Transient and permanent fusion of vesicles in Zea mays coleoptile protoplasts measured in the cell-attached configuration

Exocytosis in protoplasts from Zea mays L. coleoptiles was studied using pa tch-clamp techniques. Fusion of individual vesicles with the plasma membran e was monitored as a step increase of the membrane capacitance (C-m). Vesic le fusion was observed as (i) An irreversible step increase in C-m. (ii) Oc casionally, irreversible C-m steps were preceded by transient changes in C- m, suggesting that the electrical connection between the vesicle with the p lasma membrane opens and closes reversibly before full connection is achiev ed. (iii) Most frequently, however, stepwise transient changes in C-m did n ot lead to an irreversible C-m step. Within one patch of membrane capacitan ce steps due to transient and irreversible fusions were of similar amplitud e. This suggests that the exocytosis events do not result from the fusion o f vesicles with different sizes but are due to kinetically different states in a fusion process of the same vesicle type. The dwell time histogram of the transient fusion events peaked at about 100 msec. Fusion can be describ ed with a circular three-state model for the fusion process of two fused st ates and one nonfused state. It predicts that energy input is required to d rive the system into a prevailing direction.