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. 2019 Apr 15;151(6):798–819. doi: 10.1085/jgp.201812133

Table 1. Regression results from fitted GABA-evoked responses at wild-type and concatenated GABAARs.

GABA
Construct Subunit Emax EC50 (µM) pEC50 n GABAmax (nA)
 α1 + β2 1:50 0.99 ± 0.01 2.6 5.58 ± 0.03 12 1,500 ± 200
 α1 + β2 50:1 0.99 ± 0.02 3.2 5.49 ± 0.03 17 750 ± 120
 α1 + β2 + γ2 5:1:5 1.00 ± 0.01 22 4.66 ± 0.03 10 3,200 ± 400
Dimeric constructs
 β2-23-α1 1.00 ± 0.02 3.0 5.53 ± 0.04 12 480 ± 100
 β2-23-α1 β2 1.00 ± 0.02 2.6 5.58 ± 0.03 10 1,900 ± 200
 β2-23-α1 γ2 1.00 ± 0.02 9.2 5.04 ± 0.04 8 8,400 ± 600
 α1-10-β2 1.00 ± 0.01 2.8 5.55 ± 0.03 11 820 ± 190
 α1-10-β2 β2 1.00 ± 0.01 1.8 5.75 ± 0.02 13 1,800 ± 200
 α1-10-β2 γ2 1.00 ± 0.02 14 4.84 ± 0.04 10 8,100 ± 600
 α1-9a-β2 1.00 ± 0.01 3.2 5.49 ± 0.03 10 190 ± 60
 α1-9a-β2 β2 1.00 ± 0.02 1.7 5.77 ± 0.04 11 1,200 ± 300
 α1-9a-β2 γ2 0.99 ± 0.01 17 4.77 ± 0.03 20 11,000 ± 1,000
 α1-6a-β2 0.99 ± 0.01 5.7 5.24 ± 0.03 11 140 ± 30
 α1-6a-β2 β2 1.00 ± 0.01 2.3 5.65 ± 0.02 11 1,300 ± 300
 α1-6a-β2 γ2 1.00 ± 0.02 41 4.38 ± 0.04 10 3,100 ± 300
 α1-3a-β2 1.00 ± 0.02 10 4.99 ± 0.04 10 89 ± 28
 α1-3a-β2 β2 1.00 ± 0.02 3.2 5.49 ± 0.03 9 630 ± 110
 α1-3a-β2 γ2 0.99 ± 0.02 34 4.46 ± 0.04 12 4,300 ± 500
 α1-0a-β2 N/A 19 34 ± 10
 α1-0a-β2 β2 1.00 ± 0.01 3.5 5.45 ± 0.02 14 630 ± 100
 α1-0a-β2 γ2 0.99 ± 0.03 130 3.88 ± 0.05 11 500 ± 140
 α1-(-1a)-β2 N/A 21 No current
 α1-(-1a)-β2 β2 0.99 ± 0.01 2.6 5.58 ± 0.03 7 400 ± 140
 α1-(-1a)-β2 γ2 0.99 ± 0.02a 140a 3.84 ± 0.04a 18 7.8 ± 1.9b
 α1-(-2a)-β2 N/A 15 No current
 α1-(-2a)-β2 β2 1.00 ± 0.01 2.6 5.59 ± 0.03 11 470 ± 90
 α1-(-2a)-β2 γ2 1.00 ± 0.01a 130a 3.90 ± 0.03a 17 8.0 ± 3.6b
 α1-(-3a)-β2 N/A 21 No current
 α1-(-3a)-β2 β2 0.96 ± 0.04 4.0 5.40 ± 0.08 8 40 ± 9
 α1-(-3a)-β2 γ2 1.00 ± 0.01a 64a 4.20 ± 0.02a 24 2.3 ± 0.9b
Tetrameric constructs
 α1-10-β2-α1-β2 N/A 13 60 ± 13
 α1-10-β2-α1-β2 γ2 0.99 ± 0.02 32 4.50 ± 0.03 17 6,500 ± 1,000
 α1-10-β2-α1-γ2 N/A 15 150 ± 25
 α1-10-β2-α1-γ2 β2 0.96 ± 0.03 34 4.47 ± 0.05 19 4,700 ± 400
Pentameric constructs
 α1-(-1a)-β2-α1-β2-γ2 0.99 ± 0.01 390 3.41 ± 0.02 19 320 ± 40
 α1-(-1a)-β2-α1-γ2-β2 1.00 ± 0.01 330 3.49 ± 0.02 16 3,500 ± 400
 γ2-15a-β2-α1-β2-α1 0.99 ± 0.03 51 4.30 ± 0.04 11 4,900 ± 600
 γ2-13a-β2-α1-β2-α1 0.99 ± 0.02 70 4.16 ± 0.03 12 5,300 ± 500
 γ2-11a-β2-α1-β2-α1 0.99 ± 0.01 110 3.96 ± 0.02 17 3,300 ± 500
 γ2-10a-β2-α1-β2-α1 0.99 ± 0.02 120 3.92 ± 0.04 10 3,100 ± 700
 γ2-9a-β2-α1-β2-α1 1.00 ± 0.01 120 3.92 ± 0.02 15 890 ± 160
Mutant constructs
 α1-9a-β2 γ2(A118R) 0.99 ± 0.02 30 4.52 ± 0.04 13 8,700 ± 1,500
 α1-0a-β2 γ2(A118R) 1.00 ± 0.01 110 3.98 ± 0.02 9 600 ± 90
 α1R94A + β2 + γ2 5:1:5 0.99 ± 0.02 1,100c 2.97 ± 0.03c 10 7,100 ± 400
 γ2-11a-β2-α1-β2-α1R94A 1.01 ± 0.01 2,600c 2.58 ± 0.02c 14 3,600 ± 300
 γ2-11a-β2-α1-β2-α1R94A α1 0.97 ± 0.02c,d 2,000c,d 2.70 ± 0.03c,d 12 1,700 ± 200

