In vitro Arabidopsis pollen germination and characterization of the inwardpotassium 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-contai ning media, in vitro methods for Arabidopsis pollen germination have been s uccessfully established for the first time. The pollen germination percenta ge was nearly 75% and the average pollen tube length reached 135 pm after a 6 h incubation. A decrease in external K+ concentration from 1 mM to 35 mu M resulted in 30% inhibition of pollen germination and 40% inhibition of po llen tube growth. An increase in external K+ concentration from I 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, regul ation of the inward K+ channels in the pollen plasma membrane was investiga ted by conducting patch-clamp whole-cell recording with pollen protoplasts. K+ currents were first identified in Arabidopsis pollen protoplasts. The i nward 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 o n 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 ext ernal pH significantly affected the magnitude, conductance, voltage-indepen dent 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 discuss ed.