reduced ρ1-EM currents by roughly 35% (Figure 3B).…”

Section: Ps Blocks the Activated ρ1-em Porementioning

confidence: 99%

Divergent mechanisms of steroid inhibition in the human ρ1 GABAA receptor

Lindahl,

Fan,

Cowgill

et al. 2024

Preprint

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ρ-type γ-aminobutyric acid-A (GABAA) receptors are widely distributed in the retina and brain, and are potential drug targets for the treatment of visual, sleep and cognitive disorders. Endogenous neuroactive steroids including β-estradiol and pregnenolone sulfate negatively modulate the function of ρ1 GABAA receptors, but their inhibitory mechanisms are not clear. By combining four new cryo-EM structures with electrophysiology and molecular dynamics simulations, we characterize binding sites and negative modulation mechanisms of β-estradiol and pregnenolone sulfate at the human ρ1 GABAA receptor. β-estradiol binds in a pocket at the interface between extracellular and transmembrane domains, apparently specific to the ρ subfamily, and disturbs allosteric conformational transitions linking GABA binding to pore opening. In contrast, pregnenolone sulfate binds inside the pore to block ion permeation, with a preference for activated structures. These results illuminate contrasting mechanisms of ρ1 inhibition by two different neuroactive steroids, with potential implications for subtype-specific gating and pharmacological design.

show abstract

The Sulfated Steroids Pregnenolone Sulfate and Dehydroepiandrosterone Sulfate Inhibit the α1β3γ2L GABA_A Receptor by Stabilizing a Novel Nonconducting State

Abstract: tetradecahydro-1H-cyclopenta[a]phenanthren-1-one sulfate (dehydroepiandrosterone sulfate); P A , probability of being in the active state; PS, (1S,3aS,3bS,7S,9aR,9bS,11aS)-1-acetyl-9a,11a-dimethyl-2,

Cited by 16 publications

References 42 publications

Divergent mechanisms of steroid inhibition in the human ρ1 GABA_Areceptor

Divergent mechanisms of steroid inhibition in the human ρ1 GABA_Areceptor

Comparison of Behavioral Effects of GABAergic Low- and High-Efficacy Neuroactive Steroids in the Zebrafish Larvae Assay

Divergent mechanisms of steroid inhibition in the human ρ1 GABAA receptor

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The Sulfated Steroids Pregnenolone Sulfate and Dehydroepiandrosterone Sulfate Inhibit the α1β3γ2L GABAA Receptor by Stabilizing a Novel Nonconducting State

Abstract: tetradecahydro-1H-cyclopenta[a]phenanthren-1-one sulfate (dehydroepiandrosterone sulfate); P A , probability of being in the active state; PS, (1S,3aS,3bS,7S,9aR,9bS,11aS)-1-acetyl-9a,11a-dimethyl-2,

Cited by 16 publications

References 42 publications

Divergent mechanisms of steroid inhibition in the human ρ1 GABAAreceptor

Divergent mechanisms of steroid inhibition in the human ρ1 GABAAreceptor

Comparison of Behavioral Effects of GABAergic Low- and High-Efficacy Neuroactive Steroids in the Zebrafish Larvae Assay

Divergent mechanisms of steroid inhibition in the human ρ1 GABAA receptor

Contact Info

Product

Resources

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The Sulfated Steroids Pregnenolone Sulfate and Dehydroepiandrosterone Sulfate Inhibit the α1β3γ2L GABA_A Receptor by Stabilizing a Novel Nonconducting State

Divergent mechanisms of steroid inhibition in the human ρ1 GABA_Areceptor

Divergent mechanisms of steroid inhibition in the human ρ1 GABA_Areceptor