FIGURE 10.
Effects of CuPh on the transient currents and GABA-induced steady state currents by WT and S359C/K448A. The membrane voltage of oocytes expressing WT and S359C/K448A was stepped from a holding potential of −25 mV to voltages between −140 and +60 mV in 25-mV increments. Each potential was held clamped for 500 ms, followed by 500 ms of a potential clamped at −25 mV. A, charge movements (open bars, Qmax) and GABA-induced currents (gray bars) for WT and S359C/K448A following treatment with 20 μm CuPh were normalized to those before the exposure to the oxidant, QCuPh/QControl and ICuPh/IControl, respectively. The data are given as mean ± S.E. (error bars) of at least three oocytes. *, p < 0.05. B, voltage dependence of the GABA-induced currents by WT and S359C/K448A before and after application of 20 μm CuPh. For WT and S359C/K448A, the GABA-induced currents at each potential from 420 to 480 ms were averaged and normalized to the GABA-induced current at −140 mV. These currents were then plotted against the corresponding potential (mV). The data are the means ± S.E. (error bars) of at least three repeats. Wherever error bars are not visible, the error was smaller than the size of the symbols. The currents at −140 mV induced by 1 mm GABA ranged from −177 to −331 nA in WT and from 147 to −186 nA in S359C/K448A. C, the charge movements of oocytes expressing WT and S359C/K448A in 100 mm sodium, before and after treatment with 20 μm CuPh, were plotted as a function of the voltage. Charge movements were normalized to Qmax and were fit, using the Boltzmann distribution non-linear curve fit function in Origin version 6.1 (OriginLab Corp.). Wherever error bars are not visible, the error was smaller than the size of the symbols. The Qmax values before and after application of CuPh were 16 ± 2.5 and 16.1 ± 2.6 nanocoulombs for WT and 13.5 ± 0.6 and 11.5 ± 0.7 nanocoulombs for S359C/K448A, respectively. Data points are averaged from at least three oocytes for each transporter.