Apparent change in ion selectivity caused by changes in intracellular K+ during whole-cell recording

In whole-cell recordings from HEK293 cells stably transfected with the delayed rectifier K+ channel Kv2.1, long depolarizations produce current-dependent changes in [K+](i) that mimic inactivation and changes in ion selectivity. With 10 mM K-o(+) or K-i(+), and 140-160 mM Na-i,o(+), long depolarizations shifted the reversal potential (V-R) toward E-Na. However, similar shifts in V-R were observed when Na-i,o(+), was replaced with N-methyl-o-glucamine (NMG(+))(i,o), In that condition, [K+](o) did not change significantly, but the results could be quantitatively explained by changes in [K+](i). For example, a mean outward K+ current of 1 nA for 2 s could decrease [K+](i) from 10 mM to 3 mM in a 10 pF cell. Dialysis by the recording pipette reduced but did not fully prevent changes in [K+](i). With 10 mM K-i,o(+), 150 mM Na-i(+) end 140 mM NMG(o)(+), steps to +20 mV produced a positive shift in V-R, as expected from depletion of K-i(+), but opposite to the shift expected from a decreased K+/Na+ selectivity, Long steps to V-R caused inactivation, but no change in V-R. We conclude that current-dependent changes in [K+](i) need to be carefully evaluated when studying large K+ currents in small cells.