Cerebral endothelial cells interconnected by tight and adherens junctions constitute the structural basis of the blood-brain barrier. Extracellular calcium ions have been reported to play an important role in the formation and maintenance of the junctional complex. However, little is known about the action of calcium depletion on the structural characteristics of cerebral endothelial cells. Using atomic force microscopy we analyzed the effect of calcium depletion and readdition on the shape and size of living brain endothelial cells. It was found that the removal of extracellular calcium from confluent cell cultures induced the dissociation of the cells from each other accompanied by an increase in their height. After readdition of calcium a gradual recovery was observed until total confluency was regained. We have also demonstrated that Rho-kinase plays an important role in the calcium-depletion-induced disassembly of endothelial tight and adherens junctions. The Rho-kinase inhibitor Y27632 could prevent the morphological changes induced by a lack of calcium as well. Our results suggest that calcium depletion induces Rho-kinase-dependent cytoskeletal changes that may be partly responsible for the disassembly of the junctional complex.

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