Benzodiazepines modulate GABAA receptors by reducing a gamma-subunit-mediated inhibition of GABA sensitivity
BMC Pharmacology volume 9, Article number: A23 (2009)
Heterologous expression of α1, β2 and γ2S(γ1) subunits produces a mixed population of GABAA receptors containing α1β2 or α1β2γ2S(γ1) subunits. GABA sensitivity (lower in receptors containing γ1 or γ2S subunits) and the potentiation of GABA-activated chloride currents (IGABA) by benzodiazepines (BZDs) are dependent on γ2S(γ1) incorporation . A variable γ subunit incorporation may affect the estimation of IGABA potentiation by BZDs. We propose an approach for estimation of BZD efficiency that accounts for a mixed population of α1β2 and α1β2γ2S(γ1) receptors.
We investigated the relation between GABA sensitivity (EC50) and BZD modulation by analyzing triazolam-, clotiazepam- and midazolam-induced potentiation of IGABA in Xenopus oocytes under two-microelectrode voltage clamp.
Plotting EC50 versus BZD-induced shifts of GABA concentration-response curves (ΔEC50(BZD)) of oocytes injected with different amounts of α1, β2 and γ2S(γ1) cRNA (1:1:1-1:1:10) revealed a linear regression between γ2S(γ1)-mediated reduction of GABA sensitivity (EC50) and ΔEC50(BZD). The slope factors of the regression were always higher for oocytes expressing α1β2γ1 subunit receptors (triazolam: 1.8 ± 0.1; clotiazepam: 1.6 ± 0.1; midazolam: 2.3 ± 0.2) than for oocytes expressing α1β2γ2S receptors (triazolam: 1.4 ± 0.1; clotiazepam: 1.4 ± 0.1; midazolam: 1.3 ± 0.1). Mutant GABAA receptors (α1β2-R207Cγ2S) with lower GABA sensitivity showed higher drug efficiencies (slope factors: triazolam: 1.1 ± 0.1; clotiazepam: 1.1 ± 0.1; midazolam: 1.2 ± 0.1) whereas higher GABA sensitivity of α1-L263Sβ2γ2S mutant receptors was associated with lower efficiency (slope factor: clotiazepam: 1.7 ± 0.1).
Regression analysis enabled the estimation of BZD efficiency when variable mixtures of α1β2 and α1β2γ2S(γ1) receptors are expressed and provided new insights into the γ2S(γ1) dependency of BZD action. The method for determining the slope of the regression line also allowed the determination of the percentage of α1β2γ2S receptors. Assuming a twofold difference in the single channel conductance of α1β2 and α1β2γ2S receptors, at 70% current ratio of γ2S-containig receptors our mathematical model predicts only about 50% of γ2S subunit incorporation. Our data suggest that BZDs reduce a γ-subunit-mediated inhibition of GABA sensitivity.
Boileau AJ, Baur R, Sharkey LM, Sigel E, Czajkowski C: The relative amount of cRNA coding for γ2 subunits affects stimulation by benzodiazepines in GABAA receptors expressed in Xenopus oocytes. Neuropharmacology. 2002, 43: 695-700. 10.1016/S0028-3908(02)00036-9.
This work was supported by FWF grant 15914 (SH).
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Baburin, I., Khom, S., Timin, E. et al. Benzodiazepines modulate GABAA receptors by reducing a gamma-subunit-mediated inhibition of GABA sensitivity. BMC Pharmacol 9 (Suppl 2), A23 (2009). https://doi.org/10.1186/1471-2210-9-S2-A23