Peak current recovery as a function of intracellular Na⁺ concentration. (A) Top: Schematic rendering of the recording conditions indicating the concentrations of intracellular and extracellular substrate and cosubstrate ions and reaction scheme of a Na⁺-coupled transporter. Bottom: Original traces of representative experiments using 10, 70, or 140 mM intracellular Na⁺. (B–F) Time-dependent recovery of 5-HT–induced (10 µM) peak current amplitude after application of 30 µM PAL-287 (B; n = 4–6), 30 µM PAL-1046 (C; n = 5–7), 30 µM PAL-1045 (D; n = 5), 10 µM 5-HT (E; n = 5), and 30 µM p-chloroamphetamine (F; n = 5). The red curves in B–D are the fits of data displayed in Fig. 1. Data are means ± SD. The data points were fitted by monoexponential functions in the case of 5-HT and the PAL substrates. A biexponential function was used to fit the recovery data of p-chloroamphetamine at 70 and 140 mM Na⁺_i. For the Na⁺_i-free condition, a monoexponential fit was used. (G) Relaxation rates were obtained by the fits from B–F. The rates shown for p-chloroamphetamine are the k_fast values obtained from the biexponential fit. The k_slow values are (±SEM) 70 mM Na⁺_i: 0.50 ± 0.11 s⁻¹; 140 mM Na⁺_i: 0.49 ± 0.15 s⁻¹. Data are means ± 95% confidence intervals of the fits. The points marked in blue were obtained from data published in a previous study (Bhat et al., 2017). The gray points displayed for 140 mM Na⁺_i (which equals 140 mM Na⁺_e) were taken from Fig. 1 for comparison. Statistical significance was determined by one-way ANOVA followed by multiple comparisons using the Fisher’s LSD method or by an unpaired t test in the case of 5-HT and p-chloroamphetamine. ****, P < 0.0001; n.s., not significant.