Cells of diverse origin utilize shifts in cytosolic calcium concentrations as intracellular signals to elicit physiological responses. In endothelium, inflammatory first messengers increase cytosolic calcium as a signal to disrupt cell-cell borders and produce inter-cellular gaps. Calcium influx across the plasma membrane is required to initiate barrier disruption, although the calcium entry mechanism responsible for this effect remains poorly understood. This chapter highlights recent efforts to define the molecular anatomy of the ion channel responsible for triggering endothelial cell gap formation. Resolving the identity and function of this calcium channel will pave the way for new anti-inflammatory therapeutic targets.