M. Oike et al., CYTOSKELETAL MODULATION OF THE RESPONSE TO MECHANICAL STIMULATION IN HUMAN VASCULAR ENDOTHELIAL-CELLS, Pflugers Archiv, 428(5-6), 1994, pp. 569-576
Possible interactions of cytoskeletal elements with mechanically induc
ed membrane currents and Ca2+ signals were studied in human endothelia
l cells by using a combined patch-clamp and Fura II technique. for mec
hanical stimulation, cells were exposed to hypotonic solution (HTS). T
he concomitant cell swelling activates as Cl- current releases Ca2+ fr
om intracellular stores and activates Ca2+ influx. To interfere with t
he cytoskeleton, cells were loaded either with the F-actin-stabilizing
agent phalloidin (10 mu mol/l), or the F-actin-depolymerizing substan
ce cytochalasin B (50 mu mol/l). These were administered either in the
bath or the pipette solutions. The tubulin structure of the endotheli
al cells was modulated by taxol (50 mu mol/l), which supports polymeri
zation of tubulin, or by the depolymerizing agent colcemid (10 mu mol/
l) both applied to the bath. Immunofluorescence experiments show that
under the chosen experimental conditions the cytoskeletal modifiers em
ployed disintegrate the F-actin and microtubuli cytoskeleton. Neither
of these cytoskeletal modifiers influenced the HTS-induced Cl- current
. Ca2+ release was not affected by cytochalasin B, taxol or colcemid,
but was suppressed if the cells were loaded with phalloidin. Depletion
of intracellular Ca2+ stores by thapsigargin renders intracellular [C
a2+] sensitive to the extracellular [Ca2+], which is indicative of a C
a2+ entry pathway activated by store depletion. Neither cytochalasin B
nor phalloidin affected this Ca2+ entry. We conclude that F-actin tur
nover or depolymerization is necessary for Ca2+ release by mechanical
activation. The tubulin network is not involved. The Ca2+ release-acti
vated Ca2+ entry is not modulated by the F-actin cytoskeleton.