Adenosine 5'-triphosphate (ATP) affects multiple intra- and extracellular processes, including vascular tone and immune responses. Microvascular endothelial cells (EC) play a central role in inflammation by recruitment of inflammatory cells from blood to tissues. We hypothesized that ATP (secreted by neurons and/or released after perturbation of cutaneous cells) may influence secretion of inflammatory messengers by dermal microvascular EC through actions on purinergic P2 receptors. Addition of the hydrolysis-resistant ATP analogue, adenosine 5'-O-(3-thiotriphosphate) (ATPgammaS), to subconfluent cultures of the human microvascular endothelial cell-1 (HMEC-1) cell line led to a dose- and time-dependent increase in release of IL-6, IL-8, monocyte chemoattractant protein-1, and growth-regulated oncogene alpha. Both ATPgammaS-induced release and basal production of these proteins were significantly inhibited by the purinergic antagonists pyridoxal-5'-phosphate-6-azophenyl-2',5'-disulfonic acid (PPADS), pyridoxal-5'-phosphate-6(2'-naphthylazo-6-nitro-4',8'-disulfonate), and suramin. ATPgammaS increased expression of intercellular adhesion molecule-1 (ICAM-1), whereas suramin and PPADS decreased both ATPgammaS-induced and basal ICAM-1 expression. Using PCR, we found that HMEC-1 strongly express mRNA for the P2X(4), P2X(5), P2X(7), P2Y(2), and P2Y(11) receptors and weakly express mRNA for P2X(1) and P2X(3) receptors. Purinergic nucleotides may mediate acute inflammation in the skin and thus contribute to physiological and pathophysiological inflammation. For example, ATP may contribute to both the vasodilation and the inflammation associated with rosacea.