STUDIES ON FENESTRAL CONTRACTION IN RAT-LIVER ENDOTHELIAL-CELLS IN CULTURE

Liver endothelial cells possess fenestrae, which are pores supported b y a cytoskeleton ring composed of actin and myosin. Fenestrae are dyna mic structures that can contract or dilate, although the mechanism for this phenomenon remains to be elucidated. Staining of actin and/or of myosin permitted measurement of fenestral diameter and area in cultur ed rat liver endothelial cells using digitized video-intensified fluor escence microscopy with image analysis, Within 1 minute of incubation with 0.1 mu mol/L serotonin, fenestral diameter and area decreased by 24 +/- 5% and 56 +/- 7%, respectively. Contraction of fenestrae by ser otonin was inhibited by chelation of extracellular Ca2+ with EGTA and by addition of Ca2+ channel blockers, such as dilthiazem and verapamil . The response of fenestrae to serotonin was mimicked by addition of a Ca2+ ionophore, A23187. Serotonin inhibited cAMP production, had no e ffect on inositol phosphate production, and activated phospholipase A( 2), causing release of arachidonic acid. These results suggest that co ntraction of fenestrae is associated with Ca2+ influx. In response to 0.1 mu mol/L serotonin, intracellular Ca2+ levels increased within 3 t o 5 seconds from 150 nmol/L to > 400 nmol/L followed by rapid phosphor ylation of the 20-kd subunit of myosin light chain; both events depend ed on extracellular Ca2+.