Abstract
Objective: To determine whether fluid flow influences the action of soluble vasoactive agonists on vascular endothelium.
Methods: Confluent monolayers of bovine aortic endothelial cells (BAEC) were cultured on glass coverslips, prelabeled with the Ca2+-sensitive dye fura-2, and placed in a parallel-plate flow chamber designed to generate defined laminar fluid flow. Cytosolic free Ca2+ concentration ([Ca2+];) in individual BAEC was monitored during perfusion with medium containing adenine nucleotide under defined flow conditions.
Results: Continuous perfusion with ATP (0.3–3.0 μM) or ADP (0.1–1.0 μM) evoked repetitive oscillations in [Ca2+]; in individual BAEC. The frequency of the [Ca2+]; oscillations was dependent on both nucleotide concentration and levels of applied shear stress; at constant bulk concentration of nucleotide, the frequency increased with shear stress. Stopping flow in the continuous presence of agonists immediately extinguished the oscillatory response. Elimination of extracellular Ca2+ did not inhibit the [Ca2+]; oscillations. In the presence of nonhydrolyzable nucleotide analog, ATPγS or ADPβS, application of flow resulted in similar shear-dependent [Ca2+]; oscillations, suggesting that flow modulation of the [Ca2+]; response was not simply due to depletion of ATP or ADP in the vicinity of BAEC monolayers as a result of hydrolysis of nucleotides by ectonucleotidases.
Conclusions: These findings suggest that local hemodynamic conditions may modulate the action of vasoactive agents on the vascular endothelium in vivo.