Two mechanisms for inward rectification of current flow through the purinoceptor P2X2 class of ATP-gated channels

1. The ATP receptor subunit P2X2 was expressed in Xenopus oocytes and human embryonic kidney (HEK) 293 cells. ATP-activated currents were studied with two-electrode voltage clamp recordings from oocytes, whole-cell recordings from HEK 293 cells, and outside-out patch clamp recordings from both cell types. The steady-state current-voltage (I-V) relation showed profound inward rectification in all recording configurations. 2. Recordings from outside-out patches demonstrated that inward rectification does not require intracellular Mg2+ or polyamines, and that inward rectification was present when the same solution was used on both sides of the patch. 3. Voltage jump experiments were performed to evaluate the voltage dependence of channel gating. After fast voltage jumps, instantaneous current jumps were followed by substantial relaxations to the steady state. The time course of the current relaxations could be fitted by single exponential functions. The instantaneous I-V relation was less inwardly rectifying than the steady-state I-V relation; however, it was not linear. 4. Single channel recordings indicated that the single channel conductance became smaller when the membrane potential became more positive. This decrease could quantitatively account for inward rectification of the instantaneous I-V relation. 5. We conclude that inward rectification of P2X2 is due to two mechanisms: voltage-dependent gating and voltage dependence of the single channel conductance.