CALCIUM-CHANNEL ACTIVITY DURING POLLEN-TUBE GROWTH AND REORIENTATION

We have shown previously that the inhibition of pollen tube growth and its subsequent reorientation in Agapanthus um bellatus are preceded b y an increase in cytosolic free calcium ([Ca2+](c)), suggesting a role for Ca2+ in signaling these processes. In this study, a novel procedu re was used to measure Ca2+ channel activity in living pollen tubes su bjected to various growth reorienting treatments (electrical fields an d ionophoretic microinjection). The method involves adding extracellul ar Mn2+ to quench the fluorescence of intracellular Indo-1 at its Ca2-insensitive wavelength (isosbestic point). The spatial and temporal k inetics of Ca2+ channel activity correlated well with measurements of [Ca2+](c) dynamics obtained by fluorescence ratio imaging of Indo-1. T ip-focused gradients in Ca2+ channel activity and [Ca2+](c) were obser ved and quantified in growing pollen tubes and in swollen pollen tubes before reoriented growth. In nongrowing pollen tubes, Ca2+ channel ac tivity was very low and [Ca2+](c) gradients were absent. Measurements of membrane potential indicated that the growth reorienting treatments induced a depolarization of the plasma membrane, suggesting that volt age-gated Ca2+ channels might be activated.