Figure 2. Zn²⁺ inhibition of GABA-evoked currents from α1β3 and α1β3γ2 GABA_A receptors.

Xenopus laevis oocytes were injected with cRNA and subjected to two-electrode voltage clamp electrophysiology as described in the methods. Control currents (I_control) were evoked using a GABA concentration corresponding to ~EC₅₀ and inhibition by Zn²⁺ was evaluated by co-applications with GABA_control. (A) Representative GABA-evoked current traces from oocytes injected with the denoted cRNA mixtures. Bars above each trace indicate GABA and Zn²⁺ application periods. Dotted lines indicate the peak current amplitude by GABA_control and “/” a wash period. (B,C) Concentration-response relationships of Zn²⁺ inhibition of GABA_control-evoked currents at α1β3 or α1β3γ2 receptors stemming from injecting the indicated cRNA mixtures using free subunits (B) or a concatenated β3-α1 construct (C). Averaged Zn²⁺ inhibition values were depicted as means ± S.E.M as a function of the Zn²⁺ concentration and fitted to the Hill equation by non-linear regression. Regression results for α1 + β3 (1:1) were IC₅₀ = 0.84 (95% CI: 0.53–1.3), nH = −0.5 ± 0.32 and for β3-α1 + β3 (1:2) were IC₅₀ = 1.6 (95% CI: 1.1–2.4), nH = −0.6 ± 0.06. For the remaining cRNA mixtures, Zn²⁺ inhibition at the maximal tested concentration was too low to allow for meaningful fitting. Each data point represent experiments from n = 5–8 oocytes of ≥2 batches. (D) The depiction of α1β3 GABA_A receptor stoichiometries from Fig. 1D was modified to indicate Zn²⁺ binding in the β3(+)-β3(−) subunit interface (red/orange arrowhead).