In vitro Arabidopsis pollen germination and characterization of the inward potassium currents in Arabidopsis pollen grain protoplasts

The focus of this study is to investigate the regulatory role of K+ influx in Arabidopsis pollen germination and pollen tube growth. Using agar‐containing media, in vitro methods for Arabidopsis pollen germination have been successfully established for the first time. The pollen germination percentage was nearly 75% and the average pollen tube length reached 135 μm after a 6 h incubation. A decrease in external K+ concentration from 1 mM to 35 μM resulted in 30% inhibition of pollen germination and 40% inhibition of pollen tube growth. An increase in external K+ concentration from 1 mM to 30 mM stimulated pollen tube growth but inhibited pollen germination. To study how K+ influx is associated with pollen germination and tube growth, regulation of the inward K+ channels in the pollen plasma membrane was investigated by conducting patch‐clamp whole‐cell recording with pollen protoplasts. K+ currents were first identified in Arabidopsis pollen protoplasts. The inward K+ currents were insensitive to changes in cytoplasmic Ca2+ but were inhibited by a high concentration of external Ca2+. A decrease of external Ca2+ concentration from 10 mM (control) to 1 mM had no significant effect on the inward K+ currents, while an increase of external Ca2+ concentration from 10 mM to 50 mM inhibited the inward K+ currents by 46%. Changes in external pH significantly affected the magnitude, conductance, voltage‐independent maximal conductance, and activation kinetics of the inward K+ currents. The physiological importance of potassium influx mediated by the inward K+‐channels during Arabidopsis pollen germination and tube growth is discussed.