Ca2+-dependent exocytotic pathways in Chinese hamster ovary fibroblasts revealed by a caged-Ca2+ compound.
Ca2+-dependent exocytosis and endocytosis of Chinese hamster ovary (CHO) fibroblasts were investigated using capacitance measurement and rapid photolysis of a caged-Ca2+ compound, dimethoxynitrophenamine tetrasodium salt. CHO cells exhibited large and fast increases in membrane capacitance (1.9 +/- 1 picofarads, or 13 +/- 7% of total membrane area, mean +/- S.D., n = 37) upon Ca2+ jumps to [Ca2+]i larger than 20 microM. The fast exocytosis occurred with a delay (20-80 ms), and exhibited a rate constant that was strongly dependent on [Ca2+]i. The maximal rate constant of exocytosis was 2.8/s, and a half-maximal rate was achieved at 30 microM. The fast exocytosis was followed by rapid endocytosis in 28% of the cells. The endocytosis often began after a delay of 0.5-2 s. Ca2+ jumps also induced stepwise increases in membrane capacitance of 10-134 femtofarads in 40% of the cells, indicating fusion of large vesicles with diameters of 0.4-1.5 micron. The exocytosis of the large vesicles could selectively be induced with smaller Ca2+ jumps (6-20 microM), and occurred slowly with a rate constant of 0. 3/s. These data indicate that CHO fibroblasts possess Ca2+-dependent exocytotic mechanisms. Moreover, two parallel exocytotic pathways may exist reminiscent of those of neurons and endocrine cells. A kinetic model was constructed to account for the fast exocytosis of CHO cells.
Ninomiya
Y
Y
Department of Physiology, Faculty of Medicine, University of Tokyo, Hongo, Bunkyo-ku, Tokyo 113, Japan.
Kishimoto
T
T
Miyashita
Y
Y
Kasai
H
H
D000085:Acetates / Q000494:pharmacology; D000818:Animals; D016466:CHO Cells; D002118:Calcium / Q000378:metabolism*; D002462:Cell Membrane / Q000502:physiology; D006224:Cricetinae; D004553:Electric Conductivity; D005029:Ethylenediamines / Q000494:pharmacology; D005089:Exocytosis / Q000502:physiology*; D005347:Fibroblasts; D007700:Kinetics; D008954:Models, Biological; D010782:Photolysis
D000085:Acetates; D005029:Ethylenediamines; C057014:DM-nitrophen; D002118:Calcium