Kir2.3 isoform confers pH sensitivity to heteromeric Kir2.1/Kir2.3 channels in HEK293 cells

Background: Data on pH regulation of the cardiac potassium current I_K1 suggest species-dependent differences in the molecular composition of the underlying Kir2 channel proteins. Objective: The purpose of this study was to test the hypothesis that the presence of the Kir2.3 isoform in heterotetrameric channels modifies channel sensitivity to pH. Methods: Voltage clamp was performed on HEK293 cells stably expressing guinea pig Kir2.1 and/or Kir2.3 isoforms and on sheep cardiac ventricular myocytes at varying extracellular pH (pH_o) and in the presence of CO₂ to determine the sensitivity of macroscopic currents to pH. Single-channel activity was recorded from the HEK293 stables to determine the mechanisms of the changes in whole cell current. Results: Biophysical characteristics of whole-cell and single-channel currents in Kir2.1/Kir2.3 double stables displayed properties attributable to isoform heteromerization. Whole-cell Kir2.1/Kir2.3 currents rectified in a manner reminiscent of Kir2.1 but were significantly inhibited by extracellular acidification in the physiologic range (pK_a ∼ 7.4). Whole-cell currents were more sensitive to a combined extracellular and intracellular acidification produced by CO₂. At pH_o = 6.0, unitary conductances of heteromeric channels were reduced. Ovine cardiac ventricular cell I_K1 was pH_o and CO₂ sensitive, consistent with the expression of Kir2.1 and Kir2.3 in this species. Conclusion: Kir2.1 and Kir2.3 isoforms form heteromeric channels in HEK293. The presence of Kir2.3 subunit(s) in heteromeric channels confers pH sensitivity to the channels. The single and double stable cells presented in this study are useful models for studying physiologic regulation of heteromeric Kir2 channels in mammalian cells.