Focal adhesions composed of integrins provide an important structural basis for anchoring the endothelial lining to its surrounding matrices in the vascular wall. Complex molecular reactions occur at the endothelial cell-matrix contact sites in response to physical and chemical stress present in the circulatory system. Recent experimental evidence points to the importance of focal adhesions in the regulation of microvascular barrier function. On one hand, the adhesive interaction between integrins and their extracellular ligands is essential to the maintenance of endothelial barrier properties, and interruption of integrin-matrix binding leads to leaky microvessels. On the other hand, focal adhesion assembly and activation serve as important signalling events in modulating endothelial permeability under stimulatory conditions in the presence of angiogenic factors, inflammatory mediators, or physical forces. The focal responses show distinctive patterns with different temporal characteristics, whereas focal adhesion kinase (FAK) plays a central role in initiating and integrating various signalling pathways that ultimately affect the barrier function. The molecular basis of focal adhesion-dependent microvascular permeability is currently under investigation, and advances in the technologies of computerized quantitative microscopy and intact microvessel imaging should aid the establishment of a functional significance for focal adhesions in the physiological regulation of microvascular permeability.