Cytosolic Ca2+ overload and macromolecule permeability of endothelial monolayers.
It was investigated how cytosolic Ca2+ overload affects the cytoskeletal structure and macromolecule permeability (for albumin) of monolayers of endothelial cells (from porcine aorta). States of cytosolic Ca2+ overload were produced either 1. by metabolic inhibition (5 mM KCN plus 5 mM 2-deoxyglucose) or 2. by increasing membrane permeability with the use of a Ca2+ ionophore (10 microM A 23187). The effects of cytosolic Ca2+ overload on the structure of F-actin filaments and monolayer permeability were monitored. ATP stores were rapidly degraded (> 90% in 15 minutes) in the presence of metabolic inhibitors, but only partially reduced in the presence of A 23187 (30%) in two hours). Concomitantly with ATP loss, cytosolic Ca2+ levels were increased in metabolically inhibited cells. Two-hour exposure to the Ca2+ ionophore A 23187 mimicked the effect of two-hour metabolic inhibition on F-actin filaments and monolayer permeability, in spite of the divergence in energy metabolism. Disintegration of F-actin filaments in presence of metabolic blockers or ionophore was accompanied by appearance of F-actin clumps in the cells, but total contents of F-actin remained unaltered. Within three hours after removal of these agents, a normal F-actin structure and normal macromolecule permeability were re-established in the monolayers. The results show that cytosolic Ca2+ overload causes disintegration of F-actin filaments and a subsequent increase in macromolecule permeability. These changes are readily reversible as long as the dis-integration is based on fragmentation and not depolymerization of F-actin filaments.
Piper
H M
HM
Physiologisches Institut I, Universität Düsseldorf.
Noll
T
T
Muhs
A
A
Besselmann
M
M
Kuhne
W
W
Watanabe
H
H
D000199:Actins / Q000502:physiology; D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D001011:Aorta; D002118:Calcium / Q000502:physiology*; D015220:Calcium Channels / Q000502:physiology*; D002463:Cell Membrane Permeability / Q000502:physiology*; D046508:Culture Techniques; D003599:Cytoskeleton / Q000502:physiology*; D003600:Cytosol / Q000502:physiology*; D004730:Endothelium, Vascular / Q000502:physiology*; D013552:Swine
D000199:Actins; D015220:Calcium Channels; D000255:Adenosine Triphosphate; D002118:Calcium