Figure 6.

Extracellular ATP and ADP potentiates function of recombinant GABA_ARs. Whole-cell voltage-clamp recordings were performed in HEK293 cells transiently expressing rat recombinant α1β2γ2 GABA_ARs at a holding membrane potential of -60 mV. GABA currents were induced by fast perfusion of GABA at various concentrations. A, Representative current traces showing the potentiation of GABA (3 μM) currents by co-perfusion of ATP (0.5 mM). The increased current was blocked in the presence of bicuculline (10 μM) and returned to the control level after ATP washout. B, The dose-response curve showing the dose-dependent potentiation of the GABA (3 μM) currents by various concentration of ATP. All values were normalized to the maximal GABA currents (I_max) induced at 3 mM of ATP. Each data point is the mean ± SEM of normalized GABA currents at the indicated ATP concentrations (n = 6). The solid line is the best fit of the data to the Hill equation. The EC50 and H were 390 ± 48 μM and 1.66 ± 0.1, respectively. C, ATP potentiation of GABA_AR activity is GABA dose-dependent. GABA dose-response curves were constructed in the absence (Control) and presence (ATP) of ATP (0.5 mM) from five cells. ATP shifted the curve to the left and reduced EC50 from 11.6 ± 3.3 μM to 5.8 ± 2.1 (P < 0.01) without change in H (control: 1.57 ± 0.1; ATP: 1.62 ± 0.1; P > 0.05). All values were normalized to the I_max induced by GABA (300 μM). D, ADP mimics ATP, potentiating the activity of recombinant GABA_ARs. Representative current traces showed that application of ADP (0.5 mM) reversibly increased currents induced by GABA (3 μM). On average, it increased the currents by 147.5 ± 15.9% (P < 0.01; n = 5).