Fig 4. The α₁ subunit is critical for neurosteroid modulation of α₁β₃ GABA_A receptors.

(A) Photolabeling with 3 μM KK123, KK200, and KK202 (black) is prevented by labeling in the presence of 30 μM allopregnanolone (red). Each circle represents a replicate experiment, and the bars represent mean ± SD, n = 3. (B) Sample current traces showing modulation of WT α₁β₃, α₁^V227Wβ₃ and α₁^N408A/Y411Fβ₃ GABA_A receptors by allopregnanolone or propofol. (C1) Allopregnanolone potentiation of GABA-elicited currents is reduced in α₁^V227Wβ₃ (orange) and α₁^N408A/Y411Fβ₃ (blue) receptors compared to WT (black). Potentiation is given as the ratio of peak responses to GABA + steroid to GABA alone. Potentiation value of 1 indicates no potentiation by neurosteroids. (C2) Direct activation of α₁β₃ GABA_A receptors by allopregnanolone is reduced in α₁^V227Wβ₃ receptors (orange). Direct activation is given in percentage of the peak response to steroid to peak response to saturating GABA + 100 μM propofol. (C3) Potentiation of GABA-elicited currents by propofol, or (C4) direct activation of the receptor in the presence of propofol is not affected by the α₁^V227W or α₁^N408A/Y411F mutations. Each data point represents a replicate experiment. Bars show the mean ± SD (n = 6–8). Data were compared by a one-way analysis of variance followed by a Dunnett’s multiple comparison tests of the means. ***p < 0.001; **p < 0.01; *p < 0.05. The numerical data are included in S4 Data. AlloP, allopregnanolone; GABA_A, gamma amino-butyric acid Type A; WT, wild-type.