Data points were fitted to the Hill equation with the bottom set to 0 and Hill slope set to 1 by nonlinear regression. Normalized fitted maximal responses of GABA are presented as Emax ± SEM. Fitted potencies of GABA are presented as EC50 in µM and pEC50 ± SEM where p = −Log for the indicated number (n) of individual oocytes. The average maximal current obtained with GABAmax applications is presented as GABAmax ± SEM in nA for the tested oocytes. For α-10-β+γ2, a GABA concentration of 3,160 µM was used to establish GABAmax-evoked peak currents amplitudes, whereas a concentration of 316 µM was used for all remaining cRNA combinations of wild-type free subunits, binary constructs, and tetrameric constructs. For pentameric constructs and cRNA combinations with mutant subunits, the GABAmax concentration was adjusted according to the EC50 value of GABA CRRs: for CRRs with EC50 values <70 µM, a concentration of 316 µM was used; for CRRs with EC50 values between 70 and 500 µM, a concentration of 3,160 or 10,000 µM was used; and for CRRs with EC50 values >500 µM, a concentration of 10,000 µM was used. N/A, not applicable; No current, no GABAmax-evoked peak-current amplitude exceeded 5 nA in any of the tested oocytes.

a

Data were from n = 5 experiments performed on oocytes that had been incubated for 5–6 d following cRNA injection and selected based on GABAmax-evoked current amplitudes; note that data for these n = 5 oocytes are not included in the number of oocytes tested or GABAmax results.

b

Given the GABA EC50 value at this construct combination, the utilized 316-µM GABAmax concentration represents a submaximal EC70–80 concentration; an EC100 concentration would thus be expected to give values that are 20–30% larger.

c

Only 80–90% of the fitted curve is covered by data points (Fig. 9 C) and derived fitted values could therefore be seen as lower limits.

d

These values are derived from first-order fitting; however, data are statistically better represented by second-order fitting as described in the text.