Endothelial cells, which line the vasculature, respond to specific agonists such as adenosine triphosphate (ATP) by elevating cytosolic calcium levels and increasing production of the vasoactive compounds, prostacyclin and endothelial derived relaxing factor (EDRF). Endothelial cells express ecto-enzymes which metabolize ATP. If the activity of these enzymes is sufficiently high, then the concentration of ATP near the endothelial cell surface can be substantially lower than the bulk concentration. The ATP concentration is determined by a balance between the convection of fresh ATP from upstream and the degradation of ATP by the endothelial cells. In this report, we present a parallel plate flow system for measurement of cytosolic calcium levels ([Ca2+]i) of individual bovine aortic endothelial cells with the calcium sensitive fluorescent dye, fura-2. The cells respond to increases in the flow rate by increasing [Ca2+]i if there is ATP present in the perfusing buffer, but not in the absence of ATP. The amount of agonist in the perfusing fluid near the endothelial cell surface is estimated by solving the governing differential equation for the concentration profile of ATP in the parallel plate flow geometry. The solution indicates that one mechanism endothelial cells may use to detect changes in the flow rate is to respond to the change in the local concentration of agonist.