The neurosteroid 5β‐pregnan‐3α‐ol‐20‐one enhances actions of etomidate as a positive allosteric modulator of α1β2γ2L GABA_A receptors

Abstract

BACKGROUND AND PURPOSENeurosteroids potentiate responses of the GABAA receptor to the endogenous agonist GABA. Here, we examined the ability of neurosteroids to potentiate responses to the allosteric activators etomidate, pentobarbital and propofol. EXPERIMENTAL APPROACHElectrophysiological assays were conducted on rat α1β2γ2L GABAA receptors expressed in HEK 293 cells. The sedative activity of etomidate was studied in Xenopus tadpoles and mice. Effects of neurosteroids on etomidate-elicited inhibition of cortisol synthesis were determined in human adrenocortical cells. KEY RESULTSThe neurosteroid 5β-pregnan-3α-ol-20-one (3α5βP) potentiated activation of GABAA receptors by GABA and allosteric activators. Co-application of 1 μM 3α5βP induced a leftward shift (almost 100-fold) of the whole-cell macroscopic concentration-response relationship for gating by etomidate. Co-application of 100 nM 3α5βP reduced the EC50 for potentiation by etomidate of currents elicited by 0.5 μM GABA by about three-fold. In vivo, 3α5βP (1mg kg -1 ) reduced the dose of etomidate required to produce loss of righting in mice (ED50) by almost 10-fold. In tadpoles, the presence of 50 or 100 nM 3α5βP shifted the EC50 for loss of righting about three-or ten-fold respectively. Exposure to 3α5βP did not influence inhibition of cortisol synthesis by etomidate. CONCLUSIONS AND IMPLICATIONSPotentiating neurosteroids act similarly on orthosterically and allosterically activated GABAA receptors. Co-application of neurosteroids with etomidate can significantly reduce dosage requirements for the anaesthetic, and is a potentially beneficial combination to reduce undesired side effects. Abbreviations3α5αP, 5α-pregnan-3α-ol-20-one; 3α5βP, 5β-pregnan-3α-ol-20-one (pregnanolone); LRR, loss of righting reflex BJP British Journal of Pharmacology

Synaptic-Type α1β2γ2L GABAA Receptors Produce Large Persistent Currents in the Presence of Ambient GABA and Anesthetic Drugs

“…Interestingly, the addition of 10-1000 nM alphaxalone to the combination of 0.5 mM GABA 1 1 mM propofol did not lead to further enhancement of steady-state current. This is in contrast to our recent finding that neuroactive steroids can potentiate peak currents elicited by a combination of GABA and an allosteric activator (Li et al, 2014).…”

“…For example, we estimate that the half-maximal concentration of etomidate producing an increase of steady-state currents is 0.9 mM (Fig. 3A), whereas our recent study on the effect of etomidate on peak currents from a1b2g2L receptors revealed an EC 50 of 2 mM (Li et al, 2014). Similarly, while the EC 50 for potentiation of steady-state currents by propofol was 0.4 mM (Fig.…”

“…We previously showed that the application of a neuroactive steroid enhances peak responses from a1b2g2L GABA A receptors activated by allosteric activators or combinations of low GABA and an allosteric activator (Li et al, 2014). To test whether a steroid can enhance steady-state currents from receptors activated by GABA and an allosteric drug, we coapplied 10-1000 nM alphaxalone with 0.5 mM GABA 1 1 mM propofol.…”

Synthesis and pharmacological evaluation of neurosteroid photoaffinity ligands

“…This result indicates that these steroid anesthetics do not compete for the binding to the intersubunit sites we recently located at the extracellular end of the transmembrane domain at the β + –α − (etomidate 28 ) and at the α + – β − and γ + –β − subunit interfaces (barbiturate). 22 This result is also consistent with the earlier ones showing an enhancement of anesthetic potency of etomidate by neurosteroids 35 and of chromaffin cell‘s responses to natural neurosteroids in the presence of a variety of barbiturates. 36 Our results confirm that these sites (etomidate, barbiturate and neurosteroid) are nonidentical, but their occupancies are allosterically coupled.…”

New insights in the systemic and molecular underpinnings of general anesthetic actions mediated by γ-aminobutyric acid A receptors

“…[35] When administered together, neurosteroids and anesthetic agents can enhance GABA A receptor-mediated inhibition in a synergistic manner. [36] This finding suggests that the combined use of anesthetics and neurosteroids may significantly reduce dose requirements needed for clinical anesthesia, which in turn should reduce cardiovascular and respiratory side effects.…”