Effects of intra- and extracellular acidifications on single channel Kir2.3 currents

1. The inward rectifier K+ channel Kir2.3 is inhibited by hypercapnia, and this inhibition may be mediated by decreases in intra- and extracellular pH. To understand whether Kir2.3 has two distinct pH sensors and whether cytosol-soluble factors are involved in the modulation of this channel during intracellular acidification, single channel currents were studied by expressing Kir2.3 in Xenopus oocytes. 2. In excised inside-out patches, Kir2.3 currents had a high baseline channel open-state probability (P-o, at pH 7.4) with a strong inward rectification. Single channel conductance at hyperpolarizing membrane potential was about 17 pS with 150 mar K+ applied to both sides of the membrane. The channel showed a substate conductance of about 8 pS. 3. Reduction of intracellular pH (pH(i)) produced a fast and reversible inhibition of single channel Kir2.3 currents in inside-out patches. The extent of this inhibition is concentration dependent. A clear reduction in Kir2.3 currents was seen at pH(i) 7.0, and channel activity was completely suppressed at pH(i) 6.2 with mid-point inhibition (pK) at pK 6.77. 4. The effect of low pH(i) on Kir2.3 currents was clue to a strong: inhibition of P-o and a moderate suppression of single channel conductance. The pK values for these single channel properties were pH 6.78 and 6.67, respectively. 5. The decrease in P-o with low pH(i) resulted from an increase in the channel mean closed time without significant changes in the mean open time. Substate conductance was not seen during low pH(i). 6. Decrease in extracellular pH (pH(o)) also caused inhibition of single channel activity of Kir2.3 currents in excised outside-out patches. This effect, however, was clearly different from that of pH(i): the pK (pH 6.70) was about 0.1 pH units lower; more than 50% channel activity was retained at pH(o) 5.8; and low pH(o) affected mainly single channel conductance. 7. These results therefore indicate that (1) there are two distinct pH sensors in Kir2.3, (2) different mechanisms are involved in the modulation of Kir2.3 through these two pH sensors, and (3) cytosol-soluble factors do not appear to be engaged in this modulation.