Our major theme is that the layered structure of the endothelial barrier requires continuous activation of signalling pathways regulated by sphingosine-1-phosphate (S1P) and intracellular cAMP. These pathways modulate the adherens junction, continuity of tight junction strands, and the balance of synthesis and degradation of glycocalyx components. We evaluate recent evidence that baseline permeability is maintained by constant activity of mechanisms involving the small GTPases Rap1 and Rac1. In the basal state, the barrier is compromised when activities of the small GTPases are reduced by low S1P supply or delivery. With inflammatory stimulus, increased permeability can be understood in part as the action of signalling to reduce Rap1 and Rac1 activation. With the hypothesis that microvessel permeability and selectivity under both normal and inflammatory conditions are regulated by mechanisms that are continuously active, it follows that when S1P or intracellular cAMP are elevated at the time of inflammatory stimulus, they can buffer changes induced by inflammatory agents and maintain normal barrier stability. When endothelium is exposed to inflammatory conditions and subsequently exposed to elevated S1P or intracellular cAMP, the same processes restore the functional barrier by first re-establishing the adherens junction, then modulating tight junctions and glycocalyx. In more extreme inflammatory conditions, loss of the inhibitory actions of Rac1-dependent mechanisms may promote expression of more inflammatory endothelial phenotypes by contributing to the up-regulation of RhoA-dependent contractile mechanisms and the sustained loss of surface glycocalyx allowing access of inflammatory cells to the endothelium.