Characterization of the electrogenic Na +–K + pump in horizontal cells isolated from the carp retina

Abstract

The electrogenic Na +–K + pump current in horizontal cells acutely dissociated from the carp retina was investigated using a nystatin-perforated patch recording configuration under voltage-clamp conditions. In the presence of suitable blockers for known voltage-dependent Na +, K + and Ca 2+ conductances, the pump current was activated in a concentration-dependent manner by adding K + ions to external solution. The EC 50 value and Hill coefficient for the external K + concentration were 0.66 mM and 1.39, respectively. The pump current did not show any significant voltage dependency at the physiological potential range between −90 and 20 mV either with or without external Na + ions. In the presence of 120 mM external Na + concentration, the addition of 3 mM K + to the external solution induced a steady outward pump current even when the patch-pipette (internal) solution did not contain Na +. A large outward shift of the holding current was observed by removing external Na +. The result thus suggests that continuous Na + influxes exist across the plasma membrane in the presence of external Na +. When Na + was removed from both external and internal solutions, a transient outward pump current was observed by adding K + to the external solution, thus indicating that the transient pump current was activated by the residual intracellular Na + ions. The pump current was suppressed by ouabain in a concentration-dependent manner, and the ouabain-sensitive inhibition curve was fitted by two components. The IC 50 values of high- and low-sensitive pump currents for ouabain were 20 nM and 10.4 μM, respectively, indicating the existence of at least two isoforms of the pump in the horizontal cells.